Molecular and Cellular Biology最新文献

{"title":"Persistent Acetylation of Histone H3 Lysine 56 Compromises the Activity of DNA Replication Origins.","authors":"Roch Tremblay, Yosra Mehrjoo, Oumaima Ahmed, Antoine Simoneau, Mary E McQuaid, El Bachir Affar, Corey Nislow, Guri Giaever, Hugo Wurtele","doi":"10.1080/10985549.2023.2259739","DOIUrl":"10.1080/10985549.2023.2259739","url":null,"abstract":"<p><p>In <i>Saccharomyces cerevisiae</i>, newly synthesized histones H3 are acetylated on lysine 56 (H3 K56ac) by the Rtt109 acetyltransferase prior to their deposition on nascent DNA behind replication forks. Two deacetylases of the sirtuin family, Hst3 and Hst4, remove H3 K56ac from chromatin after S phase. <i>hst3Δ hst4Δ</i> cells present constitutive H3 K56ac, which sensitizes cells to replicative stress via unclear mechanisms. A chemogenomic screen revealed that <i>DBF4</i> heterozygosity sensitizes cells to NAM-induced inhibition of sirtuins. <i>DBF4</i> and <i>CDC7</i> encode subunits of the Dbf4-dependent kinase (DDK), which activates origins of DNA replication during S phase. We show that (i) cells harboring the <i>dbf4-1</i> or <i>cdc7-4</i> hypomorphic alleles are sensitized to NAM, and that (ii) the sirtuins Sir2, Hst1, Hst3, and Hst4 promote DNA replication in <i>cdc7-4</i> cells. We further demonstrate that Rif1, an inhibitor of DDK-dependent activation of origins, causes DNA damage and replication defects in NAM-treated cells and <i>hst3</i>Δ <i>hst4</i>Δ mutants. <i>cdc7-4 hst3Δ hst4Δ</i> cells are shown to display delayed initiation of DNA replication, which is not due to intra-S checkpoint activation but requires Rtt109-dependent H3 K56ac. Our results suggest that constitutive H3 K56ac sensitizes cells to replicative stress in part by negatively influencing the activation of origins of DNA replication.</p>","PeriodicalId":18658,"journal":{"name":"Molecular and Cellular Biology","volume":null,"pages":null},"PeriodicalIF":3.2,"publicationDate":"2023-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10791153/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"41127303","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Distinct Interaction Modes for the Eukaryotic RNA Polymerase Alpha-like Subunits.","authors":"Alana E Belkevich, Haleigh G Pascual, Aula M Fakhouri, David G Ball, Bruce A Knutson","doi":"10.1080/10985549.2023.2210023","DOIUrl":"10.1080/10985549.2023.2210023","url":null,"abstract":"<p><p>Eukaryotic DNA-dependent RNA polymerases (Pols I-III) encode two distinct alpha-like heterodimers where one is shared between Pols I and III, and the other is unique to Pol II. Human alpha-like subunit mutations are associated with several diseases including Treacher Collins Syndrome (TCS), 4H leukodystrophy, and primary ovarian sufficiency. Yeast is commonly used to model human disease mutations, yet it remains unclear whether the alpha-like subunit interactions are functionally similar between yeast and human homologs. To examine this, we mutated several regions of the yeast and human small alpha-like subunits and used biochemical and genetic assays to establish the regions and residues required for heterodimerization with their corresponding large alpha-like subunits. Here we show that different regions of the small alpha-like subunits serve differential roles in heterodimerization, in a polymerase- and species-specific manner. We found that the small human alpha-like subunits are more sensitive to mutations, including a \"humanized\" yeast that we used to characterize the molecular consequence of the TCS-causingPOLR1D G52E mutation. These findings help explain why some alpha subunit associated disease mutations have little to no effect when made in their yeast orthologs and offer a better yeast model to assess the molecular basis of POLR1D associated disease mutations.</p>","PeriodicalId":18658,"journal":{"name":"Molecular and Cellular Biology","volume":null,"pages":null},"PeriodicalIF":3.2,"publicationDate":"2023-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10251799/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9605115","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Recruitment of RBM6 to DNA Double-Strand Breaks Fosters Homologous Recombination Repair.","authors":"Samah W Awwad, Malak M Darawshe, Feras E Machour, Inbar Arman, Nabieh Ayoub","doi":"10.1080/10985549.2023.2187105","DOIUrl":"10.1080/10985549.2023.2187105","url":null,"abstract":"<p><p>DNA double-strand breaks (DSBs) are highly toxic lesions that threaten genome integrity and cell survival. To avoid harmful repercussions of DSBs, a wide variety of DNA repair factors are recruited to execute DSB repair. Previously, we demonstrated that RBM6 splicing factor facilitates homologous recombination (HR) of DSB by regulating alternative splicing-coupled nonstop-decay of the HR protein APBB1/Fe65. Here, we describe a splicing-independent function of RBM6 in promoting HR repair of DSBs. We show that RBM6 is recruited to DSB sites and PARP1 activity indirectly regulates RBM6 recruitment to DNA breakage sites. Deletion mapping analysis revealed a region containing five glycine residues within the G-patch domain that regulates RBM6 accumulation at DNA damage sites. We further ascertain that RBM6 interacts with Rad51, and this interaction is attenuated in RBM6 mutant lacking the G-patch domain (RBM6^del(G-patch)). Consequently, RBM6^del(G-patch) cells exhibit reduced levels of Rad51 foci after ionizing radiation. In addition, while RBM6 deletion mutant lacking the G-patch domain has no detectable effect on the expression levels of its splicing targets Fe65 and Eya2, it fails to restore the integrity of HR. Altogether, our results suggest that RBM6 recruitment to DSB promotes HR repair, irrespective of its splicing activity.HIGHLIGHTSPARP1 activity indirectly regulates RBM6 recruitment to DNA damage sites.Five glycine residues within the G-patch domain of RBM6 are critical for its recruitment to DNA damage sites, but dispensable for its splicing activity.RBM6 G-patch domain fosters its interaction with Rad51 and promotes Rad51 foci formation following irradiation.RBM6 recruitment to DSB sites underpins HR repair.</p>","PeriodicalId":18658,"journal":{"name":"Molecular and Cellular Biology","volume":null,"pages":null},"PeriodicalIF":5.3,"publicationDate":"2023-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10038030/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9646913","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Correction.","authors":"","doi":"10.1080/10985549.2023.2279475","DOIUrl":"10.1080/10985549.2023.2279475","url":null,"abstract":"","PeriodicalId":18658,"journal":{"name":"Molecular and Cellular Biology","volume":null,"pages":null},"PeriodicalIF":5.3,"publicationDate":"2023-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10761144/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"92155347","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Hsp70 Binding to the N-terminal Domain of Hsp104 Regulates [<i>PSI⁺</i>] Curing by Hsp104 Overexpression.","authors":"Xiaohong Zhao, Katherine Stanford, Joseph Ahearn, Daniel C Masison, Lois E Greene","doi":"10.1080/10985549.2023.2198181","DOIUrl":"10.1080/10985549.2023.2198181","url":null,"abstract":"<p><p>Hsp104 propagates the yeast prion [<i>PSI⁺</i>], the infectious form of Sup35, by severing the prion seeds, but when Hsp104 is overexpressed, it cures [<i>PSI⁺</i>] in a process that is not yet understood but may be caused by trimming, which removes monomers from the ends of the amyloid fibers. This curing was shown to depend on both the N-terminal domain of Hsp104 and the expression level of various members of the Hsp70 family, which raises the question as to whether these effects of Hsp70 are due to it binding to the Hsp70 binding site that was identified in the N-terminal domain of Hsp104, a site not involved in prion propagation. Investigating this question, we now find, first, that mutating this site prevents both the curing of [<i>PSI⁺</i>] by Hsp104 overexpression and the trimming activity of Hsp104. Second, we find that depending on the specific member of the Hsp70 family binding to the N-terminal domain of Hsp104, both trimming and the curing caused by Hsp104 overexpression are either increased or decreased in parallel. Therefore, the binding of Hsp70 to the N-terminal domain of Hsp104 regulates both the rate of [<i>PSI⁺</i>] trimming by Hsp104 and the rate of [<i>PSI⁺</i>] curing by Hsp104 overexpression.</p>","PeriodicalId":18658,"journal":{"name":"Molecular and Cellular Biology","volume":null,"pages":null},"PeriodicalIF":5.3,"publicationDate":"2023-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10153015/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9462565","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Transcription Repression of CRY2 via PER2 Interaction Promotes Adipogenesis.","authors":"Weini Li, Xuekai Xiong, Tali Kiperman, Ke Ma","doi":"10.1080/10985549.2023.2253710","DOIUrl":"10.1080/10985549.2023.2253710","url":null,"abstract":"<p><p>The circadian clock is driven by a transcriptional-translational feedback loop, and cryptochrome 2 (CRY2) represses CLOCK/BMAL1-induced transcription activation. Despite the established role of clock in adipogenic regulation, whether the CRY2 repressor activity functions in adipocyte biology remains unclear. Here we identify a critical cysteine residue of CRY2 that mediates interaction with Period 2 (PER2). We further demonstrate that this mechanism is required for repressing circadian clock-controlled Wnt signaling to promote adipogenesis. CRY2 protein is enriched in white adipose depots and robustly induced by adipogenic differentiation. Via site-directed mutagenesis, we identified that a conserved CRY2 cysteine at 432 within the loop interfacing with PER2 mediates heterodimer complex formation that confers transcription repression. C432 mutation disrupted PER2 association without affecting BMAL1 binding, leading to loss of repression of clock transcription activation. In preadipocytes, whereas CRY2 enhanced adipocyte differentiation, the repression-defective C432 mutant suppressed this process. Furthermore, silencing of CRY2 attenuated, while stabilization of CRY2 by KL001 markedly augmented adipocyte maturation. Mechanistically, we show that transcriptional repression of Wnt pathway components underlies CRY2 modulation of adipogenesis. Collectively, our findings elucidate a CRY2-mediated repression mechanism that promotes adipocyte development, and implicate its potential as a clock intervention target for obesity.</p>","PeriodicalId":18658,"journal":{"name":"Molecular and Cellular Biology","volume":null,"pages":null},"PeriodicalIF":3.2,"publicationDate":"2023-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10569361/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"41151058","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"A SNAI2/CTCF Interaction is Required for <i>NOTCH1</i> Expression in Rhabdomyosarcoma.","authors":"Prethish Sreenivas, Long Wang, Meng Wang, Anil Challa, Paulomi Modi, Nicole Rae Hensch, Berkley Gryder, Hsien-Chao Chou, Xiang R Zhao, Benjamin Sunkel, Rodrigo Moreno-Campos, Javed Khan, Benjamin Z Stanton, Myron S Ignatius","doi":"10.1080/10985549.2023.2256640","DOIUrl":"10.1080/10985549.2023.2256640","url":null,"abstract":"<p><p>Rhabdomyosarcoma (RMS) is a pediatric malignancy of the muscle with characteristics of cells blocked in differentiation. <i>NOTCH1</i> is an oncogene that promotes self-renewal and blocks differentiation in the fusion negative-RMS sub-type. However, how <i>NOTCH1</i> expression is transcriptionally maintained in tumors is unknown. Analyses of SNAI2 and CTCF chromatin binding and HiC analyses revealed a conserved SNAI2/CTCF overlapping peak downstream of the <i>NOTCH1</i> locus marking a sub-topologically associating domain (TAD) boundary. Deletion of the SNAI2-CTCF peak showed that it is essential for <i>NOTCH1</i> expression and viability of FN-RMS cells. Reintroducing constitutively activated <i>NOTCH1</i>-ΔE in cells with the SNAI2-CTCF peak deleted restored cell-viability. Ablation of SNAI2 using CRISPR/Cas9 reagents resulted in the loss of majority of RD and SMS-CTR FN-RMS cells. However, the few surviving clones that repopulate cultures have recovered <i>NOTCH1</i>. Cells that re-establish <i>NOTCH1</i> expression after SNAI2 ablation are unable to differentiate robustly as SNAI2 shRNA knockdown cells; yet, <i>SNAI2</i>-ablated cells continued to be exquisitely sensitive to ionizing radiation. Thus, we have uncovered a novel mechanism by which SNAI2 and CTCF maintenance of a sub-TAD boundary promotes rather than represses <i>NOTCH1</i> expression. Further, we demonstrate that SNAI2 suppression of apoptosis post-radiation is independent of <i>SNAI2</i>/<i>NOTCH1</i> effects on self-renewal and differentiation.</p>","PeriodicalId":18658,"journal":{"name":"Molecular and Cellular Biology","volume":null,"pages":null},"PeriodicalIF":3.2,"publicationDate":"2023-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10761179/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"50162157","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Mitochondrial Fragmentation Promotes Inflammation Resolution Responses in Macrophages via Histone Lactylation.","authors":"Leah I Susser, My-Anh Nguyen, Michele Geoffrion, Christina Emerton, Mireille Ouimet, Mireille Khacho, Katey J Rayner","doi":"10.1080/10985549.2023.2253131","DOIUrl":"10.1080/10985549.2023.2253131","url":null,"abstract":"<p><p>During the inflammatory response, macrophage phenotypes can be broadly classified as pro-inflammatory/classically activated \"M1\", or pro-resolving/alternatively \"M2\" macrophages. Although the classification of macrophages is general and assumes there are distinct phenotypes, in reality macrophages exist across a spectrum and must transform from a pro-inflammatory state to a proresolving state following an inflammatory insult. To adapt to changing metabolic needs of the cell, mitochondria undergo fusion and fission, which have important implications for cell fate and function. We hypothesized that mitochondrial fission and fusion directly contribute to macrophage function during the pro-inflammatory and proresolving phases. In the present study, we find that mitochondrial length directly contributes to macrophage phenotype, primarily during the transition from a pro-inflammatory to a proresolving state. Phenocopying the elongated mitochondrial network (by disabling the fission machinery using siRNA) leads to a baseline reduction in the inflammatory marker IL-1β, but a normal inflammatory response to LPS, similar to control macrophages. In contrast, in macrophages with a phenocopied fragmented phenotype (by disabling the fusion machinery using siRNA) there is a heightened inflammatory response to LPS and increased signaling through the ATF4/c-Jun transcriptional axis compared to control macrophages. Importantly, macrophages with a fragmented mitochondrial phenotype show increased expression of proresolving mediator arginase 1 and increased phagocytic capacity. Promoting mitochondrial fragmentation caused an increase in cellular lactate, and an increase in histone lactylation which caused an increase in arginase 1 expression. These studies demonstrate that a fragmented mitochondrial phenotype is critical for the proresolving response in macrophages and specifically drive epigenetic changes via lactylation of histones following an inflammatory insult.</p>","PeriodicalId":18658,"journal":{"name":"Molecular and Cellular Biology","volume":null,"pages":null},"PeriodicalIF":3.2,"publicationDate":"2023-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://ftp.ncbi.nlm.nih.gov/pub/pmc/oa_pdf/d5/8a/TMCB_43_2253131.PMC10569354.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"41135311","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Hypoxia-Inducible Factor-2alpha Affects the MEK/ERK Signaling Pathway via Primary Cilia in Connection with the Intraflagellar Transport Protein 88 Homolog.","authors":"Tristan Leu, Jannik Denda, Anna Wrobeln, Joachim Fandrey","doi":"10.1080/10985549.2023.2198931","DOIUrl":"10.1080/10985549.2023.2198931","url":null,"abstract":"<p><p>The ability of cells to communicate with their surrounding is a prerequisite for essential processes such as proliferation, apoptosis, migration, and differentiation. To this purpose, primary cilia serve as antennae-like structures on the surface of most mammalian cell types. Cilia allow signaling via hedgehog, Wnt or TGF-beta pathways. Their length, in part controlled by the activity of intraflagellar transport (IFT), is a parameter for adequate function of primary cilia. Here we show, in murine neuronal cells, that intraflagellar transport protein 88 homolog (IFT88) directly interacts with the hypoxia-inducible factor-2α (HIF-2α), hitherto known as an oxygen-regulated transcription factor. Furthermore, HIF-2α accumulates in the ciliary axoneme and promotes ciliary elongation under hypoxia. Loss of HIF-2α affected ciliary signaling in neuronal cells by decreasing transcription of <i>Mek1/2</i> and <i>Erk1/2</i>. Targets of the MEK/ERK signaling pathway, such as <i>Fos</i> and <i>Jun</i>, were significantly decreased. Our results suggest that HIF-2α influences ciliary signaling by interacting with IFT88 under hypoxic conditions. This implies an unexpected and far more extensive function of HIF-2α than described before.</p>","PeriodicalId":18658,"journal":{"name":"Molecular and Cellular Biology","volume":null,"pages":null},"PeriodicalIF":5.3,"publicationDate":"2023-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10153011/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9458670","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"The \"LINC\" between Δ40p53-miRNA Axis in the Regulation of Cellular Homeostasis.","authors":"Apala Pal, Pritam Kumar Ghosh, Saumitra Das","doi":"10.1080/10985549.2023.2213147","DOIUrl":"10.1080/10985549.2023.2213147","url":null,"abstract":"<p><p>Previous research has shown that Δ40p53, the translational isoform of p53, can inhibit cell growth independently of p53 by regulating microRNAs. Here, we explored the role of Δ40p53 in regulating the long noncoding RNA-micro-RNA-cellular process axis, specifically focusing on <i>LINC00176</i>. Interestingly, <i>LINC00176</i> levels were predominantly affected by the overexpression/stress-mediated induction and knockdown of Δ40p53 rather than p53 levels. Additional assays revealed that Δ40p53 transactivates <i>LINC00176</i> transcriptionally and could also regulate its stability. RNA immunoprecipitation experiments revealed that <i>LINC00176</i> sequesters several putative microRNA targets, which could further titrate several mRNA targets involved in different cellular processes. To understand the downstream effects of this regulation, we ectopically overexpressed and knocked down <i>LINC00176</i> in HCT116 p53-/- (harboring only Δ40p53) cells, which affected their proliferation, cell viability, and expression of epithelial markers. Our results provide essential insights into the pivotal role of Δ40p53 in regulating the novel <i>LINC00176</i> RNA-microRNA-mRNA axis independent of FL-p53 and in maintaining cellular homeostasis.</p>","PeriodicalId":18658,"journal":{"name":"Molecular and Cellular Biology","volume":null,"pages":null},"PeriodicalIF":5.3,"publicationDate":"2023-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10348045/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9796862","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}