Polycomb group (PcG) proteins prevent the assembly of abnormal synaptonemal complex structures during meiosis

Copyright © 2022 the Author(s). Published by PNAS. This article is distributed under Creative Commons Attribution-NonCommercial-NoDerivatives License 4.0 (CC BY-NC-ND).The synaptonemal complex (SC) is a proteinaceous scaffold that is assembled between paired homologous chromosomes during the onset of meiosis. Timely expression of SC coding genes is essential for SC assembly and successful meiosis. However, SC components have an intrinsic tendency to self-organize into abnormal repetitive structures, which are not assembled between the paired homologs and whose formation is potentially deleterious for meiosis and gametogenesis. This creates an interesting conundrum, where SC genes need to be robustly expressed during meiosis, but their expression must be carefully regulated to prevent the formation of anomalous SC structures. In this manuscript, we show that the Polycomb group protein Sfmbt, the Drosophila ortholog of human MBTD1 and L3MBTL2, is required to avoid excessive expression of SC genes during prophase I. Although SC assembly is normal after Sfmbt depletion, SC disassembly is abnormal with the formation of multiple synaptonemal complexes (polycomplexes) within the oocyte. Overexpression of the SC gene corona and depletion of other Polycomb group proteins are similarly associated with polycomplex formation during SC disassembly. These polycomplexes are highly dynamic and have a well-defined periodic structure. Further confirming the importance of Sfmbt, germ line depletion of this protein is associated with significant metaphase I defects and a reduction in female fertility. Since transcription of SC genes mostly occurs during early prophase I, our results suggest a role of Sfmbt and other Polycomb group proteins in downregulating the expression of these and other early prophase I genes during later stages of meiosis.R.G.M. is supported by Portuguese national funding through Fundação para a Ciência e a Tecnologia (FCT grant refs. PTDC/BIA-BID/28441/2017 and PTDC/BIA-BID/1606/2020). B.M. and R.D.S. are both supported by Portuguese national funding through Fundação para a Ciência e a Tecnologia, respectively, PD/BD/128342/2017 (within the scope of the ProRegeM PhD program; PD/00117/2012, CRM:0027030) and DL 57/2016/CP1361/CT0019. The Light Microscopy Unit of ABC-RI was partially supported by Portuguese national funding (FCT: PPBI-POCI-01-0145-FEDER-022122). This work was developed with the support of the research infrastructure Congento (project LISBOA-01-0145-FEDER-022170). The Transgenic RNAi Project (TRiP) collection at Harvard Medical School was supported by NIH/NIGMS R01-GM084947.info:eu-repo/semantics/publishedVersio

Altered Dosage and Mislocalization of Histone H3 and Cse4p Lead to Chromosome Loss in Saccharomyces cerevisiae

Cse4p is an essential histone H3 variant in Saccharomyces cerevisiae that defines centromere identity and is required for proper segregation of chromosomes. In this study, we investigated phenotypic consequences of Cse4p mislocalization and increased dosage of histone H3 and Cse4p, and established a direct link between histone stoichiometry, mislocalization of Cse4p, and chromosome segregation. Overexpression of the stable Cse4p mutant, cse4K16R, resulted in its mislocalization, increased association with chromatin, and a high rate of chromosome loss, all of which were suppressed by constitutive expression of histone H3 (Δ16H3). We determined that Δ16H3 did not lead to increased chromosome loss; however, increasing the dosage of histone H3 (GALH3) resulted in significant chromosome loss due to reduced levels of centromere (CEN)-associated Cse4p and synthetic dosage lethality (SDL) in kinetochore mutants. These phenotypes were suppressed by GALCSE4. We conclude that the chromosome missegregation of GALcse4K16R and GALH3 strains is due to mislocalization and a functionally compromised kinetochore, respectively. Suppression of these phenotypes by histone Δ16H3 and GALCSE4 supports the conclusion that proper stoichiometry affects the localization of histone H3 and Cse4p and is thus essential for accurate chromosome segregation

Integration of biological networks and gene expression data using Cytoscape

Cytoscape is a free software package for visualizing, modeling and analyzing molecular and genetic interaction networks. This protocol explains how to use Cytoscape to analyze the results of mRNA expression profiling, and other functional genomics and proteomics experiments, in the context of an interaction network obtained for genes of interest. Five major steps are described: (i) obtaining a gene or protein network, (ii) displaying the network using layout algorithms, (iii) integrating with gene expression and other functional attributes, (iv) identifying putative complexes and functional modules and (v) identifying enriched Gene Ontology annotations in the network. These steps provide a broad sample of the types of analyses performed by Cytoscape