Project description:We report the direct target genes of ATF4 and CHOP in response to endoplasim reticulum stress through next generation sequencing. By obtaining genowide sequence from chromatin immunoprecipitated DNA with anti-CHOP and anit-ATF4, we identified direct binding sites of ATF4 and CHOP in promoter regions of their target genes in mouse embrynic fibroblasts (MEFs) in response to ER stress. In addition, we obtained list of genes which are differentially regulated in Atf4 or Chop-deficient MEFs compared to the wild-type MEFs in response to ER stress. We found that genes related with unfolded protein response and protein synthesis were directly regulated by ATF4 and CHOP. Through this observation, we conclude that main role of ATF4 and CHOP as transcription factors is to enhance mRNA translation in respone to ER stress. This sutdy provide new insight of genetic network of ATF4 and CHOP in response to ER stress. For ChIP-seq, Chop+/+ and Chop-/- MEFs were treated with Tunicamycin, N-glycosylation inhibitor, to induce ER stress for 8hr. Atf4+/+ and Atf4-/- MEFs were also treated same condition for ChIP-Seq. For mRNA-seq, wild-type, Atf4-/-, and Chop-/- MEFs were treated with tunicamycin for 8hr for experiments.
Project description:Idiopathic pulmonary fibrosis (IPF) is a progressive and ultimately fatal interstitial lung disease characterized by alveolar epithelial cell type II (AECII) injury and enhanced fibroblast/myofibroblast proliferation. Recent data show that maladaptive endoplasmic reticulum (ER)-stress leading to AECII apoptosis could play a key role in IPF, although it is still unclear how precisely such response is initiated and signalled. C/EBP homologous protein (CHOP) has been suggested to contribute to a maladaptive ER-stress response in general and has been found to be induced and to be translocated into the nucleus of AECII in IPF in particular. We here show for the first time a novel mechanism for the regulation of CHOP expression during ER-stress in AECII via AP-1 and c-Ets-1. We found these two transcription factors to be up-regulated in AECII under ER-stress conditions and to interact with each other to jointly bind to the CHOP promoter, thereby inducing CHOP gene expression. Moreover, we show for the first time that singular CHOP overexpression in vitro and in vivo is indeed capable of inducing apoptosis of AECII and, consequently, lung fibroblast proliferation and up-regulation of pro-fibrotic markers. We therefore believe that CHOP plays a key role in AEC injury and consecutive fibrosis.
Project description:HAMLET triggers a p38- and ER stress-dependent death response in carcinoma cells. Transcriptome and proteome analysis detected an increase in p38 expression and phosphorylation exclusively in carcinoma cells and p38 inhibitors delayed the death response to HAMLET in carcinoma and lymphoma cells. ER stress gene expression was also increased in tumor cells and HAMLET triggered rapid XBP1 mRNA splicing, eIF2a phosphorylation, and ATF6 cleavage as well as Hsc70 and CHOP activation, suggesting that ER stress caused by HAMLET may trigger p38 phosphorylation and death. The p38 inhibitor reduced the transcription of both p38 and ER stress gene transcription. Healthy differentiated cells, in contrast, showed no alteration in p38 signaling but a rapid innate immune response was detected and the cells survived HAMLET challenge. 2 cell lines, time course, HAMLET treatment
Project description:Primary mouse embryonic fibroblasts (pass 3 to 4) were treated with tunicamycin (2 micrograms/ml) for specified times - genotypes were WILDTYPE and CHOP-/-. Each time point n=4. C/EBP homologous protein CHOP is activated by ER stress, and CHOP deletion protects against its lethal consequences.
Project description:We report the direct target genes of ATF4 and CHOP in response to endoplasim reticulum stress through next generation sequencing. By obtaining genowide sequence from chromatin immunoprecipitated DNA with anti-CHOP and anit-ATF4, we identified direct binding sites of ATF4 and CHOP in promoter regions of their target genes in mouse embrynic fibroblasts (MEFs) in response to ER stress. In addition, we obtained list of genes which are differentially regulated in Atf4 or Chop-deficient MEFs compared to the wild-type MEFs in response to ER stress. We found that genes related with unfolded protein response and protein synthesis were directly regulated by ATF4 and CHOP. Through this observation, we conclude that main role of ATF4 and CHOP as transcription factors is to enhance mRNA translation in respone to ER stress. This sutdy provide new insight of genetic network of ATF4 and CHOP in response to ER stress.
