Chaperonin-containing TCP-1 (CCT or TRiC) is a multi-subunit complex that folds many of the proteins essential for cancer development. CCT is expressed in diverse cancers and could be an ideal therapeutic target if not for the fact that the complex is encoded by eight distinct genes, complicating the development of inhibitors. Few definitive studies addressed the role of specific subunits in promoting the chaperonin’s function in cancer. To this end, we investigated the activity of CCT2 (CCTβ) by overexpressing or depleting the subunit in breast epithelial and breast cancer cells. We found that increasing total CCT2 in cells by 1.3-1.8-fold using a lentiviral system, also caused CCT3, CCT4, and CCT5 levels to increase. Likewise, silencing
Cohesin is an important structural regulator of the genome, regulating both three-dimensional genome organization and gene expression. The core cohesin trimer interacts with various HEAT repeat accessory subunits, yielding cohesin complexes of distinct compositions and potentially distinct functions. The roles of the two mutually exclusive HEAT repeat subunits PDS5A and PDS5B are not well understood.
Here, we determine that PDS5A and PDS5B have highly similar localization patterns across the mouse embryonic stem cell (mESC) genome and they show a strong overlap with other cohesin HEAT repeat accessory subunits, STAG1 and STAG2. Using CRISPR/Cas9 genome editing to generate individual stable knockout lines for PDS5A and PDS5B, we find that loss of one PDS5 subunit does not alter the distribution of the other PDS5 subunit, nor the core cohesin complex. Both PDS5A and PDS5B are required for proper gene expression, yet they display only partially overlapping effects on gene targets. Remarkably, gene expression following dual depletion of the PDS5 HEAT repeat proteins does not completely overlap the gene expression changes caused by dual depletion of the STAG HEAT repeat proteins, despite the overlapping genomic distribution of all four proteins. Furthermore, dual loss of PDS5A and PDS5B decreases cohesin association with NIPBL more »
This work reveals the importance of PDS5A and PDS5B for proper cohesin function. Loss of either subunit has little effect on cohesin localization across the genome yet PDS5A and PDS5B are differentially required for gene expression.
- Publication Date:
- NSF-PAR ID:
- 10369720
- Journal Name:
- Epigenetics & Chromatin
- Volume:
- 15
- Issue:
- 1
- ISSN:
- 1756-8935
- Publisher:
- Springer Science + Business Media
- Sponsoring Org:
- National Science Foundation
More Like this
-
Abstract cct2 gene expression by ~50% caused other CCT subunits to decrease. Cells expressing CCT2 were more invasive and had a higher proliferative index. CCT2 depletion in a syngeneic murine model of triple negative breast cancer (TNBC) prevented tumor growth. These results indicate that the CCT2 subunit is integral to the activity of the chaperonin and is needed for tumorigenesis. Hence CCT2 could be a viable target for therapeutic development in breast and other cancers. -
Regulation of gene expression starts from the transcription initiation. Regulated transcription initiation is critical for generating correct transcripts with proper abundance. The impact of epigenetic control, such as histone modifications and chromatin remodelling, on gene regulation has been extensively investigated, but their specific role in regulating transcription initiation is far from well understood. Here we aimed to better understand the roles of genes involved in histone H3 methylations and chromatin remodelling on the regulation of transcription initiation at a genome-scale using the budding yeast as a study system. We obtained and compared maps of transcription start site (TSS) at single-nucleotide resolution by nAnT-iCAGE for a strain with depletion of MINC (Mot1-Ino80C-Nc2) by Mot1p and Ino80p anchor-away (Mot1&Ino80AA) and a strain with loss of histone methylation (set1Δset2Δdot1Δ) to their wild-type controls. Our study showed that the depletion of MINC stimulated transcription initiation from many new sites flanking the dominant TSS of genes, while the loss of histone methylation generates more TSSs in the coding region. Moreover, the depletion of MINC led to less confined boundaries of TSS clusters (TCs) and resulted in broader core promoters, and such patterns are not present in the ssdΔ mutant. Our data also exhibits that themore »
-
Abstract During meiotic prophase I, accurate segregation of homologous chromosomes requires the establishment of chromosomes with a meiosis-specific architecture. The sister chromatid cohesin complex and the enzyme Topoisomerase II (TOP-2) are important components of meiotic chromosome architecture, but the relationship of these proteins in the context of meiotic chromosome segregation is poorly defined. Here, we analyzed the role of TOP-2 in the timely release of the sister chromatid cohesin subunit REC-8 during spermatogenesis and oogenesis of Caenorhabditis elegans. We show that there is a different requirement for TOP-2 in meiosis of spermatogenesis and oogenesis. The loss-of-function mutation top-2(it7) results in premature REC-8 removal in spermatogenesis, but not oogenesis. This correlates with a failure to maintain the HORMA-domain proteins HTP-1 and HTP-2 (HTP-1/2) on chromosome axes at diakinesis and mislocalization of the downstream components that control REC-8 release including Aurora B kinase. In oogenesis, top-2(it7) causes a delay in the localization of Aurora B to oocyte chromosomes but can be rescued through premature activation of the maturation promoting factor via knockdown of the inhibitor kinase WEE-1.3. The delay in Aurora B localization is associated with an increase in the length of diakinesis bivalents and wee-1.3 RNAi mediated rescue of Aurora Bmore »
-
Abstract Background Spiders have evolved two types of sticky capture threads: one with wet adhesive spun by ecribellate orb-weavers and another with dry adhesive spun by cribellate spiders. The evolutionary history of cribellate capture threads is especially poorly understood. Here, we use genomic approaches to catalog the spider-specific silk gene family (spidroins) for the cribellate orb-weaver
Uloborus diversus .Results We show that the cribellar spidroin, which forms the puffy fibrils of cribellate threads, has three distinct repeat units, one of which is conserved across cribellate taxa separated by ~ 250 Mya. We also propose candidates for a new silk type, paracribellar spidroins, which connect the puffy fibrils to pseudoflagelliform support lines. Moreover, we describe the complete repeat architecture for the pseudoflagelliform spidroin (Pflag), which contributes to extensibility of pseudoflagelliform axial fibers.
Conclusions Our finding that Pflag is closely related to Flag, supports homology of the support lines of cribellate and ecribellate capture threads. It further suggests an evolutionary phase following gene duplication, in which both Flag and Pflag were incorporated into the axial lines, with subsequent loss of Flag in uloborids, and increase in expression of Flag in ecribellate orb-weavers, explaining the distinct mechanical properties of the axial lines of these two groups.
-
Abstract Pan-genome analyses of metagenome-assembled genomes (MAGs) may suffer from the known issues with MAGs: fragmentation, incompleteness and contamination. Here, we conducted a critical assessment of pan-genomics of MAGs, by comparing pan-genome analysis results of complete bacterial genomes and simulated MAGs. We found that incompleteness led to significant core gene (CG) loss. The CG loss remained when using different pan-genome analysis tools (Roary, BPGA, Anvi’o) and when using a mixture of MAGs and complete genomes. Contamination had little effect on core genome size (except for Roary due to in its gene clustering issue) but had major influence on accessory genomes. Importantly, the CG loss was partially alleviated by lowering the CG threshold and using gene prediction algorithms that consider fragmented genes, but to a less degree when incompleteness was higher than 5%. The CG loss also led to incorrect pan-genome functional predictions and inaccurate phylogenetic trees. Our main findings were supported by a study of real MAG-isolate genome data. We conclude that lowering CG threshold and predicting genes in metagenome mode (as Anvi’o does with Prodigal) are necessary in pan-genome analysis of MAGs. Development of new pan-genome analysis tools specifically for MAGs are needed in future studies.
