Hypomorphic maternal LSD1 in mice leads to long-term defects in survival and imprinting Öffentlichkeit
Curlee, Marcus (Fall 2020)
Published
Abstract
Oocytes contain proteins which make epigenetic modifications during the maternal-to-zygotic transition, which is the shift from expression of the maternal genome to expression of the zygotic genome. At fertilization, these maternally provided proteins are relied on for the reprogramming of each parental genome, a necessary step for transitioning from the high levels of differentiation in each gamete to a totipotent zygote. LSD1 is an H3K4 demethylase which is present in the female germline and is necessary for successful epigenetic reprogramming. The complete loss of maternal LSD1 is lethal in mice at the 1-2 cell stage, with transcription resembling that of the maternal genome. A partial loss of maternal LSD1 allows mice embryos to survive early development, but their improper reprogramming causes several negative health effects, such as perinatal lethality and defective genomic imprinting. This means that incomplete epigenetic reprogramming can lead to defects which manifest postnatally. In order to further study these long-term effects, our lab developed a new hypomorphic maternal Lsd1 allele with a mutation in its tower domain. This mutation primarily inhibits LSD1’s binding with CoREST, with only minimal impact on LSD1’s in vitro demethylase activity. Hypomorphic maternal loss of LSD1 increases perinatal lethality and leads to imprinting defects which are maintained throughout development. These results partially phenocopy those of our lab’s previous maternal LSD1 partial loss studies, suggesting that LSD1’s maternal activity may be CoREST-dependent and that this mouse mutant can serve as a more efficient tool for the in-depth study of phenotypes observed in mice with deficient maternal LSD1. Our results also provide another lens through which to view human mutations in epigenetic enzymes, due to the fact that some phenotypes may be a result of defects which act maternally.
Table of Contents
INTRODUCTION 1-12
Introduction to Epigenetics 1-3
H3K4me as an Active Mark 3-4
LSD1 Removes H3K4me1/2 and Can Erase Transcriptional Memory 4-5
LSD1 Function Depends on Complex Composition 5-6
Epigenetic Enzymes Can be Mutated in Humans 6-7
Epigenetic Reprogramming at Fertilization and the Maternal-to-Zygotic Transition 7-8
LSD1 and Epigenetic Reprogramming 8-9
Introduction to Genomic Imprinting 9-11
Outstanding Questions 11-12
MATERIALS AND METHODS 13-15
Mouse Husbandry and Genotyping 13
Bisulfite Analysis and Bisulfite-PCR Optimization 13-14
Immunofluorescence 14-15
Perinatal Lethality 15
RESULTS 16-20
Partial Maternal Loss of LSD1 Causes Long-Term Defects 16-17
The M448V Mutation Reduces Binding with CoREST 17-19
Hypomorphic Maternal LSD1 Causes Perinatal Lethality 19
Hypomorphic Maternal LSD1 Leads to Imprinting Defects 19-20
DISCUSSION 21-30
Partial Loss of Maternal LSD1 Causes Defects Postnatally 21
Maternal Epigenetic Reprogramming via LSD1 May Be CoREST-Dependent 21-24
Hypomorphic Maternal LSD1 Leads to Imprinting Defects 24-26
A Model for LSD1 and Epigenetic Reprogramming at Fertilization 27-28
Future Directions 28-30
FIGURES 31-41
Figure 1. Partial loss of maternal LSD1 leads to long-term defects. 31-32
Figure 2. Hypomorphic LSD1 mutation and genetic crosses. 32-33
Figure 3. Increased perinatal lethality in Lsd1M448V progeny. 33
Figure 4. Zac1 imprinting defects in Lsd1M448V progeny heart tissue. 34
Figure 5. Zac1 imprinting defects in Lsd1M448V progeny brain tissue. 35
Figure 6. Zac1 imprinting defects in Lsd1M448V progeny liver tissue. 36
Figure 7. CoREST is broadly expressed in the nucleus at three different stages of oogenesis. 37
Figure 8. Differentially expressed genes in spr-5; met-2 progeny are similarly misregulated in met-2; spr-1 progeny, leading to progressive sterility. 38
Figure 9. Schematic of Negative and Positive Snrpn GPF Imprinting Reporter Mice. 39-40
Figure 10. Hypomorphic maternal LSD1 in mammals leads to the incomplete removal of H3K4me transcriptional memory. 40-41
REFERENCES 42-45
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