MRG15 is required for pre-mRNA splicing and spermatogenesis

Abstract

Splicing can be epigenetically regulated and involved in cellular differentiation in somatic cells, but the interplay of epigenetic factors and the splicing machinery during spermatogenesis remains unclear. To study these interactions in vivo, we generated a germline deletion of MORF-related gene on chromosome 15 (MRG15), a multifunctional chromatin organizer that binds to methylated histone H3 lysine 36 (H3K36) in introns of transcriptionally active genes and has been implicated in regulation of histone acetylation, homology-directed DNA repair, and alternative splicing in somatic cells. Conditional KO (cKO) males lacking MRG15 in the germline are sterile secondary to spermatogenic arrest at the round spermatid stage. There were no significant alterations in meiotic division and histone acetylation. Specific mRNA sequences disappeared from 66 germ cell-expressed genes in the absence of MRG15, and specific intronic sequences were retained in mRNAs of 4 genes in the MRG15 cKO testes. In particular, introns were retained in mRNAs encoding the transition proteins that replace histones during sperm chromatin condensation. In round spermatids, MRG15 colocalizes with splicing factors PTBP1 and PTBP2 at H3K36me3 sites between the exons and single intron of transition nuclear protein 2 (Tnp2). Thus, our results reveal that MRG15 is essential for premRNA splicing during spermatogenesis and that epigenetic regulation of pre-mRNA splicing by histone modification could be useful to understand not only spermatogenesis but also, epigenetic disorders underlying male infertile patients.infertility | fertility defects | splicing defects | epigenetics | spermiogenesis S permatogenesis is a complex process involving several biological events and dramatic changes of chromatin structure. Male germ cells undergo stem cell self-renewal, mitotic divisions in spermatogonial proliferation, genomic rearrangement by meiotic homologous recombination at the spermatocyte stage, and morphological changes of round spermatids into elongated spermatids to form mature spermatozoa (1-3). During spermiogenesis, nucleosomal histone proteins are replaced with transition nuclear proteins (TNPs) and subsequently, protamines, the major nucleosomal proteins in spermatozoa. Moreover, epigenetic modifications, such as histone methylation, dramatically change throughout spermatogenesis (3, 4).Pre-mRNA splicing generates protein diversity and is involved in the regulation of cellular differentiation (5, 6). Recent advances have shown that histone modifications regulate alternative splicing through recruitment of splicing regulators via chromatin binding proteins, such as MORF-related gene on chromosome 15 (MRG15) (7). Histone H3 lysine 36 (H3K36) is methylated proximal to tissuespecific splicing regions (8-12). MRG15 specifically recognizes the methylated H3K36 and recruits polypyrimidine tract binding protein (PTB) at intronic splicing silencer elements near an exon to suppress exon insertions into mRNA (7). MRG15 is also a component of histone acetyltransfe...

Physical and functional interaction between SET1/COMPASS complex component CFP-1 and a Sin3 HDAC complex

“…C. elegans HDA-1 is one of three class I histone deacetylases (HDACs) in C. elegans and a component of several other chromatin complexes, as in other species (Kelly and Cowley, 2013;Passannante et al, 2010). MRG-1, the C. elegans counterpart of the chromo-domain (CD) protein Eaf3/MRG15, is also found in additional chromatin complexes (Bleuyard et al, 2017;Chen et al, 2010;Huang et al, 2017;Iwamori et al, 2016;Smith et al, 2013), and ATHP-1 (AT Hook plus PHD finger transcription factor), a counterpart of Rco1/Pf1, contains an AT Hook domain that is not found in either Rco1 or Pf1 ( Figure S2D).…”

PTBP2-dependent alternative splicing regulates protein transport and mitochondria morphology in post-meiotic germ cells.

“…Strikingly, two spermatogenic cell types (spermatocytes and spermatids) are responsible for most of the AS isoform complexity found in the testis (Soumillon et al, 2013). The functions and regulation of AS in these and other spermatogenic cells is just beginning to be understood (O'Bryan et al, 2013;Schmid et al, 2013;Iwamori et al, 2016;Hannigan et al, 2017). Using available RNA-sequencing (RNA-seq) data from spermatogonia, spermatocytes, round spermatids, and spermatozoa (Soumillon et al, 2013), we previously identified 1030 stagespecific changes in AS mRNA isoforms as germ cells move through the spermatogenic pathway (affecting expression of 912 genes) (Hannigan et al, 2017).…”

ASH1L REGULATES THE STRUCTURAL DEVELOPMENT OF NEURONAL CIRCUITRY BY MODULATING BDNF/TrkB SIGNALING IN HUMAN NEURONS

“…Histone H3K36 methylation regulates pre-mRNA splicing in S. cerevisiae (Sorenson et al, 2016), and MRG15, which forms a complex with ASH1L (Lee et al, 2019), also regulates pre-mRNA splicing (Iwamori et al, 2016). Since ASH1L dimethylates H3K36 (Miyazaki et al, 2013;Zhu et al, 2016a) and forms a complex with MRG15, we asked if ASH1L could regulate the alternative splicing of NTRK2 (TrkB) mRNA, which has seven distinct splicing isoforms.…”