ABSTRACT
In mammals, chromatin organization undergoes drastic reorganization during oocyte development. However, the dynamics of three-dimensional chromatin structure in this process is poorly characterized. Using low-input Hi-C (genome-wide chromatin conformation capture), we found that a unique chromatin organization gradually appears during mouse oocyte growth. Oocytes at late stages show self-interacting, cohesin-independent compartmental domains marked by H3K27me3, therefore termed Polycomb-associating domains (PADs). PADs and inter-PAD (iPAD) regions form compartment-like structures with strong inter-domain interactions among nearby PADs. PADs disassemble upon meiotic resumption from diplotene arrest but briefly reappear on the maternal genome after fertilization. Upon maternal depletion of Eed, PADs are largely intact in oocytes, but their reestablishment after fertilization is compromised. By contrast, depletion of Polycomb repressive complex 1 (PRC1) proteins attenuates PADs in oocytes, which is associated with substantial gene de-repression in PADs. These data reveal a critical role of Polycomb in regulating chromatin architecture during mammalian oocyte growth and early development.
Subject(s)
Chromatin/chemistry , Oocytes/growth & development , Oogenesis/genetics , Polycomb-Group Proteins/physiology , Animals , Blastocyst/chemistry , Cell Cycle Proteins/physiology , Chromosomal Proteins, Non-Histone/physiology , Embryo, Mammalian/chemistry , Gene Silencing , Histone Code , Mice , Oocytes/chemistry , Transcription, Genetic , CohesinsABSTRACT
Abstract Dynamic DNA methylation is a prerequisite for many developmental processes and maintenance of cellular integrity. In mammals however, mechanisms of active DNA demethylation have for long been elusive. The discovery of the ten-eleven translocation (Tet) family of enzymes that oxidize 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC), 5-formylcytosine (5fC) or 5-carboxylcytosine (5caC) provided new means by which DNA methylation could actively be reversed. This review focuses on the possible mechanisms of DNA demethylation via Tet proteins and their metabolites 5hmC, 5fC and 5caC. Additionally, it discusses the roles of the three Tet protein family members Tet1, Tet2 and Tet3 as developmental regulators, probably in part independent of their enzymatic activity. By contrast, recent evidence suggests a function of 5hmC as an epigenetic mark on its own, going beyond the expectation of only acting as an intermediate in an active DNA demethylation pathway.
Subject(s)
Cytosine/analogs & derivatives , DNA Methylation , DNA-Binding Proteins/metabolism , Embryonic Stem Cells/metabolism , Epigenesis, Genetic , Gene Expression Regulation, Developmental , Proto-Oncogene Proteins/metabolism , 5-Methylcytosine/analogs & derivatives , Animals , Chromatin/genetics , Chromatin/metabolism , CpG Islands/genetics , Cytosine/analysis , Cytosine/metabolism , Gene Expression Regulation, Developmental/genetics , Genome/genetics , Mice , Polycomb-Group Proteins , Promoter Regions, Genetic/genetics , Repressor Proteins/metabolism , Sin3 Histone Deacetylase and Corepressor Complex , Transcription, Genetic/geneticsABSTRACT
The t complex responder (Tcr) encoded by the mouse t haplotype is able to cause phenotypic differences between t and + sperm derived from t/+ males, leading to non-Mendelian inheritance. This capability of Tcr contradicts the concept of phenotypic equivalence proposed for sperm cells, which develop in a syncytium and actively share gene products. By analyzing a Tcr minigene in hemizygous transgenic mice, we show that Tcr gene products are post-meiotically expressed and are retained in the haploid sperm cells. The wild-type allele of Tcr, sperm motility kinase-1 (Smok1), behaves in the same manner, suggesting that Tcr/Smok reveal a common mechanism prone to evolve non-Mendelian inheritance in mammals.
Subject(s)
Gene Expression Regulation , Inheritance Patterns/genetics , Protein Kinases/genetics , Protein Kinases/metabolism , Spermatids/metabolism , Animals , Female , Humans , Male , Mice , Mice, Inbred C57BL , Mice, Transgenic , Molecular Sequence Data , Phenotype , Spermatids/ultrastructureABSTRACT
Transmission ratio distortion (TRD), the preferential inheritance of the t haplotype from t/+ males, is caused by the cooperative effect of four t-complex distorters (Tcd1-4) and the single t-complex responder (Tcr) on sperm motility. Here we show that Fgd2, encoding a Rho guanine nucleotide exchange factor, maps to the Tcd2 region. The t allele of Fgd2 is overexpressed in testis compared with wild type. A loss-of-function allele of Fgd2 generated by gene targeting reduces the transmission ratio of the t haplotype t(h49), directly demonstrating the role of Fgd2 as Distorter. Fgd2 identifies a second Rho G protein signaling pathway promoting TRD.
Subject(s)
Guanine Nucleotide Exchange Factors/metabolism , Microtubule-Associated Proteins/metabolism , Nuclear Proteins/metabolism , Signal Transduction , Spermatozoa/metabolism , Alleles , Animals , Crosses, Genetic , Female , Gene Targeting , Guanine Nucleotide Exchange Factors/genetics , Haplotypes , Male , Mice , Models, Genetic , Sperm Motility , Testis/metabolism , t-Complex Genome RegionABSTRACT
The recently described protocadherin gene clusters encode cadherin-related proteins, which are highly expressed in the vertebrate nervous system. Here, we report biochemical studies addressing proteolytic processing of gamma-protocadherins. These type-I transmembrane proteins are cleaved by a metalloproteinase in vivo, generating a soluble extracellular fragment and a carboxyl-terminal fragment associated with the cellular membrane. In addition, we show that the carboxyl-terminal fragment is a substrate for further cleavage mediated by presenilin. Consequently, accumulation of the fragment is found when gamma-secretase is inactivated either by the specific presenilin-inhibitor L685,458 or in double mutant murine embryonic fibroblasts lacking both presenilin genes. The gamma-secretase-generated carboxyl-terminal fragment is largely unstable but accumulates when proteasomal degradation is inhibited. Interestingly, the proteolytic fragment generated by gamma-secretase can localize to the nucleus. This is the first report providing experimental evidence for a cell surface receptor signaling function of protocadherins regulated by proteolytic events.