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UC Riverside UC Riverside Electronic Theses and Dissertations Title Molecular Evolution of Silk Genes in Mesothele and Mygalomorph Spiders, With Implications for the Early Evolution and Functional Divergence of Silk Permalink https://escholarship.org/uc/item/8q80p6s5 Author Starrett, James Richard Publication Date 2012 Peer reviewed|Thesis/dissertation eScholarship.org Powered by the California Digital Library University of California UNIVERSITY OF CALIFORNIA RIVERSIDE Molecular Evolution of Silk Genes in Mesothele and Mygalomorph Spiders, With Implications for the Early Evolution and Functional Divergence of Silk A Dissertation submitted in partial satisfaction of the requirements for the degree of Doctor of Philosophy in Genetics, Genomics, and Bioinformatics by James Richard Starrett September 2012 Dissertation Committee: Dr. Cheryl Y. Hayashi, Chairperson Dr. Renyi Liu Dr. Mark Springer i Copyright by James Richard Starrett 2012 i i The Dissertation of James Richard Starrett is approved: ____________________________________________ ____________________________________________ ____________________________________________ Committee Chairperson University of California, Riverside ii i Acknowledgements The first chapter of this dissertation is a reprint of the material as it appears in PLoS ONE 7(6): e38084. doi:10.1371/journal.pone.0038084, published 22 June 2012. It is reproduced with permission from James Starrett, Jessica E. Garb, Amanda Kuelbs, Ugochi O. Azubuike, and Cheryl Y. Hayashi and is an open-access article distributed under the terms of the Creative Commons Attribution License. Co-authors Jessica E. Garb, Amanda Kuelbs, and Ugochi O. Azubuike provided research assistance. Co-author Cheryl Y. Hayashi supervised the research and provided lab materials. Research was supported by National Science Foundation (NSF) Doctoral Dissertation Improvement Grant DEB-0910365 to James Starrett and Cheryl Y. Hayashi, NSF DEB-0236020 and Army Research Office W911NF0610455 grants to Cheryl Y. Hayashi, and NSF DEB-0108575 grant to Marshal Hedin and Jason Bond. Funding was also provided by a Biology Newell Travel Grant Award, UCR Distinguished Graduate Fellowship in Biology, UCR Dissertation-Year Fellowship Award, and UCR Genetics, Genomics, and Bioinformatics Graduate Fellowship. I would like to thank Peter Arensburger, Nadia Ayoub, John Brown, Matthew Collin, Chinonyerem Oguguo, and Yonghui Zhao for providing help in the lab and with analyses. Nadia Ayoub, Matthew Collin, Jessica Garb, Marshal Hedin, Seung-Chul Kim, Dmitri Maslov, Robert Meredith, Morris Maduro, and Jordan Satler provided valuable comments on earlier versions of the dissertation chapters. John Gatesy offered comments that improved the dissertation considerably. Jason Bond, Dave Carlson, Frederick Coyle, iv James Cokendolpher, John Gatesy, Helene Lee, Norman Platnick, and Casey Richart helped with specimen collecting. Marshal Hedin, Dean Leavitt, Jordan Satler, and Steven Thomas were especially helpful in the field. I would like to thank my dissertation committee: Cheryl Hayashi, Mark Springer, and Renyi Liu for their valuable advice, support, and considerable flexibility. I am grateful to Cheryl Hayashi and Marshal Hedin for their significant contributions to the successes I have had so far in my academic pursuits. Both have dedicated a tremendous amount of time, energy, resources, and ideas to my projects. I would not be where I am without the opportunities they have provided me, and I have always been encouraged by their consistent enthusiasm and willingness to discuss all things spider, natural history, and molecular evolution related. v Dedication To my friends and family, for all of your support. If you ever need a kidney... vi ABSTRACT OF THE DISSERTATION Molecular Evolution of Silk Genes in Mesothele and Mygalomorph Spiders, With Implications for the Early Evolution and Functional Divergence of Silk by James Richard Starrett Doctor of Philosophy, Graduate Program in Genetics, Genomics, and Bioinformatics University of California, Riverside, September 2012 Dr. Cheryl Y. Hayashi, Chairperson The evolution of adaptively significant gene families is an important subject in the field of evolutionary genetics. For spiders, the gene families encoding silk proteins have received considerable attention due to the high performance capabilities of the fibers they produce. However, silk gene research has largely focused on spiders of the infraorder Araneomorphae, leaving much of the phylogenetic diversity of spiders unsampled for their silk genes. Here, I sample silk genes from spiders of the suborder Mesothelae and infraorder Mygalomorphae, which are distantly related to araneomorph spiders. Phylogenetic analyses of the spidroin genes indicate that numerous duplications occurred in the spidroin gene family after opisthotheles (mygalomorphs plus araneomorphs) split from mesotheles. However, while mesotheles appear to possess a single spidroin gene, they possess numerous copies of genes homologous to Egg Case Proteins, which are vi i currently only known from one araneomorph species. Together these results indicate that the common ancestor of extant spiders possessed a diversity of silk genes. In addition to higher level species sampling of silk genes, I sequence repetitive and carboxy terminal regions of spidroins from all species of the trapdoor spider genus, Aliatypus. Gene tree analyses and tests of selection suggest that contrasting evolutionary forces influenced the different regions of the spidroin gene. I also investigate the expression of silk transcripts from the tarsi and silk glands of tarantulas. Tarantulas exude silk-like secretions from their tarsi, which is hypothesized to increase surface adhesion. I discovered that while known spidroin silk genes are not expressed in the tarsi, novel silk-like genes are expressed in tarantula silk glands and tarsi. Gene families of adaptive significance may also show phylogenetic signal. Here, I sample hemocyanin gene family sequences from a phylogenetically diverse sample of spider species and infer gene trees and species trees. Phylogenetic analyses reveal that despite instances of lineage specific duplication and loss of hemocyanin paralogs, hemocyanins have phylogenetic utility for most spider groups. This dissertation shows the importance of research on gene family evolution, the roles of gene families in adaptation, and the utility of gene families in phylogenetics. vi ii Table of Contents Introduction………………………………………………………………………………1 References…………………………………………………………………………4 Chapter 1 Abstract……………………………………………………………........................7 Introduction…………………………………………………………......................9 Materials and Methods…………………………………………………………...13 Results……………………………………………………………………………21 Discussion…………………………………………………………......................27 References………………………………………………………………………..39 Tables…………………………………………………………………….………46 Figures……………………………………………………………………..…......51 Chapter 2 Abstract…………………………………………………………………………..64 Introduction…………………………………………………………....................66 Materials and Methods………………………………………………...................68 Results and Discussion………………………………………………..................73 ix References………………………………………………………….................….81 Tables…………………………………………………………….........................85 Figures…………………………………………………………...................…….89 Chapter 3 Abstract………………………………………………………..................………97 Introduction……………………………………………………...................…….99 Materials and Methods…………………………………….................………....102 Results…………………………………………………..................……………105 Discussion…………………………………………………...................……….109 References………………………………………………….…...........................115 Tables…………………………………………………..................…………….118 Figures……………………………………………………...................…….…..125 Chapter 4 Abstract………………………………………………….....................………...137 Introduction……………………………………....................……….…….……139 Materials and Methods………………………………....................…..…….…..143 Results……………………………………………..................…….…….…..…152 Discussion…………………………………………….....................…….……..157 x References…………………………………………….…….....................……..166 Tables………………………………………………………......................…….173 Figures……………………………………………....................….…….………179 Conclusion………………………………….....................………….…………………190 xi List of Tables Table 1.1. Node support for spidroin phylogenetic analyses............................................46 Table 1.2. Continuous character data and alternative reconciliation based outgroups for maximum likelihood constrained spidroin tree................................................48 Table 2.1. Aliatypus species list with collection locality information..............................85 Table 2.2. Selection test results for the terminal repeat plus carboxy terminal encoding region from codeml analyses...........................................................................86 Table 2.3. TreeSAAP selection test results for the physicochemical property, alpha helical tendency, in the terminal repeat plus carboxy terminal encoding region...............................................................................................................87 Table 3.1. Bowtie2 read counts for Aphonopelma seemanni..........................................118 Table 3.2. Bowtie2 read counts for Poecilotheria regalis..............................................121 Table 4.1. Spider taxon sample for hemocyanins...........................................................173