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Methods in
Molecular Biology 2693

Stuart K. Calderwood
Thomas L. Prince Editors

Chaperones
Methods and Protocols
Second Edition
METHODS IN MOLECULAR BIOLOGY

Series Editor
John M. Walker
School of Life and Medical Sciences
University of Hertfordshire
Hatfield, Hertfordshire, UK

For further volumes:

http://www.springer.com/series/7651
For over 35 years, biological scientists have come to rely on the research protocols and
methodologies in the critically acclaimed Methods in Molecular Biology series. The series was
the first to introduce the step-by-step protocols approach that has become the standard in all
biomedical protocol publishing. Each protocol is provided in readily-reproducible step-by-
step fashion, opening with an introductory overview, a list of the materials and reagents
needed to complete the experiment, and followed by a detailed procedure that is supported
with a helpful notes section offering tips and tricks of the trade as well as troubleshooting
advice. These hallmark features were introduced by series editor Dr. John Walker and
constitute the key ingredient in each and every volume of the Methods in Molecular Biology
series. Tested and trusted, comprehensive and reliable, all protocols from the series are
indexed in PubMed.
 Chaperones

Methods and Protocols

Second Edition

Edited by

Stuart K. Calderwood
Harvard Medical School, Beth Israel Deaconess Medical Center, Boston, MA, USA

Thomas L. Prince
Danville, PA, USA
Editors
Stuart K. Calderwood Thomas L. Prince
Harvard Medical School Danville, PA, USA
Beth Israel Deaconess Medical Center
Boston, MA, USA

ISSN 1064-3745 ISSN 1940-6029 (electronic)

Methods in Molecular Biology
ISBN 978-1-0716-3341-0 ISBN 978-1-0716-3342-7 (eBook)
https://doi.org/10.1007/978-1-0716-3342-7

© The Editor(s) (if applicable) and The Author(s), under exclusive license to Springer Science+Business Media, LLC, part
of Springer Nature 2023
This work is subject to copyright. All rights are solely and exclusively licensed by the Publisher, whether the whole or part
of the material is concerned, specifically the rights of translation, reprinting, reuse of illustrations, recitation,
broadcasting, reproduction on microfilms or in any other physical way, and transmission or information storage and
retrieval, electronic adaptation, computer software, or by similar or dissimilar methodology now known or hereafter
developed.
The use of general descriptive names, registered names, trademarks, service marks, etc. in this publication does not imply,
even in the absence of a specific statement, that such names are exempt from the relevant protective laws and regulations
and therefore free for general use.
The publisher, the authors, and the editors are safe to assume that the advice and information in this book are believed to
be true and accurate at the date of publication. Neither the publisher nor the authors or the editors give a warranty,
expressed or implied, with respect to the material contained herein or for any errors or omissions that may have been
made. The publisher remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Cover Caption: The predicted protein structure of HSF1, a key component of the heat shock response and guardian of
the proteome, is shown on the cover. Model structure generated by AlphaFold (Jumper, J. et al. Nature, 2021; Varadi,
M. et al. Nucleic Acid Research, 2021). HSF1 color scheme based on studies by Kijima, T. et al. Scientific Reports, 2018.

This Humana imprint is published by the registered company Springer Science+Business Media, LLC, part of Springer
Nature.
The registered company address is: 1 New York Plaza, New York, NY 10004, U.S.A.
Dedication

(SKC) This volume is dedicated to the memory of my sister Carole who passed away in
2021. I would particularly like to thank my wife Laura for her patience and support in the
long hours science demands.

v
Preface

Life is a dynamic and complex interplay of energy and mass within space and time. Biological
life depends on proteins to facilitate the processes that allow communities of cells to
cooperate and compete to propagate their genetic information. Proteins are linear polymers
that fold into intricate three-dimensional structures that provide the essential functions of
life. Here nature demonstrates the common notion that structure determines function.
Most proteins are able to spontaneously fold into their proper structure given adequate
space. However, in the crowded confines of the cell, many proteins require the aid of a
special cadre of proteins known as molecular chaperones. These chaperone proteins help
other client proteins avoid misfolding, aggregation, and loss of function. Furthermore,
molecular chaperones maintain cellular proteostasis by facilitating protein transport and
degradation. The cellular stress response is the biological process for enabling the explosive
expression of molecular chaperones in response to proteostatic perturbations. And since
heat is one of the oldest and most studied forms of proteotoxic stress, many molecular
chaperones are referred to as heat shock proteins (HSPs). Expression of HSP genes is
initiated by the activation of heat shock factors (HSFs) that bind to the DNA promoters
of chaperones and other stress-related genes to initiate or extend transcription. HSPs once
expressed may be post-translationally modified as they reestablish proteostasis while also
contributing to the modulation of immunity, extracellular communication, and cell growth.
Life is shaped by molecular chaperones and the stress response. Almost all aspects of health
and disease are affected by the ability to maintain proteostasis. Unchecked chaperone and
HSF activity enables malignant cell growth that gives way to cancer. In contrast, reduced
chaperone activity allows for accumulated protein aggregation that leads to neurodegenera-
tion. All while, the stress response influences inflammation and immunity.
Our pursuit to better understand molecular chaperones and the stress response along
with the desire to pass this knowledge on to future generations of scientist is represented by
this book. In this volume, we have complied 23 chapters of methods and topics focused on
investigating the role of chaperones. Some chapters are succinct; some chapters are exten-
sive; all address important topics within the field. We appreciate the expertise, time, and
effort the authors have provided.
We commence the volume analyzing the initiation and regulation of the stress response.
In Chap. 1, Ackerman et al. describe the use of a plasmid-based reporter for monitoring the
heat shock response. In Chaps. 2 and 3, Heyoun Bunch analyzes the stress response at the
level of transcription and RNA elongation. Holton et al. in Chap. 4 describes their workflow
for studying RNA-seq data relative to the stress response. Ritwick Sawarkar in Chap. 5
explores the role of Hsp90 in gene expression through chromosome-immunoprecipitation.
Yuka Okusha next details the use of microRNA to regulate HSP levels.
Chapters 7 through 14 revolve around major features of chaperone function and
biology: post-translational modifications and protein-protein interactions. Jin et al. explain
the effects of key HSF1 phosphorylation sites on transcriptional activity. In Chap. 8,
Chakraborty et al. describe a fluorescent complementation assay to analyze Hsp90 and
client protein interactions. Edkins and Blatch further elucidate and compare complementa-
tion assays for protein-protein interactions studies. Tonui et al. in Chap. 10 explain their
protein aggregation assay for measuring chaperone holdase activity. Sager et al. explore the

vii
viii Preface

detection of Hsp90 post-translational modifications. In Chap. 12, Ono and Eguchi discuss
the challenges of addressing HSP depletion and isoform compensation. Baldan et al. in
Chap. 13 describe their proximity ligation assay to analyze HSP protein-protein interaction
dynamics. The study of the epichaperone complexes through the use of native polyacryl-
amide gel electrophoresis is described by Roychowdhury et al in Chap. 14.
Chapter 15 through 21 focus on the emerging role of extracellular HSPs. Borges et al.
provide an update on molecular chaperone and extracellular receptor interactions. In
Chap. 15, Weng et al. expound the use of Hsp70 in antigen presentation and possible
vaccine generation. Votra et al. assess the effect of Hsp90 activity on extracellular proteases.
Ono and Eguchi follow up with proteomic profiling of chaperone-rich extracellular vesicles
with mass spectrometry. In Chap. 19, Chang et al. provide a protocol for analyzing
extracellular Hsp90 levels. Bavisotto et al. in Chap. 20 characterize the role of extracellular
Hsp60 in health and disease.
The final Chaps. 22, 23, and 24 explore the use chaperones as biomarkers. Kawai et al
develop a method for profiling HSP isoforms in cancer specimen. This is followed by a
chapter on chaperone focused cancer database analysis for prognostic biomarkers. Finally,
Dezfouli et al. utilize flow cytometry and enzyme-linked immunoassays to profile Hsp70
localization in cancer.

Boston, MA, USA Stuart K. Calderwood

Danville, PA, USA Thomas L. Prince
Acknowledgments

This book is dedicated to the next generation of scientists including my (T.L.P.) boys Jessup
and Lincoln along with my wife, Bourdana, and to the memory of my mother Suzanne
Prince. I would also like to acknowledge the Thomas Beaver Free Library in Danville, PA,
for providing me a quiet place to get work done.

ix
Contents

Dedication . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . v
Preface . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . vii
Acknowledgments. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . ix
Contributors. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . xiii

1 Monitoring of the Heat Shock Response with

a Real-Time Luciferase Reporter . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1
Andrew Ackerman, Toshiki Kijima, Takanori Eguchi,
and Thomas L. Prince
2 Studying RNA Polymerase II Promoter-Proximal Pausing
by In Vitro Immobilized Template and Transcription Assays . . . . . . . . . . . . . . . . . 13
Heeyoun Bunch
3 Role of Heat Shock Factors in Stress-Induced Transcription: An Update . . . . . . 25
Heyoun Bunch and Stuart K. Calderwood
4 A Workflow Guide to RNA-Seq Analysis of Chaperone
Function and Beyond. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 39
Kristina M. Holton, Richard M. Giadone, Benjamin J. Lang,
and Stuart K. Calderwood
5 Chromatin Immunoprecipitation (ChIP) of Heat Shock
Protein 90 (Hsp90) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 61
Ritwick Sawarkar
6 Transfection and Thermotolerance Methods for Analysis
of miR-570 Targeting the HSP Chaperone Network . . . . . . . . . . . . . . . . . . . . . . . . 73
Yuka Okusha and Stuart K. Calderwood
7 Targeted Replacement of HSF1 Phosphorylation Sites at S303/S307
with Alanine Residues in Mice Increases Cell Proliferation
and Drug Resistance . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 81
Xiongjie Jin, Demetrius Moskophidis, and Nahid F. Mivechi
8 Bimolecular Fluorescence Complementation Assay to Evaluate
HSP90-Client Protein Interactions in Cells . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 95
Abir Chakraborty, Gregory L. Blatch, and Adrienne L. Edkins
9 Complementation Assays for Co-chaperone Function . . . . . . . . . . . . . . . . . . . . . . . 105
Adrienne L. Edkins and Gregory L. Blatch
10 Optimized Microscale Protein Aggregation Suppression Assay:
A Method for Evaluating the Holdase Activity of Chaperones . . . . . . . . . . . . . . . . 113
Ronald Tonui, Ruth O. John, and Adrienne L. Edkins
11 Detecting Posttranslational Modifications of Hsp90 Isoforms . . . . . . . . . . . . . . . . 125
Rebecca A. Sager, Sarah J. Backe, Len Neckers, Mark R. Woodford,
and Mehdi Mollapour
12 Multiple Targeting of HSP Isoforms to Challenge Isoform Specificity
and Compensatory Expression . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 141
Kisho Ono and Takanori Eguchi
xi
xii Contents

13 Using a Modified Proximity Ligation Protocol to Study

the Interaction Between Chaperones and Associated Proteins . . . . . . . . . . . . . . . . 163
Simone Baldan, Anatoli B. Meriin, and Michael Y. Sherman
14 Use of Native-PAGE for the Identification of Epichaperomes in Cell Lines . . . . 175
Tanaya Roychowdhury, Anand R. Santhaseela, Sahil Sharma, Palak
Panchal, Anna Rodina, and Gabriela Chiosis
15 Molecular Chaperone Receptors: An Update . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 193
Thiago J. Borges, Ayesha Murshid, Jimmy Theriault,
and Stuart K. Calderwood
16 A Novel Heat Shock Protein 70-Based Vaccine Prepared
from DC Tumor Fusion Cells: An Update . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 209
Desheng Weng, Stuart K. Calderwood, and Jianlin Gong
17 Methods to Assess the Impact of Hsp90 Chaperone Function
on Extracellular Client MMP2 Activity . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 221
SarahBeth D. Votra, Deema Alsalih, and Dimitra Bourboulia
18 Proteomic Profiling of the Extracellular Vesicle Chaperone in Cancer . . . . . . . . . 233
Kisho Ono and Takanori Eguchi
19 A Modified Differential Centrifugation Protocol for Isolation
and Quantitation of Extracellular Heat Shock Protein 90 (eHsp90) . . . . . . . . . . . 251
Cheng Chang, Xin Tang, and Wei Li
20 Immunohistochemistry of Human Hsp60 in Health and Disease:
Recent Advances in Immunomorphology and Methods for Assessing
the Chaperonin in Extracellular Vesicles . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 263
Celeste Caruso Bavisotto, Francesco Cappello, Everly Conway de Macario,
Alberto J. L. Macario, and Francesca Rappa
21 Multiplex Immunostaining Method to Distinguish HSP Isoforms
in Cancer Tissue Specimens . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 281
Hotaka Kawai, Kisho Ono, and Takanori Eguchi
22 Large-Scale Databases and Portals on Cancer Genome to Analyze
Chaperone Genes Correlated to Patient Prognosis . . . . . . . . . . . . . . . . . . . . . . . . . . 293
Kisho Ono and Takanori Eguchi
23 Immunohistochemical, Flow Cytometric, and ELISA-Based
Analyses of Intracellular, Membrane-Expressed, and Extracellular
Hsp70 as Cancer Biomarkers . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 307
Ali Bashiri Dezfouli, Stefan Stangl, Gemma A. Foulds, Philipp Lennartz,
Geoffrey J. Pilkington, A. Graham Pockley, and Gabriele Multhoff

Index . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 325
Contributors

ANDREW ACKERMAN • Geisinger Clinic, Danville, PA, USA

DEEMA ALSALIH • Department of Urology, SUNY Upstate Medical University, Syracuse, NY,
USA
SARAH J. BACKE • Department of Urology, SUNY Upstate Medical University, Syracuse, NY,
USA
SIMONE BALDAN • MRC Toxicology Unit, University of Cambridge, Cambridge, United
Kingdom
GREGORY L. BLATCH • Biomedical Biotechnology Research Unit (BioBRU), Department of
Biochemistry and Microbiology, Rhodes University, Makhanda, South Africa; Biomedical
Research and Drug Discovery Research Group, Faculty of Health Sciences, Higher Colleges
of Technology, Sharjah, United Arab Emirates; The Vice Chancellery, The University of
Notre Dame Australia, Fremantle, WA, Australia
GREGORY L. BLATCH • Biomedical Biotechnology Research Unit (BioBRU), Department of
Biochemistry and Microbiology, Rhodes University, Makhanda, South Africa; Biomedical
Research and Drug Discovery Research Group, Faculty of Health Sciences, Higher Colleges
of Technology, Sharjah, United Arab Emirates; The University of Notre Dame Australia,
Fremantle, WA, Australia
THIAGO J. BORGES • Center for Transplantation Sciences, Department of Surgery,
Massachusetts General Hospital, Harvard Medical School, Boston, MA, USA
DIMITRA BOURBOULIA • Department of Urology, SUNY Upstate Medical University,
Syracuse, NY, USA; Upstate Cancer Center, SUNY Upstate Medical University, Syracuse,
NY, USA; Department of Biochemistry and Molecular Biology, SUNY Upstate Medical
University, Syracuse, NY, USA
HEEYOUN BUNCH • Department of Applied Biosciences, Kyungpook National University,
Daegu, Republic of Korea
STUART K. CALDERWOOD • Molecular and Cellular Radiation Oncology, Beth Israel
Deaconess Medical Center, Harvard Medical School, Boston, MA, USA; Department of
Radiation Oncology, Beth Israel Deaconess Medical Center, Boston, MA, USA; Beth Israel
Deaconess Medical Center, Harvard Medical School, Boston, MA, USA
FRANCESCO CAPPELLO • Department of Biomedicine, Neurosciences and Advanced
Diagnostic (BiND), Human Anatomy Section, University of Palermo, Palermo, Italy
CELESTE CARUSO BAVISOTTO • Department of Biomedicine, Neurosciences and Advanced
Diagnostic (BiND), Human Anatomy Section, University of Palermo, Palermo, Italy
ABIR CHAKRABORTY • Biomedical Biotechnology Research Unit (BioBRU), Department of
Biochemistry and Microbiology, Rhodes University, Makhanda, South Africa
CHENG CHANG • Department of Dermatology the Norris Comprehensive Cancer Centre,
University of Southern California Keck Medical Center, Los Angeles, CA, USA
GABRIELA CHIOSIS • Program in Chemical Biology, Memorial Sloan Kettering Cancer
Center, New York, NY, USA
EVERLY CONWAY DE MACARIO • Department of Microbiology and Immunology, School of
Medicine, University of Maryland at Baltimore-Institute of Marine and Environmental
Technology (IMET) – Rita Rossi Colwell Center, Baltimore, MD, USA

xiii
xiv Contributors

ALI BASHIRI DEZFOULI • Radiation Immuno-Oncology Group, Center for Translational

Cancer Research Technische Universit€ a t München (TranslaTUM), Department of
Radiation Oncology, Klinikum rechts der Isar, Technische Universit€ a t München (TUM),
Munich, Germany
ADRIENNE L. EDKINS • Biomedical Biotechnology Research Unit (BioBRU), Department of
Biochemistry and Microbiology, Rhodes University, Makhanda, South Africa
TAKANORI EGUCHI • Department of Dental Pharmacology, Graduate School of Medicine,
Dentistry and Pharmaceutical Sciences, Okayama University, Okayama, Japan;
Department of Dental Pharmacology, Faculty of Medicine, Dentistry and Pharmaceutical
Sciences, Okayama University, Okayama, Japan
GEMMA A. FOULDS • John van Geest Cancer Research Centre, School of Science and
Technology, Nottingham Trent University, Nottingham, UK
RICHARD M. GIADONE • Department of Stem Cell and Regenerative Biology, Harvard
University, Cambridge, MA, USA; Harvard Stem Cell Institute, Cambridge, MA, USA;
Stanley Center for Psychiatric Research, Broad Institute of MIT and Harvard, Cambridge,
MA, USA
JIANLIN GONG • Department of Medicine, Boston University School of Medicine, Boston, MA,
USA
KRISTINA M. HOLTON • Department of Stem Cell and Regenerative Biology, Harvard
University, Cambridge, MA, USA; Harvard Stem Cell Institute, Cambridge, MA, USA;
Stanley Center for Psychiatric Research, Broad Institute of MIT and Harvard, Cambridge,
MA, USA
XIONGJIE JIN • Molecular Chaperone Biology, Medical College of Georgia, Augusta, GA,
USA; Georgia Cancer Center, Augusta University, Augusta, GA, USA
RUTH O. JOHN • Biomedical Biotechnology Research Unit (BioBRU), Department of
Biochemistry and Microbiology, Rhodes University, Makhanda, South Africa
HOTAKA KAWAI • Department of Oral Pathology and Medicine, Faculty of Medicine,
Dentistry and Pharmaceutical Sciences, Okayama University, Okayama, Japan
TOSHIKI KIJIMA • Department of Urology, Dokkyo Medical University, Tochigi, Japan
BENJAMIN J. LANG • Department of Radiation Oncology, Beth Israel Deaconess Medical
Center, Boston, MA, USA
PHILIPP LENNARTZ • Radiation Immuno-Oncology Group, Center for Translational Cancer
Research Technische Universit€ a t München (TranslaTUM), Department of Radiation
Oncology, Klinikum rechts der Isar, Technische Universit€ a t München (TUM), Munich,
Germany
WEI LI • Department of Dermatology the Norris Comprehensive Cancer Centre, University of
Southern California Keck Medical Center, Los Angeles, CA, USA
ALBERTO J. L. MACARIO • Department of Microbiology and Immunology, School of Medicine,
University of Maryland at Baltimore-Institute of Marine and Environmental Technology
(IMET) – Rita Rossi Colwell Center, Baltimore, MD, USA
ANATOLI B. MERIIN • Department of Biochemistry, Boston University, Boston, MA, USA
NAHID F. MIVECHI • Molecular Chaperone Biology, Medical College of Georgia, Augusta,
GA, USA; Georgia Cancer Center, Augusta University, Augusta, GA, USA; Department
of Radiation Oncology, Augusta University, Augusta, GA, USA
MEHDI MOLLAPOUR • Department of Urology, SUNY Upstate Medical University, Syracuse,
NY, USA; Department of Biochemistry and Molecular Biology, SUNY Upstate Medical
University, Syracuse, NY, USA
 Contributors xv

DEMETRIUS MOSKOPHIDIS • Molecular Chaperone Biology, Medical College of Georgia,

Augusta, GA, USA; Georgia Cancer Center, Augusta University, Augusta, GA, USA;
Department of Medicine, Augusta University, Augusta, GA, USA
GABRIELE MULTHOFF • Radiation Immuno-Oncology Group, Center for Translational
Cancer Research Technische Universit€ a t München (TranslaTUM), Department of
Radiation Oncology, Klinikum rechts der Isar, Technische Universit€a t München (TUM),
Munich, Germany
AYESHA MURSHID • Molecular and Cellular Radiation Oncology, Beth Israel Deaconess
Medical Center, Harvard Medical School, Boston, MA, USA
LEN NECKERS • Urologic Oncology Branch, Center for Cancer Research, National Cancer
Institute, Bethesda, MD, USA
YUKA OKUSHA • Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA,
USA; JSPS Overseas Research Fellow, Tokyo, Japan
KISHO ONO • Department of Oral and Maxillofacial Surgery, Okayama University
Hospital, Okayama, Japan
PALAK PANCHAL • Program in Chemical Biology, Memorial Sloan Kettering Cancer Center,
New York, NY, USA
GEOFFREY J. PILKINGTON • Brain Tumour Research Centre, School of Pharmacy and
Biomedical Sciences, Institute of Biomedical and Biomolecular Sciences, University of
Portsmouth, Portsmouth, UK
A. GRAHAM POCKLEY • John van Geest Cancer Research Centre, School of Science and
Technology, Nottingham Trent University, Nottingham, UK
THOMAS L. PRINCE • Ranok Therapeutics, Danville, PA, USA
FRANCESCA RAPPA • Department of Biomedicine, Neurosciences and Advanced Diagnostic
(BiND), Human Anatomy Section, University of Palermo, Palermo, Italy
ANNA RODINA • Program in Chemical Biology, Memorial Sloan Kettering Cancer Center,
New York, NY, USA
TANAYA ROYCHOWDHURY • Program in Chemical Biology, Memorial Sloan Kettering Cancer
Center, New York, NY, USA
REBECCA A. SAGER • Department of Urology, SUNY Upstate Medical University, Syracuse,
NY, USA
ANAND R. SANTHASEELA • Program in Chemical Biology, Memorial Sloan Kettering Cancer
Center, New York, NY, USA
RITWICK SAWARKAR • Department of Genetics, Medical Research Council (MRC), University
of Cambridge, Cambridge, UK
SAHIL SHARMA • Program in Chemical Biology, Memorial Sloan Kettering Cancer Center,
New York, NY, USA
MICHAEL Y. SHERMAN • Department of Molecular Biology, Ariel University, Ariel, Israel
STEFAN STANGL • Radiation Immuno-Oncology Group, Center for Translational Cancer
Research Technische Universit€a t München (TranslaTUM), Department of Radiation
Oncology, Klinikum rechts der Isar, Technische Universit€
a t München (TUM), Munich,
Germany
XIN TANG • Department of Dermatology the Norris Comprehensive Cancer Centre,
University of Southern California Keck Medical Center, Los Angeles, CA, USA
JIMMY THERIAULT • Molecular and Cellular Radiation Oncology, Beth Israel Deaconess
Medical Center, Harvard Medical School, Boston, MA, USA
RONALD TONUI • Biomedical Biotechnology Research Unit (BioBRU), Department of
Biochemistry and Microbiology, Rhodes University, Makhanda, South Africa
xvi Contributors

SARAHBETH D. VOTRA • Department of Urology, SUNY Upstate Medical University,

Syracuse, NY, USA
DESHENG WENG • Department of Medicine, Boston University School of Medicine, Boston,
MA, USA
MARK R. WOODFORD • Department of Urology, SUNY Upstate Medical University, Syracuse,
NY, USA; Department of Biochemistry and Molecular Biology, SUNY Upstate Medical
University, Syracuse, NY, USA
Exploring the Variety of Random
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Joy stared at him. What he said made her speechless with
astonishment at first, but she controlled herself enough to say.
“Stop by an’ see me an’ Ma, sometimes. Please, Cun April. We gits
so awful lonesome after dark.”
April promised he would. Promised in words that were very gentle.
Then he stalked on, a tall lonely shadow, moving under the trees.
April came to see Big Sue that very night, dropping by so
unexpectedly that the sight of him made her dumb for a while. She
tried to be natural, to hide her agitation, but her breath caught fast in
her throat every time she opened her mouth to talk, and her words
were uncertain and stammering.
But April paid little heed to her. He seemed scarcely to know she
was there, for his eyes spent much of the time looking at Joy. Breeze
thought he saw them flash once or twice, but it may have been the
firelight in them.
April declared he had eaten supper and cared for nothing either to
eat or drink, but Joy fixed him a cup of water, sweetened with wild
honey, flavored with bruised mint leaves, from the mint-bed by the
back door. When he tasted it he smiled, and the dull fire in the
chimney blazed up, and everything seemed brighter, more joyful
than in many a long day.
When April got up to go Joy followed him to the door. She made him
shake hands with her and promise to come back very soon.
After that when Joy walked out in the dusk she always let Breeze
know she’d rather go by herself. Not that she ever hurt his feelings,
but she made some sort of flimsy excuse to be rid of him. He hadn’t
shut up the coop where the hen and youngest biddies slept, or he
hadn’t cut up enough fat kindling wood, or couldn’t he go fetch a
fresh bucket of water from the spring?
Then Breeze discovered that April walked with Joy. He had forgotten
Big Sue altogether.
Once Breeze saw them walking shoulder to shoulder, arm touching
arm. They talked so softly their words were drowned by the rustle of
the leaves under their feet. When April stopped and bent his face so
close to Joy’s that she drew back a little, Breeze’s heart almost quit
beating. He let them go on unwatched, hidden by the deepening
twilight.
When Joy came home Big Sue grumbled as she handed her a
panful of supper and a spoon.
“How come you so love to walk out in de night? It ain’ good. You’ll
ketch a fever or somet’ing worse. You ain’ been home to eat supper
wid me since last Sat’day night was a week.”
But Joy sat mute, looking into the fire, with eyes that gleamed back
at the flames.
After that, Joy was always gentle, but except for her evening walk
she went nowhere, not even for the mail. For days at a time she
scarcely uttered a word. Lying on the bed, or sitting by the fire, she
did nothing but think, all the time. When visitors came she said she
wasn’t well, and went to lie down in the shed-room. Even Big Sue’s
constant scolding got few words out of the girl.
Late one afternoon Big Sue went to see Maum Hannah, whose
crippled knee was being troublesome. In the cabin a bright fire
blazed merrily, and Breeze and Joy shelled parched pindars to make
some molasses candy before time to cook supper. Breeze ran to
Zeda’s house to borrow a pinch of cooking soda to make the candy
foam up light. When he came back he found April talking to Joy in a
strangled husky voice. Both were standing up by the fire, the shelled
nuts were scattered on the floor; the smell of the molasses boiling
over and burning, made a bitter stench in the room.
“Wha’ you say, Joy?” April asked it very low.
Joy stood dumb, motionless, then she lifted her eyes to his face. “Is
you want me fo’ true, Cun April?”
His eyes were on her, so bold, so full of admiration, that she shrank
back in confusion, although her white teeth were flashing with
excitement.
April leaned closer and whispered, and her beaming eyes darted up
sidewise to see by his face if he meant all he was saying. She
reflected in silence, with a downcast look. But when she answered
him softly, she looked straight up again into his eyes.
His breath came quick. His eyes glinted fiercely. Joy drew back, but
she was nodding yes all the time. April caught her and squeezed her
to him and kissed her. She started struggling to free herself, but Big
Sue’s steps sounded outside and April hurried away out of the door.
Joy’s eyes followed him until the darkness had swallowed him, and
only the tramp of his feet could reach her ears. She pulled a chair up
to the fire and sat down, with her eyes fixed on the flames. She sat
there a long time. Once she smiled to herself, then she frowned, but
her eyes stayed glittering like a high spring tide under a full noon
sun.
“Joy,” Big Sue called her name sternly, “I b’lieve you’s conjured. I
know April is. Dat death-sheet is had him walkin’ his feet off ever
since Leah was buried. You’s a fool to let dat man talk wid you. I
wish to Gawd e’d stay way f’om my house.”

When the boat blew for the landing early next morning on its way to
town, Breeze and Big Sue had gone in Uncle Isaac’s cart to the lime
mill near the seashore to get lime enough to whitewash the front of
her house fresh for Christmas. Every cabin on the whole plantation
was being scoured and scrubbed and dressed up with papers. Big
Sue wanted hers to be the finest of all. Breeze had wrung next
year’s supply of straw brooms out of the old unplanted fields and had
swept the yard clean with a new dogwood brush-broom.
Joy had helped some, but in a half-hearted way. She wouldn’t even
ride out with them to get the lime. Her excuse was that Julia looked
wild. Breeze knew she didn’t mean it, for no mule ever moved more
sluggishly. Breeze had to get a stick and frail Julia to make her trot at
all.
Noon had passed when they got back home with their load.
Big Sue called Joy to see what nice white fresh-burned lime it was.
Like flour. Not a lump in it. But Joy was not at home and Big Sue
grumbled.
“Gone to Zeda’s again. Joy keeps hankerin’ to hear news from
Sherry. E may as well quit dat. Sherry’s gone! Fo’ good! E ain’ got
Joy to study ’bout! Not no mo’! No!”
When the sun went down, a great red ball, floods of brilliant light
gushed up around it, foretelling a cold night and a windy day to-
morrow. Water birds flew over the rice-fields, crying out in dread. The
trees were full of sighs. The open window blinds creaked dismally. A
puff of smoke came down the chimney. Winter was coming.
Dusk fell and the night closed in dark. Joy’s supper waited on the
hearth. Where could she be so late?
Breeze went to ask Zeda, but she wasn’t at home. Maum Hannah’s
house was dark, so he stopped at Bina’s to ask if any one there had
seen Joy lately.
Bina looked at him with searching eyes, “You is tryin’ to be smart,
enty? A-actin’ fool to ketch sense!” She sucked her teeth scornfully,
but Breeze didn’t understand what she meant.
“Don’ stan’ up an’ lie to me, boy! You know Joy an’ April went off on
de boat dis mawnin’.”
Breeze could scarcely believe his ears heard Bina right.
Joy and April gone? Together? Where had they gone?
Why hadn’t Joy told somebody?
He flew to tell Big Sue.
Instead of meeting the news with an outburst of grief, Big Sue
chuckled, “Who’d ’a’ thought my Joy could catch April! An’ Leah not
yet cold in her grave! Lawd! April’s old enough to be Joy’s daddy!
Well, all I got to say is dis! April was born fo’ luck. E ever did git de
best o’ ev’yt’ing on dis plantation.”

The boat was due to return three days hence. When the time came
the whole plantation was at the landing to meet it.
As the old battered hulk hove in sight, around the bend, a hush fell
on the crowd, and every eye was fixed on the lower deck where April
and Joy stood, side by side, smiling happily. April took off his hat and
waved it. Joy fluttered a handkerchief to greet them.
They were both dressed fit to kill. Joy, gay as a peacock, in a dress
striped with yellow bands, and a hat with green ribbons and red
flowers. April looked youthful in a brand-new suit that showed off his
broad shoulders and slim waist well. He held Joy’s hand and led her
carefully over the unsteady gangplank, and she fell into Big Sue’s
arms while April looked on smiling and rubbing his hands awkwardly.
The crowd crushed around them, wishing them happiness, hoping
they’d live like Isaac and Rebecca, wishing them joy and a gal and a
boy. Breeze pressed forward too until he could touch Joy’s hand,
and she bent down and gave him a smacking kiss, then a hug.
“Looka li’l’ Breeze, Cun April,” she said, and April reached out and
shook his hand, and Joy added; “I done told you I was gwine be you’
mammy, Breeze, and Cun April’s you’ daddy, now.”
The people crammed too close around them. Breeze could scarcely
breathe. He got out quickly as he could, and went to the store steps
to wait with old Louder, who sat wagging his tail, and making short
whines of pleasure. Breeze and Big Sue, and most of the neighbors,
went with them to April’s cabin, where a huge fire was built, and the
whole room made light as day.
Big Sue and Bina bustled around cooking supper, and April’s
children and Breeze all helped. Sweetened bread and fried bacon
and coffee with plenty of cream and sugar, were passed around. The
cabin was filled with the fragrance of the food. But Joy couldn’t eat.
Big Sue pressed her to take something, but she said she couldn’t
swallow a bite to save her life.
April had eyes only for Joy. He leaned over and whispered softly, “Is
anyt’ing ail you, honey?”
But she shook her head. She was only weary, too weary to eat.
Some of the young folks suggested a dance, but April said they must
come back another night; Joy was weary. The boat trip was long,
and the chill of the river wind had her trembling yet.
When everybody had something to eat and drink, they said good
night, and tramped out into the night, Breeze and Big Sue last of all.
The dark roads and paths swarmed with merry people, the air rang
with songs and laughter.
“April sho’ is a fool over Joy!” Big Sue grunted as they turned into the
path toward home. “A pure fool. A ol’ fool is de worst fool too.”
Joy and April took supper with Big Sue Christmas Eve, and they
helped fill Breeze’s stocking. He knew, for soon after supper he was
sent to bed. They were in a hurry to get to Maum Hannah’s house
where an all-night meeting was to be held.
Breeze wanted to go too. He wanted to stay up for all the singing
and shouting, and see the cows kneel down and pray at midnight,
and the sun rise shouting in the east in the morning. But Big Sue
said he was too sleepy-headed for her to fool with him, and if he
didn’t go to bed like a good boy old Santy Claw would leave his
stocking empty.
They all said good night and went out of the door and Breeze
thought they had gone for good. He was about to hop up and look at
his stocking when Joy ran back in, and, falling on the bed where he
was, burst out crying.
What on earth! Big Sue and April hurried in, and did all they could to
quiet her. Was she sick? Had somebody hurt her feelings? April
petted her and called her tender names, but she cried on even when
her tears were spent and broken sobs shook her of their own free
will.
Big Sue called April into the other room and whispered to him. He
came back and asked Joy if she wouldn’t rather stay quietly with
Breeze and rest? He’d stay too if she liked, or go to meeting with Big
Sue. Whatever she wanted was the thing he wanted too. She got up
and wiped her eyes. She’d go home and go to bed. He could do
whatever he liked. Her words sounded cold, almost bitter.
But soon the next morning she came to show Big Sue the Christmas
presents April had given her. A watch to wear on her wrist, and a
diamond ring! The two must have cost twenty-five dollars, if not
more.
The winter days passed slowly, many of them dull, gray, with an
overcast sky, where low clouds sailed and cast their murky color
over the ground. The first March day came in bright and warm, with a
wind that roared over the land, whipping the trees, snapping off their
rotten limbs, lifting old shingles off of roofs, sweeping yards and
woods clean, thrashing fields until clouds of dust and sand rose and
floated in the sky. But everybody rejoiced that winter was over and
gone. And besides, a windy March is lucky. Every pint of March dust
brings a peck of September corn, and a pound of October cotton. Let
it blow!
Such a high wind could never last. A March that comes in like a lion
will go out as quiet as a new-born lamb. Let it blow! But watch the
fires! One little spark can easily be fanned into a flame.
New leaves quivered and glittered on the restless boughs. Old
leaves, dead for months on the ground, hopped out from their
resting-places and skipped and flew, making brown leaf whirlwinds
that spun around dizzily, then settled in new sheltered places.
The wind lulled a little at sunset, and the night fell black and
cloudless. A multitude of stars crowded the sky, foretelling rain close
at hand. The rain was waiting for the blustery gale to hold still so the
clouds could gather and agree. In the night the wind rose and beat
against the cabin’s sides. It shook the walls, and whistled and
whined through the cracks. The front door banged wide open, as the
nail that held the bar frame was jerked out by its force. Finally Big
Sue made Breeze get up and get a hatchet and a long nail out of the
tool-box Santy Claw had given him, and she held the door while he
nailed it up.
Big Sue was frightened. She kept talking to Breeze, trying to keep
him awake with her, but he was too sleepy-headed to listen. When
he woke at dawn a flood of rain was pouring down, and thunder
roared louder than the rain or wind.
As a fearful crash shook the earth. Big Sue opened the back door
and peeped out and quavered, “Git up, Breeze! Lightnin’ is struck dat
big pine yonder, close to April’s house! It’s afire! Dat bolt shooken de
whole earth. I bet April’ll find it. Lawd! E’s been diggin’ at de roots o’
struck trees to git a bolt a long time! An’ now one mighty nigh hit
him!”
“What’s a bolt, Cun Big Sue?”
The wind howled as she answered, “Why, son, a thunderbolt is a’
iron rod. If you finds one, you’ll have de power to rule life an’ death!”
The cabin was closed tight, yet so fierce was the lightning it blazed
through cracks right into the room. Blood-red streaks of light took
turns with others that were blue. Breeze shut his eyes and put the
pillow over his head. He finally dozed off, and slept until the morning
had come, clear of rain and wind, and filled with the warm breath of
the earth.
He was alone. Big Sue had gone to see April’s struck pine, so he
dressed and ran to see it too.
A crowd of people were around the burning tree, and others were
coming. All were talking excitedly. God must have His eye on April to
aim a thunderbolt so close to his house. He had a narrow escape.
His house might catch fire yet, for pieces of burning limbs were
falling, and water could not put out fire started with lightning. Nothing
could, but new milk from a cow with her first calf. Where would April
get enough of that to do any good?
April was brazenly unafraid. He laughed at the notion of getting a
heifer’s milk. He said he’d make water outen this fire, or any other
fire, that bothered his cabin. They’d see.
April sat in front of the fire on his hearth, and when Big Sue fixed his
breakfast in a pan and handed it to him, he called to the neighbors,
standing outside, “Yunnuh come an’ eat some breakfast wid me.
We’s got a-plenty fo’ ev’ybody.” At first all of them answered, “No,
thank you,” but when April insisted, a half-dozen or more went in and
took a piece of bread, or a mouthful of sweetened water.
“How’s Joy?” Bina asked Big Sue politely.
“Joy’s awful nervish since dat tree got struck. I made em stay in bed
dis mawnin’.”
“Joy ain’ been well in a good while,” Bina commented.
Big Sue’s eyes snapped. “Joy ever was a delicate child, Bina. You
know dat good as me.”
The thick high trees, lapping their branches overhead, sheltered the
cabins from a sun that burned down, fierce and bright, drawing a
strong steamy stench up from the heated mud flats left naked by the
outgone tide.
The fields were all too wet for plowing, and the blacksmith shop was
the center for the day’s work. Plowshares needed to be filed and
sharpened. Plow-stocks mended. Mules’ feet trimmed. Manes and
tails clipped short. A few of the older, thinner beasts had got lousy.
The hair must be cut off them and their hides wet with tea made out
of china-berry leaves.
The men laughed and talked and chewed tobacco and smoked, as
they worked leisurely at their different tasks. A difference of opinion
rose as to the best place to twitch a mule to make him stand still for
his hair to be cut off. A twine-string could be twisted around an ear,
or tied to the upper lip. Uncle Bill preferred the lip. He said mules
have pockets inside their ears and a string twisted tight enough to
hold the beast quiet, will tear that pocket in two. April objected to the
twitch on the lip, for it often caused a painful swelling.
The question was still unsettled when Brudge came running hard as
he could, crying out that Joy had been taken with a death-sickness.
She was lying on the bed in a trance. She couldn’t speak a word.
Brudge almost popped out his eyeballs showing how her eyes were
rolled away back in her head. Her hands and feet were cold as
clabber. Big Sue said April must hurry or Joy would be gone before
he got there!
April did not wait to hear the end of Brudge’s talk, but flew home
ahead of them all with Breeze close at his heels. Lamentations and
outcries met them as they got nearer. Big Sue’s above all the rest.
Joy was dying. Nothing but a death-sickness could strike a young
woman down so hard.
Breeze was almost petrified with terror, but he dragged himself on to
the cabin, which was already filled with the neighbors. Joy lay on the
bed covered over with a quilt, up to her very neck. Her eyes were
shut. Her head moved from side to side. Her lips whispered things
nobody could hear at all.
Big Sue sat near the bed in a low chair, her fat body rocking. Big
tears rolled down her cheeks as she chanted over and over.
“Do, Jedus! Don’ let Joy dead!
“Oh, my Gawd! Help my chile! Help em!
“Oh, Lawd! Oh, my Gawd!
“Don’ let Joy dead dis mawnin’!”
April broke through the crowd surrounding the bed, and taking one of
Joy’s hands from under the cover felt her pulse, then leaned over to
hear what she was saying. “No. No, honey,” he crooned, “you
wouldn’ dead an’ leave me. No. No. I couldn’ do widout you nohow. I
wouldn’ ’a’ left you last night in dat storm, but I was ’fraid de stables
would blow down an’ kill all de mules an’ horses. De storm is gone.
De lightnin’ didn’ hurt nobody. Death is gone away off now. E can’
take you. No!”
Breeze pricked up his ears. Was death about to take Joy?
As her life fluttered uncertainly, Big Sue’s wailing and misery were
less hard to bear than April’s fierce resolute manner.
Joy had to get well. No matter what ailed her. If she was conjured,
Uncle Isaac had to take off the spell. If the storm had scared her until
her heart-string was strained, she must keep still and rest until it
went back into place. Nobody must come in the room to worry her
with talking. Send for Maum Hannah. No matter if she was at the end
of the earth instead of the end of the “Neck,” go fetch her! Hurry!
Don’t tarry and waste any more time! Fetch Maum Hannah! Joy had
a death-sickness!
Uncle Bill hitched up the fastest horse in the barnyard to the lightest
cart, and went flying down the road for Maum Hannah, who had
gone to a sick woman some miles away. When he got back, several
hours later, the horse was lathered with sweat, and all but broken
winded, but Joy was still alive.
The room was chock-full, the door choked with people, both windows
were dark with heads. Big Sue’s mourning that had fallen into a low
mumbling prayer to Jesus now changed and livened to:
“Do, Maum Hannah! Help my Joy!
“Do, Maum Hannah! Don’ let Joy dead!
“Do, Maum Hannah!”
Maum Hannah hurried up the steps as fast as her crippled knee
would let her. She was all out of breath, but instead of pitying Big
Sue, she stopped still and eyed her with an impatient grunt. “Do shut
you’ mouth, Big Sue! You ought to be shame’ to cut all dis crazy! You
can’ fool dese people. No! Everybody knows wha’ ails Joy, ’ceptin’
April. An’ e ought to take you out an’ duck you good in de creek fo’
makin’ such a fool out o’ him! Dat fine horse is most dead! Bill made
em run so fas’, de wind likened to ’a’ cut my breath off. You people
go home. Gi’ Joy a chance to turn dat chile loose. Joy done well to
hold em dis long but e can’ hold em no longer. Yunnuh go on! Go on,
Breeze! Yunnuh clear de room!”
Big Sue stopped grieving and stared, but Maum Hannah’s talk stung
April to the quick. He stepped up to her angrily, but she stopped
taking off her cloak long enough to pat his arm, “Don’ be vexed wid
me, son. I’m tellin’ yunnuh what Gawd loves, de truth! Joy’s done
well to hold dat chile dis long. You married in de Christmas, enty?
Well, Joy can’ hold em six more months. I know dat. Dat gal’s got to
turn em loose, no matter if it do hurt you’ feelin’s!”
Joy trembled like a leaf in a storm. Her dazed eyes turned from
Maum Hannah to April, who was silent, except for one word. “Bitch,”
he snarled, and his eyes blazed like lightning flashes, as he turned
and left the room.
Breeze left too, but he scarcely knew where he was, or where he
was going. April had cursed Joy and she a-dying!
He dragged himself home and fell across Joy’s own bed, for Big Sue
was not there to stop him. He wept until his tears failed him. He tried
his best to pray, “O Gawd, don’ let Joy die—” but he went fast
asleep. He slept heavily until a harsh hoarse voice waked him. He
came instantly to his senses, and tried to stammer out some excuse,
but Big Sue’s grim swollen face made his words falter, and the slap
her hand laid on his jaw brought shining stars in front of his eyes.
“Git up and go borrow a piece o’ fire f’om somebody! Hurry, too,
befo’ I kill you!”
He ran to April’s house, but stopped at the step for a tiny baby was
crying inside. He ran all the way to Zeda’s and borrowed a piece of
fire, then flew home. As he made up the fire for Big Sue, she walked
around the room unsteadily, mumbling between her teeth. If April
mistreated her Joy now, she herself would put a “hand” on him; one
so strong that it would wither his hands! And his feet! She couldn’t
keep still or stop talking. Her tongue lashed April and Joy too, and
each word was a poison sting. Who was he to blame Joy? He had
children scattered from one end of this “Neck” to the other. Now he
cursed Joy as if she were lowest of the low. It was a shame! A heavy
shame! Joy must leave him at once!
The wind had risen and whistled through the trees, tossing the
branches, making them moan. Big Sue talked on and on. Breeze
was glad when she went back to April’s cabin, although she left him
without a bite to eat. He’d go somewhere and get supper. Maybe
Uncle Bill was at home. He’d go see.
To his surprise April was there too, sitting by the fire, miserably
dejected, while Uncle Bill talked to him, trying to cheer him.
Breeze had hardly got inside when Zeda arrived and, brushing past
Uncle Bill, walked up to April and put a hand on his shoulder. “Look
at me, April. I got somet’ing to say to you.” Bitter spite hurried her
words.
But April, instead of looking up as she bade him, leaned forward and
spat in the fire.
“Wha’ ails you, now, Zeda?” he asked curtly.
“I kin easy say what ails me; dat new-born child yonder ’side Joy is
my gran’! But e’s you’ gran’, too! Joy had dat chile for Sherry, an’ you
ain’ gwine put no dis-grace-ment on em. No. If nobody else can’
hinder you, I kin. I already got you’ feet so dey can’ rest. Wid Leah’s
death-sheet.”
April heard her, and although he didn’t answer, his jaw set his teeth
hard enough to bite a ten-penny nail in two. Zeda smiled.
“You may as well give in, April,” she persisted. “Sherry’s you own, an’
who is Breeze, but you’ own? Ev’body knows dat. It’s a wonder
somebody ain’ cut you’ throat long time ago. If you wa’n’t so lucky
you’d ’a’ been in hell wid some o’ dem women you sent dere.”
“How come you meddlin’ in my business so raven?” April suddenly
flashed out.
“Dat li’l’ chile is my business. Joy had em fo’ Sherry, a li’l’ boy-chile,
too. You go on home an’ tell Joy to hurry up an’ git well. ’Tain’ no use
to hold hard feelin’s ’gainst em. No! Joy’s had you a gran’son.”
When he did not stir, she blazed out: “You’ neck is stiff, enty! So’s my
own! An’ I hope a misery’ll gnaw you’ heart in two. I hope you’ll die of
thirst an’ hunger. I hope ev’y lawful yard-chile you had by Leah’ll
perish. I hope you’ feet’ll rot——”
“You shut you’ mouth, Zeda. If you cuss me again I’ll choke you’
tongue down you’ throat.” April got up and fled from her bitter words.
 XVIII
JOY AND APRIL

For days after Joy’s child was born, Big Sue kept to her cabin. Joy
had disgraced her, made her ashamed to show her face in company.
She’d never forgive Joy as long as she lived. Never. Joy saw Leah
drop dead in her face, yet she went straight on and married Leah’s
husband. A shame! Joy would sup sorrow yet. She might bewitch
April and make a plumb fool out of him, but she’d pay for bringing
disgrace on her mother who had worked her knuckles to the bone to
keep Joy in school!
If Joy had behaved herself, she might have married anybody instead
of a man old as her daddy, and conjured to boot. That death-sheet
had put a spell on April. Sure as preaching. He’d never be the same
man again. He’d have run Joy out of his house if he had been in his
right mind.
She talked so fast and loud one morning she didn’t see Uncle Bill
until he was at the door-step. “How come you tiptoes around so easy
dis mornin’!” she asked tartly.
“Gawd knows how I’m a-walkin’, I’m so fretted.”
“Wha’ dat ail you now?”
“Joy sent me to tell you.”
“How come Joy don’ fetch e own answer?”
“Joy’s too troub-led.”
Big Sue shot a look at him and sucked her teeth. “Joy’s mighty late
gittin’ troub-led,” and a hard, wicked smile touched her mouth.
“Joy’s troub-led about April. April ain’ well, Miss Big Sue.”
Big Sue sniffed and said April was due to have something wrong
with him, wicked as he had lived, hard as he had been with
everybody that crossed him. What kind of sickness did April have?
“Somet’ing ails his feets.”
“Dat ain’ surprisin’. April slept wid a death-sheet on ’em a whole
night.”
“Uncle Isaac took dat spell off em.”
“Well, who put dis spell on em, den?”
Uncle Bill sat down on the step. He was so troubled in his mind, it
was difficult for him to say what ailed April. At first it favored
chilblains; then ground-itch, for April went out barefooted in the dew
every morning God sent, and any little scratch that lets dew get
inside your skin may give you ground-itch. But none of the chilblain
or ground-itch cures helped him at all. His appetite was clean gone.
He had eaten nothing but spoon-victuals for a week. He was thin as
a fence rail.
Big Sue made an ugly mouth. What did she care? Why hadn’t April
married a settled woman who could cook decent rations instead of a
scatter-brained girl like Joy who didn’t know the name of one pot
from another? He needn’t be sending word here about victuals. Let
April eat what Joy fixed for him. Love would season up lumpy
hominy and make doughy bread taste good.
Uncle Bill sat frowning, chewing his tobacco wearily, studying. Joy
had said she hated to worry Big Sue. She was sorry for all that had
happened. Joy was a good girl. She had slipped up once, and made
a bad mistake, but any young inexperienced girl is likely to miss and
do that. April did right to excuse her.
Big Sue sneered. Joy had worked one sharp trick. Leah herself
couldn’t have fooled April any slicker. Joy ever was tricky, though.
Just like Silas for the world. Likely as not, Joy had April conjured
right now.
Uncle Bill pursed up his lips so tight, they looked as if they’d never
open and speak again, and his eyes were full of worry.
“Whyn’ you go see Uncle Isaac? E might could help April?” Big Sue
asked presently.
“I done seen em. When de bear-grass poultices and de violet-leaves
tea failed, I went an’ got Uncle Isaac. Joy sent me. I don’ like
charms. I don’ trust ’em. I know a Christian man ain’ got no business
foolin’ wid ’em. But Joy was so fretted, I done it to please her. I kept
a-studyin’ over it; one mind said do it; another mind said, no, I better
ask all de Christian people to hold a prayer-meetin’ an’ ask Gawd to
help April; I listened at dem two minds arguin’ one whole night befo’ I
give in to Joy. An’ now I wish to Gawd I didn’ heed em.”
“How come so? April wouldn’ wear em, I bet you!” Big Sue was
listening with interest now, anxious to know what happened, but
Uncle Bill took his time.
April did everything Joy said. Wilful and unruly as he was with every
one else, he tried to please Joy. And yet when Joy brought that
charm to him and began coaxing him to let her tie it around his neck,
he balked. Joy had to outtalk him.
For a whole day and night April wore it, a little cloth bag, tied with a
white horse hair; but because it didn’t cure his feet right away, he
jerked it off and threw it in the fire. Such a pity. Even strong charms
take time to start working. April ever was a short-patienced man. He
made trouble for himself by hastiness. A man can be hot-blooded
and pettish with people but not with charms or magic.
Joy snatched the bag out of the flames, but it was scorched and a
hole burned in one side. A speck of the mixture inside it spilled out
on the coals and smoked such a strong smoke, April sneezed three
times!
Right then, the gristles in April’s feet got hard. Hard as a rock! God
only knew if they’d ever go back to their rightful softness.
Uncle Isaac made Joy take the bag off to the woods and bury it at
the foot of a locust tree, but April got worse and worse. His feet were
numb and hard and dry. Joy wanted to send for a white doctor. They
might get one to come on the boat from town, and with the crop so
promising they’d have money to pay him next fall. But April wouldn’t
have it. He said Maum Hannah knew more than any white doctor.
Big Sue kept shaking her head and grunting shamelessly until Uncle
Bill got up painfully to go. Something in his sad face must have
moved her, for all of a sudden she scrambled to her feet, letting her
scraps fall on the floor. “I made some nice little sweetened breads
dis mawnin’. Take some to April. I sho’ am sorry ’bout his feets. You
tell em so. I’m gwine broil em a fat pullet, too.”
“Ev’y man has to manage his own dueness, but how ’bout gwine
along wid me, to see April, Miss Big Sue? You done chastise Joy
long enough. De gal’s in trouble.”
“I can’ go, not so well, right now, Uncle Bill, but Breeze kin go if e’ll
thread me two or three needles first.” She started to say more, but
she changed her mind and kept silent, her eyes cast down on her
sewing. When she did speak it was to say Joy had been mighty shut-
mouthed about April. Joy had funny ways.
Breeze and Uncle Bill found April with a quilt around him, sitting
alone by the fire, looking at his feet. Looking and looking. His heavy
black brows overshadowed his sad eyes as they lifted and hovered
over Breeze, then Uncle Bill. But as soon as he shook hands and
said “thank you” for the food, they fell, and settled on his feet, which
were bare and on the hearth very close to the fire.
The weather had turned off cool in the night, but there was no
reason for April to keep his feet so close to the fire. Uncle Bill told
him he’d scorch them, but April shook his head and said they felt no
heat at all. Not a bit. They had gone to sleep or something. They felt
like blocks of wood. And he moved them stiffly, as if they were.
He complained that he had no appetite. He was tired too. Sitting still
was the hardest work he had ever done in his life. If he could read,
or if he had somebody to talk to, if he had something pleasant to
think about, it would help pass the time. But he couldn’t read, and he
didn’t want anybody to stay at home out of the field. Cotton needs
fast hoeing during these warm wet days. He wished he could stop off
thinking. Stop short off. He’d like to go to sleep and never wake up
any more. He’d go crazy if he had to stay still and look at his feet
much longer. What in God’s name ailed them! Nobody seemed to
know!
Uncle Bill tried to tell him the plantation news, but April’s eyes stayed
on his feet. Uncle Bill offered to teach him to read if he wanted to
learn. Now would be a good time for April to learn how to write. He
ought to learn to write his name if no more. Every man ought to know
how to write his name. But April said he never had much faith in
books and reading. Black people were better off without it. It takes
their mind off their work. It makes them think about things they can’t
have. They’re better off without knowing how. Uncle Bill didn’t argue.
All of a sudden a coal popped out of the hearth with a sharp
explosion. It fell right between April’s feet, as if it could see and did it
on purpose. It lay there, red, bright, like a dare. April opened his tired
eyes wide, and leaned forward and looked at it, for instead of dying
out it burned freer. April carefully raised one long black bony foot and
placed its heel on the coal. He waited a moment; then he lifted it up
and stared at Uncle Bill. His scared eyes told what had happened.
Breeze knew too.
April had felt no heat. His foot was dead. It couldn’t feel fire! April
grabbed the fire shovel, and scraped up a batch of live coals from
under the fire and dropped them on the hearth. He’d see if fire had
stopped being hot. Uncle Bill didn’t raise a finger to stop him when
he lifted his other foot and pressed its heel down on the coals and
mashed hard on them.
The bitter smell of his burned flesh stung the air. April’s eyes glared,
and he laughed a harsh discordant laugh. But a sob quickly caught
him by the throat and choked him. He leaned over and picked up a
live coal in his fingers, then dropped it quickly, for his fingers were
alive. They could feel. The coal burned them. But his feet were dead.
They couldn’t feel even fire!
“Oh, Gawd!” he moaned, and his long fingers knotted and clenched,
his strong tobacco-yellowed teeth ground together.
Joy came in from the field to feed her little baby. Nobody heard her
bare footsteps, until she spoke to Uncle Bill and Breeze. She went
up to April and put a hand on his arm, and asked how his feet were.
She leaned over and looked at them, but he drew them underneath
his chair. He didn’t want her to see. He reached for the quilt on the
floor beside him and covered them over.
“My feets is all right,” he told her gruffly.
But Joy sniffed the air once or twice, she searched the fire with her
eyes, then she swept the hearth clean of the coals. She patted
April’s shoulder, and said gentle things to him. He must have
patience. She’d make some fresh violet-leaves tea and soak his feet.
She was sure that would help them.
Bright tears ran out of April’s eyes, down his thin hard cheeks, and
fell on the bony clasped hands that held tight to each other in his lap.
Breeze could hardly bear to see those tears. Uncle Bill got up and
tried to say something, but his voice broke, and he began punching
the fire. For April was crying out loud. Saying he had given out! He
couldn’t go on any longer!
Joy put her arms around him and held his head on her bosom, and
patted his face and tried to hush him. She wiped his tears away with
her homespun apron, and smoothed his eyelids softly. Her fingers
were trembling, but April became quieter. She stroked his head and
begged him to go back to bed and lie down and rest.
He was hard and sullen, and frowned as if she had insulted him.
He’d stay right where he was. Bed and chair were the same to him
now. Joy stood with her eyes on the red embers, never answering
back a single time, even when anger made the words strangle in his
throat. It was hard for him to bend his neck under such a galling,
hellish yoke.
Until now he had never asked a favor of anybody in his whole life.
He had always worked, and made others work. His women and
children too. And now his feet, the feet that had carried him faithfully
through all these years, the only ones he could ever have, had failed
him. They made game of him. And it was more than he could bear.
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