Rescue of placental phenotype in a mechanistic model of Beckwith-Wiedemann syndrome

Background  

Several imprinted genes have been implicated in the process of placentation. The distal region of mouse chromosome 7 (Chr 7) contains at least ten imprinted genes, several of which are expressed from the maternal homologue in the placenta. The corresponding paternal alleles of these genes are silenced in cis by an incompletely understood mechanism involving the formation of a repressive nuclear compartment mediated by the long non-coding RNA Kcnq1ot1 initiated from imprinting centre 2 (IC2). However, it is unknown whether some maternally expressed genes are silenced on the paternal homologue via a Kcnq1ot1-independent mechanism. We have previously reported that maternal inheritance of a large truncation of Chr7 encompassing the entire IC2-regulated domain (DelTel7 allele) leads to embryonic lethality at mid-gestation accompanied by severe placental abnormalities. Kcnq1ot1 expression can be abolished on the paternal chromosome by deleting IC2 (IC2KO allele). When the IC2KO mutation is paternally inherited, epigenetic silencing is lost in the region and the DelTel7 lethality is rescued in compound heterozygotes, leading to viable DelTel7/IC2KO mice.     

Antisense noncoding RNA promoter regulates the timing of de novo methylation of an imprinting control region

Epigenetic marks at cis acting imprinting control regions (ICRs) regulate parent of origin-specific expression of multiple genes in imprinted gene clusters. Epigenetic marks are acquired during gametogenesis and maintained faithfully thereafter. However, the mechanism by which differential epigenetic marks are established and maintained at ICRs is currently unclear. By using Kcnq1 ICR as a model system, we have investigated the functional role of genetic signatures in the acquisition and maintenance of epigenetic marks. Kcnq1 ICR is methylated on the maternal chromosome but remains unmethylated on the paternal chromosome. Here, we show that a paternal allele of Kcnq1 ICR lacking the Kcnq1ot1 promoter remains unmethylated during spermatogenesis; however, it becomes methylated specifically during pre-implantation development. Analysis of the chromatin structure at the paternal ICR in spermatogenic cells and in E13.5 embryonic tissues revealed that the ICRs of both wild type and mutant mice are enriched with H3K4me2 in spermatiogenic cells of the testicular compartment, but the mutant ICR lost H3K4me2 specifically in epididymal sperm and an increase in repressive marks was observed in embryonic tissues. Interestingly, we also detected a decrease in nucleosomal histone levels at the mutant ICR in comparison to the wild-type ICR in epididymal sperm. Taken together, these observations suggest that the Kcnq1ot1 promoter plays a critical role in establishing an epigenetic memory in the male germline by ensuring that the paternal allele remains in an unmethylated state during pre-implantation development.     

An antisense RNA regulates the bidirectional silencing property of the Kcnq1 imprinting control region

The Kcnq1 imprinting control region (ICR) located in intron 10 of the Kcnq1 gene is unmethylated on the paternal chromosome and methylated on the maternal chromosome and has been implicated in the manifestation of parent-of-origin-specific expression of six neighboring genes. The unmethylated Kcnq1 ICR harbors bidirectional silencer activity and drives expression of an antisense RNA, Kcnq1ot1, which overlaps the Kcnq1 coding region. To elucidate whether the Kcnq1ot1 RNA plays a role in the bidirectional silencing activity of the Kcnq1 ICR, we have characterized factor binding sites by genomic footprinting and tested the functional consequence of various deletions of these binding sites in an episome-based system. Deletion of the elements necessary for Kcnq1ot1 promoter function resulted in the loss of silencing activity. Furthermore, interruption of Kcnq1ot1 RNA production by the insertion of a polyadenylation sequence downstream of the promoter also caused a loss of both silencing activity and methylation spreading. Thus, the antisense RNA plays a key role in the silencing function of the ICR. Double-stranded RNA (dsRNA)-mediated RNA interference is unlikely to be involved, as the ICR is active irrespective of the simultaneous production of dsRNA from the genes it silences.     

The C. elegans Polycomb gene SOP-2 encodes an RNA binding protein

Epigenetic silencing of Hox cluster genes by Polycomb group (PcG) proteins is thought to involve the formation of a stably inherited repressive chromatin structure. Here we show that the C. elegans-specific PcG protein SOP-2 directly binds to RNA through three nonoverlapping regions, each of which is essential for its localization to characteristic nuclear bodies and for its in vivo function in the repression of Hox genes. Functional studies indicate that the RNA involved in SOP-2 binding is distinct from either siRNA or microRNA. Remarkably, the vertebrate PcG protein Rae28, which is functionally and structurally related to SOP-2, also binds to RNA through an FCS finger domain. Substitution of the Rae28 FCS finger for the essential RNA binding region of SOP-2 partially restores localization to nuclear bodies. These observations suggest that direct binding to RNA is an evolutionarily conserved and potentially important property of PcG proteins.     

Polycomb group蛋白复合体

Polycomb group (PcG) 蛋白是一组通过染色质修饰调控靶基因的转录抑制子, 从生化和功能上它可以分成两个主要的核心蛋白复合体PRC1(Polycomb repressive complex 1)和PRC2(Polycomb repressive complex 2)。研究发现PcG蛋白不仅控制个体正确的发育模式, 而且与细胞的增殖、分化和肿瘤发生有关。文章就PcG蛋白的组成、作用机制及功能进行综述, 并对PcG未来的研究方向进行展望。