University of Pennsylvania ScholarlyCommons Publicly Accessible Penn Dissertations 2015 Transgenerational Epigenetic Mechanisms of Stress Axis Programming Alison Buch Rodgers University of Pennsylvania, [email protected] Follow this and additional works at: https://repository.upenn.edu/edissertations Part of the Neuroscience and Neurobiology Commons Recommended Citation Rodgers, Alison Buch, "Transgenerational Epigenetic Mechanisms of Stress Axis Programming" (2015). Publicly Accessible Penn Dissertations. 1979. https://repository.upenn.edu/edissertations/1979 This paper is posted at ScholarlyCommons. https://repository.upenn.edu/edissertations/1979 For more information, please contact [email protected]. Transgenerational Epigenetic Mechanisms of Stress Axis Programming Abstract Altered stress reactivity is a predominant feature of sex-biased neuropsychiatric diseases, with hyperreactivity and hyporeactivity of the hypothalamic-pituitary-adrenal (HPA) stress reported in affective, neurotic, and psychotic disorders. Stress dysregulation may precede symptom onset, and has been characterized as a vulnerability that predisposes males and females to disease. Environmental factors, particularly prior stress history, contribute to the developmental programming of stress reactivity in exposed individuals and their offspring. Epigenetic marks in the brain are uniquely positioned to act at the interface of stress and lasting changes in behavior and physiology, and epigenetic signatures in germ cells are similarly poised to communicate experience-dependent information across generations. In this dissertation, mouse models are used to examine the consequences of chronic stress on later stress reactivity, and to define the molecular mechanisms involved in transgenerational and brain programming. Results of the first set of studies revealed that paternal exposure to chronic variable stress elicits a reduced HPA axis response in offspring, and that nine microRNAs (miRs) increased in the sperm of stressed sires mediate this transmission, as zygote microinjection of the nine miRs remarkably recapitulated of the paternal stress phenotype. Broad repression of the hypothalamic transcriptome following paternal stress or miR-microinjection suggested that epigenetic reprogramming underlies altered offspring stress reactivity. Further, targeted degradation of maternal mRNA transcripts in miR- microinjected zygotes provided the first step in the complex egulatr ory cascade by which sperm miRs impact the adult offspring brain. The final study offers evidence of the direct reprogramming of neurodevelopment by chronic stress, highlighting the disruption of sex differences in the prefrontal cortex by peripubertal stress as particularly relevant to the onset of sex-biased neuropsychiatric diseases. Together, research presented in this dissertation support that epigenetic mechanisms mediate the transgenerational transmission of chronic stress and allow lifetime adversity to program later stress dysregulation. Degree Type Dissertation Degree Name Doctor of Philosophy (PhD) Graduate Group Neuroscience First Advisor Tracy L. Bale Keywords environment, epigenetic, hypothalamus, stress, transgenerational Subject Categories Neuroscience and Neurobiology This dissertation is available at ScholarlyCommons: https://repository.upenn.edu/edissertations/1979 TRANSGENERATIONAL EPIGENETIC MECHANISMS OF STRESS AXIS PROGRAMMING Alison Buch Rodgers A DISSERTATION in Neuroscience Presented to the Faculties of the University of Pennsylvania in Partial Fulfillment of the Requirements for the Degree of Doctor of Philosophy 2015 Supervisor of Dissertation _____________________ Tracy L. Bale, PhD Professor of Neuroscience Graduate Group Chairperson _____________________ Joshua I. Gold, PhD Professor of Neuroscience Dissertation Committee Zhaolon (Joe) Zhou, PhD, Assistant Professor of Neuroscience Max B. Kelz, MD PhD, Associate Professor of Anesthesiology Edward S. Brodkin, MD, Associate Professor of Psychiatry R. Christopher Pierce, PhD, Professor of Neuroscience ACKNOWLEDGEMENT When I welcomed the new class of biomedical graduate students to Penn last summer, I charged them to remember, as my PI has said, “grad school isn’t meant to be easy.” Perhaps those weren’t the most encouraging words, but they were honest. Completing this thesis was one of the most trying, mentally and emotionally exhausting endeavors that I have undertaken. But I was never alone in my struggles or my successes over the last five years, and the individuals who have seen me through graduate school through deserve both acknowledgement and praise. My mentor, Tracy Bale, spent significant energies developing my scientific creativity and independence, challenging me to grow from a student into a neuroscientist. With high expectations that spoke to her confidence in me, she fostered my confidence in my own science. Her enthusiasm for our projects was infectious, helping to free me from the mud of skepticism and doubt. I am most thankful for her decision to swap sticks for carrots, for her encouragement to enjoy writing as a creative outlet, and for her incredible support as I entered motherhood and with it, a new chapter of work-life balance. Katie Morrison has been my co-star, role model, and confidant who shared her skills and expertise as a lab ninja, and her sympathy and insight as a friend, so generously. She’s seen me at my most raw—frustratingly frustrated and standing on tables—but always encouraged me to stick with it and helped me do just that. I am so grateful for my new best friend, our train rides, and our story. I also thank the sum of all the current and former Bale Lab-ites, who have decorated my graduate career with conversations and camaraderie. Notably, Chris Morgan collaborated on so many projects with me, marking the many time we spent ii together by the annual the vet student courtyard games. Greg Dunn pioneered the miR injection experiments and has shaped my definition of success. Alexis Gerber Howerton was responsible for my joining the lab in the first place, and Chris Howerton was my standing consult on all things scientific and statistical. Jen Chan adopted my projects and continues to nurture them with her careful thought and attention. My thesis committee, Joe Zhou, Max Kelz, Ted Brodkin, and Chris Pierce, guided my research with helpful and constructive feedback at our meetings. Dan Beiting taught and advised big data statistics with unparalleled patience. The Neuroscience Graduate Group and its chair, Josh Gold, established a unique environment that engages students in research and extracurricular activities, particularly through their support of GLIA, with administrators Jane Hoshi and Christine Clay keeping us all together. My friends and fellow grad students offered alternating laughs and consolation, as required, especially Nicole Yohn, who gave me more credit than I deserve and whose organizational skills complemented my own; we made a good team. My family has always reminded me that what I do is unusual and interesting, especially Grandpa Fleming, who made me feel nothing less than extraordinary. My daughter, Avery, brought so much joy to my last year and has helped me overcome perfectionism more in a few short months than others have in years. And finally, my husband, Dan, has kept me grounded in a way no one else can. He just keeps loving me. iii ABSTRACT TRANSGENERATIONAL EPIGENETIC MECHANISMS OF STRESS AXIS PROGRAMMING Ali B. Rodgers Tracy L. Bale Altered stress reactivity is a predominant feature of sex-biased neuropsychiatric diseases, with hyperreactivity and hyporeactivity of the hypothalamic-pituitary-adrenal (HPA) stress reported in affective, neurotic, and psychotic disorders. Stress dysregulation may precede symptom onset, and has been characterized as a vulnerability that predisposes males and females to disease. Environmental factors, particularly prior stress history, contribute to the developmental programming of stress reactivity in exposed individuals and their offspring. Epigenetic marks in the brain are uniquely positioned to act at the interface of stress and lasting changes in behavior and physiology, and epigenetic signatures in germ cells are similarly poised to communicate experience-dependent information across generations. In this dissertation, mouse models are used to examine the consequences of chronic stress on later stress reactivity, and to define the molecular mechanisms involved in transgenerational and brain programming. Results of the first set of studies revealed that paternal exposure to chronic variable stress elicits a reduced HPA axis response in offspring, and that nine microRNAs (miRs) increased in the sperm of stressed sires mediate this transmission, as zygote microinjection of the nine miRs remarkably recapitulated of the paternal stress phenotype. Broad repression of the hypothalamic transcriptome following paternal stress or miR-microinjection suggested that epigenetic reprogramming underlies altered offspring stress reactivity. Further, iv targeted degradation of maternal mRNA transcripts in miR-microinjected zygotes provided the first step in the complex regulatory cascade by which sperm miRs impact the adult offspring brain. The final study offers evidence of the direct reprogramming of neurodevelopment by chronic stress, highlighting the disruption of sex differences in the prefrontal cortex by peripubertal stress as particularly relevant to the onset of sex-biased neuropsychiatric diseases. Together,