Download Epigenetics

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