C/EBP proteins are known to have a conservedC-terminal structure,basic leucine zipper domain(bZIP), that is necessary for the formation of DNA-binding capable homodimers or heterodimers with other proteins or members of the C/EBP protein family.[7] CHOP is a relatively small (29kDa) protein that differs from most C/EBP proteins in several amino acid substitutions, which impacts its DNA-binding ability.[8]
Due to a variety of upstream and downstream regulatory interactions, CHOP plays an important role inER stress-inducedapoptosis caused by a variety of stimuli such as pathogenic microbial or viralinfections, amino acid starvation, mitochondrial stress,neurological diseases, andneoplastic diseases.
DuringER stress, CHOP is mainly induced via activation of theintegrated stress response pathways through the subsequent downstream phosphorylation of a translation initiation factor,eukaryotic initiation factor 2α (eIF2α), and induction of a transcription factor, activation transcription factor 4 (ATF4),[12] which converges on thepromoters of target genes, including CHOP.
Integrated stress response, and thus CHOP expression, can be induced by
Under ER stress, activated transmembrane proteinATF6 translocates to the nucleus and interacts with ATF/cAMP response elements and ER stress-response elements,[17] binding the promoters and inducing transcription of several genes involved inunfolded protein response (including CHOP,XBP1 and others).[18][19] Thus,ATF6 activates the transcription of both CHOP andXBP-1, whileXBP-1 can also upregulate the expression of CHOP.[20]
ER stress also stimulates transmembrane protein IRE1α activity.[21] Upon activation, IRE1α splices the XBP-1 mRNA introns to produce a mature and active XBP-1 protein,[22] that upregulates CHOP expression[23][24][25]IRE1α also stimulates the activation of theapoptotic-signaling kinase-1 (ASK1), which then activates the downstream kinases,Jun-N-terminal kinase (JNK) and p38 mitogen-activated protein kinase (p38 MAPK),[26] which participate inapoptosis induction along with CHOP.[27] TheP38 MAP kinase family phosphorylates Ser78 and Ser81 of CHOP, which induces cell apoptosis.[28] Moreover, research studies found that theJNK inhibitors can suppress CHOP upregulation, indicating thatJNK activation is also involved in the modulation of CHOP levels.[29]
As a transcription factor, CHOP can regulate the expression of many anti-apoptotic andpro-apoptotic genes, including genes encoding theBCL2-family proteins,GADD34 andTRB-3.[30][31] In the CHOP-induced apoptotic pathway, CHOP regulates the expression ofBCL2 protein family, that includes anti-apoptotic proteins (BCL2,BCL-XL,MCL-1, andBCL-W) and pro-apoptotic proteins (BAK,BAX, BOK,BIM, PUMA and others).[32][33]
Under ER stress, CHOP can function as either atranscriptional activator orrepressor. It formsheterodimers with other C/EBP family transcription factors via bZIP-domain interactions to inhibit the expression of genes responsive to C/EBP family transcription factors, while enhancing the expression of other genes containing a specific 12–14 bp DNAcis-acting element.[34] CHOP candownregulate the expressions of anti-apoptoticBCL2 proteins, andupregulate the expression of proapoptotic proteins (BIM, BAK and BAX expression).[35][36] BAX-BAK oligomerization causescytochrome c andapoptosis-inducing factor (AIF) release from mitochondria, eventually causingcell death.[37]
TRB3 pseudokinase is upregulated by the ER stress-inducible transcriptional factor,ATF4-CHOP.[38] CHOP interacts with TRB3, which contributes to the induction of apoptosis.[39][40][41] The expression of TRB3 has a pro-apoptotic capacity.[42][43] Therefore, CHOP also regulates apoptosis by upregulating the expression of the TRB3 gene.
A summary of CHOP upstream and downstream pathways
The PERK-ATF4-CHOP pathway can induceapoptosis by binding to thedeath receptors and upregulating the expression ofdeath receptor 4 (DR4) andDR5. CHOP also interacts with the phosphorylated transcription factorJUN to form a complex that binds to the promoter region ofDR4 in lung cancer cells.[44] The N-terminal domain of CHOP interacts with phosphorylatedJUN to form a complex that regulates the expression ofDR4 andDR5.[44] CHOP also upregulates the expression ofDR5 by binding to the5′-region of the DR5 gene.[45]
In addition, CHOP also mediates apoptosis through increasing the expression of theERO1α (ER reductase)[10] gene, which catalyzes the production ofH2O2 in theER. The highly oxidized state of the ER results in H2O2 leakage into the cytoplasm, inducing the production ofreactive oxygen species (ROS) and a series of apoptotic andinflammatory reactions.[10][48][49][50]
Theoverexpression of CHOP can lead tocell cycle arrest and result in cell apoptosis. At the same time, CHOP-induced apoptosis can also trigger cell death by inhibiting the expression ofcell cycle regulatory protein, p21. Thep21 protein inhibits theG1 phase of the cell cycle as well as regulates the activity of pre-apoptotic factors. Identified CHOP-p21 relationship may play a role in changing the cell state from adapting to ER stress towards pre-apoptotic activity.[51]
In general, the downstream targets of CHOP regulate the activation of apoptotic pathways, however, the molecular interaction mechanisms behind those processes remain to be discovered.
Chop gene deletion has been demonstrated protective against diet induced metabolic syndromes in mice.[60][61] Mice withgermlineChop gene knockout have better glycemic control despite unchanged obesity. A plausible explanation for the observed dissociation between obesity and insulin resistance is that CHOP promotes insulin hypersecretion from pancreatic β cells.[62]
Furthermore,Chop depletion by a GLP1-ASO delivery system[63] was shown to have therapeutic effects of insulin reduction and fatty liver correction,[64] in preclinical mouse models.[62]
Since CHOP has an important role of apoptosis induction during infection, it is an important target for further research that will help deepen the current understanding ofpathogenesis and potentially provide an opportunity for invention of newtherapeutic approaches. For example,small molecule inhibitors of CHOP expression may act as therapeutic options to prevent ER stress and microbial infections. Research had shown that small molecule inhibitors of PERK-eIF2α pathway limitPCV2 virus replication.[65]
The regulation of CHOP expression plays an important role in metabolic diseases and in some cancers through its function in mediating apoptosis. The regulation of CHOP expression could be a potential approach to affecting cancer cells through the induction of apoptosis.[51][29][44][74] In the intestinal epithelium, CHOP has been demonstrated to be downregulated under inflammatory conditions (in inflammatory bowel diseases and experimental models of colitis). In this context, CHOP seems to rather regulate the cell cycle than apoptotic processes.[75]
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