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A Genomic and Transcriptomic Analysis of Wood Decay and Copper Tolerance in the Brown Rot Fungus Fibroporia Radiculosa

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A Genomic and Transcriptomic Analysis of Wood Decay and Copper Tolerance in the Brown Rot Fungus Fibroporia Radiculosa Mississippi State University Scholars Junction Theses and Dissertations Theses and Dissertations 1-1-2011 A genomic and transcriptomic analysis of wood decay and copper tolerance in the brown rot fungus Fibroporia radiculosa Juliet D. Tang Follow this and additional works at: https://scholarsjunction.msstate.edu/td Recommended Citation Tang, Juliet D., "A genomic and transcriptomic analysis of wood decay and copper tolerance in the brown rot fungus Fibroporia radiculosa" (2011). Theses and Dissertations. 143. https://scholarsjunction.msstate.edu/td/143 This Dissertation - Open Access is brought to you for free and open access by the Theses and Dissertations at Scholars Junction. It has been accepted for inclusion in Theses and Dissertations by an authorized administrator of Scholars Junction. For more information, please contact [email protected]. Template Created By: James Nail 2010 A GENOMIC AND TRANSCRIPTOMIC ANALYSIS OF WOOD DECAY AND COPPER TOLERANCE IN THE BROWN ROT FUNGUS FIBROPORIA RADICULOSA By Juliet Dao-May Tang A Dissertation Submitted to the Faculty of Mississippi State University in Partial Fulfillment of the Requirements for the Degree of Doctor of Philosophy in Forest Resources in the Department of Forest Products Mississippi State, Mississippi December 2011 Template Created By: James Nail 2010 Copyright 2011 By Juliet Dao-May Tang Template Created By: James Nail 2010 A GENOMIC AND TRANSCRIPTOMIC ANALYSIS OF WOOD DECAY AND COPPER TOLERANCE IN THE BROWN ROT FUNGUS FIBROPORIA RADICULOSA By Juliet Dao-May Tang Approved: _________________________________ _________________________________ Susan V. Diehl Shane C. Burgess Professor of Forest Products Dean of the College of Agriculture and (Director of Dissertation) Life Sciences, University of Arizona (Committee Member) _________________________________ _________________________________ Andy D. Perkins M. Lynn Prewitt Assistant Professor of Computer Science Assistant Research Professor of and Engineering Forest Products (Committee Member) (Committee Member) _________________________________ _________________________________ Darrel D. Nicholas Daniel G. Peterson Professor of Forest Products Associate Professor of Plant and Soil (Committee Member) Sciences (Committee Member) _________________________________ _________________________________ Rubin Shmulsky George M. Hopper Professor and Head of Forest Products Dean of the College of Forest Graduate Coordinator Resources Template Created By: James Nail 2010 Name: Juliet Dao-May Tang Date of Degree: December 9, 2011 Institution: Mississippi State University Major Field: Forest Resources Major Professor: Dr. Susan V. Diehl Title of Study: A GENOMIC AND TRANSCRIPTOMIC ANALYSIS OF WOOD DECAY AND COPPER TOLERANCE IN THE BROWN ROT FUN- GUS FIBROPORIA RADICULOSA Pages in Study: 122 Candidate for Degree of Doctor of Philosophy Brown rot fungi are notoriously copper-tolerant, which makes them difficult to control with copper-based wood preservatives. Brown rot fungi are also unique because they have evolved a bilateral strategy for decay. Their initial attack involves the produc- tion of hydroxyl free radicals to increase wood porosity, followed by an enzymatic on- slaught of glycoside hydrolases that free the sugars locked within cellulose and hemicel- lulose. Our molecular understanding of these biological processes, however, has been hampered by our limited knowledge of the underlying genetic mechanisms. To address this knowledge gap, high-throughput, short-read sequencing was used to conduct a comprehensive analysis of the genomics and transcriptomics of wood decay and copper tolerance in the brown rot fungus Fibroporia radiculosa. The results were impressively informative. In the genomic study, the sequences of 9262 genes were pre- dicted and gene function was assigned to 5407 of the genes. An examination of target motifs showed that 1213 of the genes encoded products with extracellular functions. By mining these genomic annotations, 187 genes were identified with putative roles in ligno- cellulose degradation and copper tolerance. Template Created By: James Nail 2010 The transcriptomic study quantified gene expression of the fungus growing on wood treated with a copper-based preservative. At day 31, the fungus was adapting to the preservative, and the wood showed no strength loss. At day 154, the preservative ef- fects were gone, and the fungus was actively degrading the wood, which exhibited 52% strength loss. A total of 917 differentially expressed genes were identified, 108 of which appeared to be regulating wood decay and preservative tolerance. Genes that showed in- creased expression at day 31 were involved in oxalate metabolism, hydroxyl free radical production by the enzyme laccase, energy production, xenobiotic detoxification, copper resistance, stress response, and pectin degradation. Genes that exhibited higher expres- sion at day 154 were involved in wood polysaccharide degradation, hexose transport, ox- alate catabolism, catabolism of laccase substrates, proton reduction, re-modeling the glu- can sheath, and shoring up the plasma membrane for acid shock. These newly discovered genes represent a significant step towards accelerating a genome-wide understanding of brown rot decay and tolerance to wood preservatives. Key words: systems biology, wood degradation, copper tolerance DEDICATION I dedicate this work to my loving husband, David C. Cross. iii ACKNOWLEDGEMENTS I have learned more than I ever could have hoped for during my graduate studies, and I owe it to my mentor, Susan Diehl. She has given me so many opportunities, the most important of which was to take me on as a graduate student, and then to give me a project that I feel has turned me into a real scientist. Because of her generous support, I have been able to publicize my work internationally and meet the icons of wood protec- tion. Moreover, she is an honest person, who truly cares about the success of her stu- dents. I will always admire her for these qualities. The other person I owe a world of gratitude to is Chuan-Yu Hsu. She is one of my Forever Friends, who also happens to be the best molecular biologist I know. I could always ask her anything, and I know her advice and expertise added enormous value to the quality of my research. I would also like to thank my co-authors, Shane Burgess, Andy Perkins, Darrel Nicholas, Tad Sonstegard, and Steve Schroeder. Shane funded the genomics study, Andy was my bioinformatics guru, Darrel helped fund the transcriptomics study and taught me how to run decay tests, and Tad and Steve ran my samples through the Genome Analyzer for the DNA sequence analysis. Their contributions to my research are deeply appreci- ated. Thanks are also due to Cetin Yuceer for letting me use his laboratory when I needed it, to Dan Peterson for telling me that careers are built on genomes, and to Lynn Prewitt, who served on my committee. iv TABLE OF CONTENTS Page DEDICATION................................................................................................................... iii! ACKNOWLEDGEMENTS............................................................................................... iv! TABLE OF CONTENTS.....................................................................................................v! LIST OF TABLES............................................................................................................ vii! LIST OF FIGURES ......................................................................................................... viii CHAPTER I. INTRODUCTION ...........................................................................................11! Wood Protection ..............................................................................................11! Biology of Brown Rot Decay ..........................................................................12! Biology of Copper Tolerance...........................................................................15! Knowledge Gaps..............................................................................................16! Challenges of the Wood Protection Industry...................................................18! Study Objectives ..............................................................................................19! References Cited ..............................................................................................21! II. SHORT-READ SEQUENCING FOR GENOMIC ANALYSIS OF THE BROWN ROT FUNGUS FIBROPORIA RADICULOSA...........25! Abstract............................................................................................................25! Introduction......................................................................................................26! Materials and Methods.....................................................................................29! Fungus Isolate ............................................................................................29! DNA Library Preparation ..........................................................................29! Short-Read Sequencing..............................................................................30! Stringency Filters.......................................................................................31! Assembly....................................................................................................31! Optimal Assembly Determination .............................................................32! Gene
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