University of Pennsylvania ScholarlyCommons Publicly Accessible Penn Dissertations 2017 Evolutionary Instability Of Genomic Mutation Rate In Rapidly Adapting Asexual Mutator Escherichia Coli Populations Mitra Eghbal University of Pennsylvania, [email protected] Follow this and additional works at: https://repository.upenn.edu/edissertations Part of the Biology Commons, Genetics Commons, and the Microbiology Commons Recommended Citation Eghbal, Mitra, "Evolutionary Instability Of Genomic Mutation Rate In Rapidly Adapting Asexual Mutator Escherichia Coli Populations" (2017). Publicly Accessible Penn Dissertations. 2850. https://repository.upenn.edu/edissertations/2850 This paper is posted at ScholarlyCommons. https://repository.upenn.edu/edissertations/2850 For more information, please contact [email protected]. Evolutionary Instability Of Genomic Mutation Rate In Rapidly Adapting Asexual Mutator Escherichia Coli Populations Abstract Alleles conferring higher mutation rates (mutators) can fix in asexual populations through a process called ‘mutator hitchhiking’. Theory predicts that repeated mutator hitchhiking can occur in an adapting asexual mutator population. I tested this prediction in two settings: a mutL- population under lethal selection and a mutS- population under soft selection. In both experiments, the starting mutation rate was 100-fold higher than wild-type. In the lethal selection experiment, two replicate populations were exposed to a sequence of three different antibiotics. In both replicates, all survivors sampled after the final antibiotic exposure had undergone further genomic mutation rate increases. Whole-genome and Sanger sequencing revealed that an identical spontaneous 1-bp insertion in mutT (a known mutator gene) rose to probable fixation in both populations. Complementation tests demonstrated that the mutT- defect was responsible for the increased mutation rates. In the soft selection experiment, 30 isogenic populations were propagated in limited glucose media. After 900 generations, five clones were isolated from each population. Relative to the ancestor, 9% of the clones had increased mutation rates and 68% had unchanged mutation rates; surprisingly, 23% had decreased mutation rates. Most populations (21 of 30) had at least one clone whose mutation rate either increased or decreased. One population exhibited apparent fixation for a mutator and one other population exhibited apparent fixation for an antimutator. Some of the sequenced clones with altered mutation rates had mutations in known (anti)mutator loci. I conclude that the mutators likely arose by hitchhiking and that the antimutators likely confer pleiotropic direct fitness benefits. Competitions between thev e olved clones and the ancestor demonstrated that all clones and populations had increased in fitness since generation 0. No relationship was detected, however, between mutation rate and relative fitness. These experiments provide evidence supporting the prediction of repeated mutator hitchhiking. More broadly, the work described in this dissertation reveals multiple ways in which the mutation rates of asexual populations may be evolutionarily unstable, with potential implications for evolving asexual systems, including infectious agents and cancer. Degree Type Dissertation Degree Name Doctor of Philosophy (PhD) Graduate Group Biology First Advisor Paul Sniegowski Subject Categories Biology | Genetics | Microbiology This dissertation is available at ScholarlyCommons: https://repository.upenn.edu/edissertations/2850 EVOLUTIONARY INSTABILITY OF GENOMIC MUTATION RATE IN RAPIDLY ADAPTING ASEXUAL MUTATOR ESCHERICHIA COLI POPULATIONS Mitra M. Eghbal A DISSERTATION in Biology Presented to the Faculties of the University of Pennsylvania in Partial Fulfillment of the Requirements for the Degree of Doctor of Philosophy 2017 Supervisor of Dissertation _______________________________________ Paul Sniegowski, Professor, Department of Biology, University of Pennsylvania Graduate Group Chairperson _______________________________________ Michael Lampson, Associate Professor, Department of Biology, University of Pennsylvania Dissertation Committee: Timothy Linksvayer, Associate Professor, Department of Biology, University of Pennsylvania Mechthild Pohlschröder, Professor, Department of Biology, University of Pennsylvania Paul Schmidt, Professor, Department of Biology, University of Pennsylvania Sarah Tishkoff, Professor, Department of Biology, University of Pennsylvania EVOLUTIONARY INSTABILITY OF GENOMIC MUTATION RATE IN RAPIDLY ADAPTING ASEXUAL MUTATOR ESCHERICHIA COLI POPULATIONS COPYRIGHT 2017 Mitra M. Eghbal This thesis is dedicated to whoever reads it, be they compelled by curiosity or by coercion. Reading my thesis entails a dedication of your time. It is only right that I reciprocate that dedication. iii ACKNOWLEDGMENTS In a certain light, mutations could be viewed as a form of biological luck (or lack thereof). Studying mutation rate evolution for several years really makes you appreciate the rarity of good luck. Several years is also enough time for me to realize my own good luck. In my time as a graduate student, I’ve been lucky to be in contact with a number of people who influenced my life for the better. Thank you to the members of my lab. Although, at almost any given point in time, we were a small lab, there was a changing cast of many delightful characters over the years. I’ll start off with the grad students. My senior graduate student lab mates, Eugene and Chris, were great resources (for science and for mischief), especially when I first arrived in the lab. Years after his graduation, Eugene still offers me excellent advice from afar. In the latter years of my schooling, I was especially grateful for the company of Tanya and Ben. Their perpetual curiosity facilitated the exploration of a number of scientific questions together. I also appreciated the fact that we had complementary areas of scientific expertise and were able to assist each other as needed. Tanya and Ben, may you dream evermore of many dragons and many robots (respectively), who will welcome you as one of their own and extend the respect and affection that you both deserve. iv Our lab was lucky to be visited by a master’s student (Damien) and rotation students (Alexandra and Ozan) whose intellectual enthusiasm and warm-heartedness were greatly appreciated, and with whom I continued to have many productive and enjoyable chats about science (and science fiction) for years even after they departed our lab. I am honored that our lab was visited by a philosophy PhD student, Emily, who was eager to learn about our work and also eager to offer us a glimpse of her academic world. Exposure to the philosophy of science community has helped me see many more complexities behind that hazy beast that we call the ‘scientific process’. Our lab was fortunate to have a representative philosopher as approachable, articulate, and enjoyable as Emily. Although we only had one postdoc in our lab in the time I’ve been here, she certainly had an impact on my research. Kathleen was very helpful in getting me oriented in the wild frontiers of genomic sequencing. The current state of genomics in our lab wouldn’t be the same without her. I am grateful to Jude for his work on the hard selection project for over a year and for keeping my mind fresh with his passion for science. I can hardly believe that he’s already heading to college this fall, but I know that Penn is lucky to have this bright and hard- working fellow. I am also grateful to Holly and Clement for their diligent work on the soft selection fluctuation assays. Spending several weeks of a summer doing labwork is a sacrifice for any high schooler, yet it was a sacrifice they made very good-naturedly. v In the past year, I have had the fortune of working with several other talented and industrious ‘apprentices’ (as I like to think of them): Arlene, Meredith, and Breanna. Performing hundreds of fitness assays is no quick task, and if they hadn’t helped out, well, I don’t know where I’d be. Probably out of my mind. My lab mentees have given me great cause for optimism with regard to the future of science, and I am proud of them all. Thank you also to Kate, Matt, Angela, Dave, Dan, Ankur, Brooks, and Emilie for being a pleasure to work alongside and for being such good representatives of the next generation of science. I am deeply grateful to my committee, both as a group and individually: Mecky, Paul, Tim, and Sarah. I really appreciated their thought-provoking questions and their huge range of expertise. Thank you for the many hours that you spent listening to me talk about my research and offering your advice. My thesis (and, in fact, my career) has definitely benefited from your help. Many thanks to Phil, especially for his help on the fluctuation assay component of my thesis. His excellent advice from his perspective as a theorist was truly appreciated by this experimentalist, and his unflappably mellow, friendly, and unpretentious demeanor was a welcome counterweight to the stresses of graduate research. vi I want to thank the Kohli lab (especially Rahul, Matt, and Charlie) and the other members of the bacterial mutagenesis journal club. They were instrumental in helping me think more broadly about all facets of mutagenesis, which prepared me for the next step of my scientific career. I want to thank Sam from the Goulian lab for kindly sharing her impressive knowledge of molecular microbiology techniques. Thanks to