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Ralph G. Meyer - Bios * Heidelberg University (Ruprecht-Karls-Universitaet Heidelberg) Ask questions! Please send me an email, or stop by, or call me **anytime 9 am-6 pm, Mo-Fr** with your questions Ralph Meyer, Ph.D., BTB Rm. 211 [email protected] (435) 797 1774 Reproduction Genetics Epigenetics + = Environment There are great summer research projects available to all students! What does “Epigenetics” mean? • gene expression changes not caused by changes in DNA sequence • genes are “turned on” or “turned off” by chemical alterations to: - DNA (e.g.methylation) or RNA - histone modification (methylation, acetylation, etc.) • epigenetic changes can be heritable • if DNA (genome) is the hardware of a computer, epigenetics (epigenome) is like the “software”. • provides an additional layer of information on the genetic info. Spermatogenesis Rationale: • 15% of couples fertility problems • 50% of these: male factor – nonobstructive: 70% idiopathic infert. • reduced fertilization capacity of sperm • reduced embryonal survival/miscarriage • Chromatin integrity and proper nucleoprotein composition are prerequisites for normal sperm cell function. Incomplete chromatin condensation & persisting DNA strand breaks Compromised sperm nucleoprotein composition = increased ratio of nuclear histones to protamines (normal histone contents: 1-5% in mice, 5-15% in humans) Molecular mechanisms of sperm DNA remodeling not well understood Chromatin remodeling in spermiogenesis “Spermiogenesis”: postmeiotic phase of male germ cell development PAR (after Zhao et al., 2004 ) What does all this have to do with animal reproduction in the real world? Cutting to the chase: the PAR machinery is important for proper sperm histone to protamine exchange (Meyer-Ficca et al., 2011) So what? The “histone code” regulates gene expression Activating: e.g. H3K4me3 Silencing: e.g. H3K9me3, H3K27me3 Why are sperm histones relevant? • Sperm have no gene activity – are just packages • Not all histones lost: some remain after fertilization • Histones carry epigenetic modifications • Epigenetic marks on histones silence or activate genes! • Do sperm histones regulate embryo gene expression? • What happens if PARylation inhibited? PARP Inhibitors are important experimental drugs for numerous diseases • Ischemia-Reperfusion Injury • Intestine • Eye • Kidney • Skeletal Muscle • Myocardial Infarction • Stroke • Systemic Inflammation and Circulatory Shock • Cancer • Arthritis • Inflammatory Bowel Disease • Diabetes • Antiretroviral Therapy AG-014699 AZD2281 PARP inhibition causes poor sperm chromatin condensation and excessive histone retention in sperm PJ34 a b c e f Saline d (Meyer-Ficca et al., 2011b, 2014) Research Project I (funded UAES grant) Sperm chromatin composition and the fertility of domestic animals: • design of a simple antibody-based detection method, not unlike test-strip kits that are currently used for pregnancy tests • testing sperm quality in stallions as a model for other domestic species. Strategy: test for marker proteins identified so far and by an open proteomics approach. I am looking for students who would like to do summer research projects working with me on this project. Also looking for graduate students. Could sperm histones regulate early embryonic gene expression? Could sperm histones regulate early embryonic gene expression? Histones retained in sperm occupy promoter regions of genes relevant to early development of the embryo (Arpanahi A et al, Genome Res 2009, Hammoud SS et al, Nature 2009, Brykczynska, U. et al., Nat. Struct. Biol. 2010) Need to change amount and positions of histones associated with promoter regions in sperm to test if expression of genes during early embryonic development is actually changed. PARP inhibition in mice ideal model to test the hypothesis that sperm histones regulate embryonic gene expression Testing the hypothesisconsequences of changing histones in sperm for embryo gene expression? ♀ Wild-type ♀ Individual 2-cell embryos Which genes are active? ♂ Wild-type ♂ Parg(110)-/- ♂ PJ34 inj. ♂ Compare to identify genes that were regulated in embryo by histones in sperm Which genes have histones in sperm? Result(s): histones mediate gene regulation in the embryo Testis Sperm Progeny Spermiogenesis Zygote/1CE 2CE ? A A A ? B B B ? C C Chromatin remodeling PARP1 PARP2 TOP2B Protamines C Genome activation Chromatin remodeling ? (♂PN) Histones DE ♀ Histones Protamines Ihara et al., PLoS Genetics 2014 Research Project II (NIH project) Epigenetic inheritance through the male germ line • or: we are what Daddy ate…. molecular mechanisms of phenotypic variation and the ability to pass on acquired metabolic traits to progeny. (novel transgenic mouse models of humanized vitamin B3 metabolism, dietary deficiency). • Epigenetic modifications of sperm histones retained by altered PAR metabolism (genome-wide) • Roles of histone modifications in the regulation of gene expression in the preimplantation embryo (global) • DNA methylation analyses (genome-wide) Looking for students who would like to get involved with this research. Research Project III (Mirella Meyer-Ficca, PhD., NIH project 2) • Understand early epigenetic reprogramming in primordial germ cells (PGC) in utero- important for sperm development in the adult. • Impact of the environment – potential adverse effect on male fertility • Potential transgenerational inheritance • Poly(ADP-ribose) (PAR) metabolism: BER (DNA demethyl.) • PAR gauges environmental stress and VitB3 status • PAR: chromatin remodeling in epigenetic events Central hypothesis: Poly(ADP-ribose) metabolism is necessary for proper epigenetic pattern formation in the male germ line. Primordial germ cells are also reprogrammed with the help of PARP and PAR B6/OG2 (Oct4-GFP) PJ34/ABT-888* * showed efficacy in GR/PGC DNA demethylation remethylation E9.5 10.5 12.5 13.5 15.5 17.5 (~21d) Birth PGC Prospermatogonia 60 Days ♂ ♂ FACS Days pp 90 120 Histology Sperm GR Testes Is sperm quality in adults affected by adverse exposure of males prior to birth? Past /other projects Targeting poly(ADP-ribose) metabolism for development of a nonsurgical sterilant (Found Animals Foundation, D0809W20) Objective: Humanely stem feral companion animal overpopulation in a trap-and-release setting Approach: single-dose drug combination of cytotoxic anticancer drug with PARP inhibitor Motomasa Ihara, MD., PhD Julia Lonchar Jessica Bader Hope Douglas The activity of poly(ADP-ribose) polymerase-1 in response to equine intestinal ischemiareperfusion injury (ongoing) Objective: Evaluate PARP inhibitors as first-line drugs to prevent ischemia-reperfusion injury in horses with strangulating intestinal lesions (colic) Approach: measure PAR formation in horse tissues, test efficacy of PARP inhibitors in equine, in vitro and in vivo Hope Douglas Mirella Meyer-Ficca, PhD Samantha Hart, BVMS (Hons), MS, DACVS Ralph Meyer, PhD Louise Southwood, BVSc, PhD, DACVS, DACVECC Clinical Problem • Colic is an important cause of mortality in horses: • Annual cost $115.3 million • Overall fatality rate 11% • Strangulating lesions increase morbidity and mortality Clinical Problem Ischemia-Reperfusion and Inflammation Mucosal Damage Endotoxin Absorption Shock Death Ask questions! Please send me an email, or stop by, or call me **anytime 9 am-6 pm, Mo-Fr** with your questions Ralph Meyer, Ph.D., BTB Rm. 211 [email protected] (435) 797 1774