Dual role for inositol-requiring enzyme 1α in promoting the development of hepatocellular carcinoma during diet-induced obesity in mice.

Obesity is associated with both endoplasmic reticulum (ER) stress and chronic metabolic inflammation. ER stress activates the unfolded protein response (UPR) and has been implicated in a variety of cancers, including hepatocellular carcinoma (HCC). It is unclear whether individual UPR pathways are mechanistically linked to HCC development, however. Here we report a dual role for inositol-requiring enzyme 1α (IRE1α), the ER-localized UPR signal transducer, in obesity-promoted HCC development. We found that genetic ablation of IRE1α in hepatocytes not only markedly reduced the occurrence of diethylnitrosamine (DEN)-induced HCC in liver-specific IRE1α knockout (LKO) mice when fed a normal chow (NC) diet, but also protected against the acceleration of HCC progression during high-fat diet (HFD) feeding. Irrespective of their adiposity states, LKO mice showed decreased hepatocyte proliferation and signal transducer and activator of transcription 3 (STAT3) activation, even in the face of increased hepatic apoptosis. Furthermore, IRE1α abrogation blunted obesity-associated activation of hepatic inhibitor of nuclear factor kappa B kinase subunit beta (IKKß)-nuclear factor kappa B (NF-κB) pathway, leading to reduced production of the tumor-promoting inflammatory cytokines tumor necrosis factor (TNF) and interleukin 6 (IL-6). Importantly, higher IRE1α expression along with elevated STAT3 phosphorylation was also observed in the tumor tissues from human HCC patients, correlating with their poorer survival rate. CONCLUSION: IRE1α acts in a feed-forward loop during obesity-induced metabolic inflammation to promote HCC development through STAT3-mediated hepatocyte proliferation. (Hepatology 2018).

Functional analysis of the inhibitor of apoptosis genes in Antheraea pernyi nucleopolyhedrovirus.

The inhibitor of apoptosis proteins (IAP) plays an important role in cell apoptosis. We cloned two novel IAP family members, Ap-iap1 and Ap-iap2, from Antheraea pernyi nucleopolyhedrovirus (ApNPV) genome. Ap-IAP1 contains two baculoviral IAP repeat (BIR) domains followed by a RING domain, but Ap-IAP2 has only one BIR domain and RING. The result of transient expression in Spodoptera frugiperda (Sf21) showed that Ap-iap1 blocked cell apoptosis induced by actinomycin D treatment and also rescued the p35 deficient Autographa californica nucleopolyhedrovirus (AcNPV) to replicate in Sf9 cells, while Ap-iap2 does not have this function. Several Ap-IAP1 truncations were constructed to test the activity of BIRs or RING motif to inhibit cell apoptosis. The results indicated that BIRs or RING of Ap-IAP1 had equally function to inhibit cell apoptosis. Therefore deletion of above both of the above domains could not block apoptosis induced by actinomycin D or rescue the replication of AcMNPV Delta p35. We also screened two phage-display peptides that might interact with Ap-IAP1.

Functional analysis of the orf390 gene of the white spot syndrome virus.

orf390 (WSSV449) is a novel apoptosis inhibitor gene in the genome of the White Spot Syndrome Virus (WSSV). In the present study, we focus on the function of orf390 gene. Stable expression of orf390 prevented SF9 insect cells from undergoing actinomycin D-induced apoptosis. ORF390 also rescued the replication of a p35-deficient-mutant (AcMNPVDeltap35k/pol+) in SF9 cells. In addition, ORF390 inhibits the activities of caspase-3 and -9 in vivo and in vitro. Here we demonstrate that the anti-apoptotic activity of ORF390 is dependent on two putative caspase-9 cleavage sites (VETD233 downward arrowG and LEHD303 downward arrowG) and one caspase-3 cleavage site (DEVD272 downward arrowG). Our results support the conclusion that these three sites play a key role in the suppression of apoptosis mediated by ORF390. These data further suggest that orf390 encodes a novel anti-apoptotic protein involved in cell survival and apoptosis regulation.

Overexpression of Pygopus2 protects HeLa cells from vinblastine-induced apoptosis.

Pygopus, a very important component of the Wnt signaling transcriptional complex, has multiple functions in both Wnt-dependent and -independent pathways. Human Pygopus2 (Pygo2) is expressed in many cancers and plays an important role in tumor growth. In the present study, we generated human carcinoma (HeLa) cell lines stably expressing Pygo2, which counteracts vinblastine- induced apoptosis. The anti-apoptotic function was determined by DNA fragmentation, sub-G1 appearance, loss of mitochondrial membrane potential (Deltapsim) and the activation of caspase-9 and caspase-3. In addition, we found that Pygo2 effectively blocks vinblastineinduced c-Jun and AP-1 activation, maintains the anti-apoptotic protein Bcl-2 in an unphosphorylated state, and thus can render cells resistant to apoptosis. However, Pygo2 does not alter the vinblastine-induced cell cycle changes. Here, we describe an anti-apoptotic activity exerted by Pygo2 through blocking activation of the JNK/AP-1 signaling pathway induced by vinblastine.

Selectively oncolytic mutant of HSV-1 lyses HeLa cells mediated by Ras/RTN3.

The selectively oncolytic mtHSV, a HSV icp34.5 mutant with lacz gene insertion, was proved that it was targeted for treating tumors but not other organs, however, its oncolytic mechanism is under confirmation. The results showed that HeLa cells could be lysed efficiently by mtHSV in vitro. In the flow cytometry and Western blot experiment, Ras protein was obviously downregulated on plasma membrane (PM) while the whole Ras protein didn't change along with upregulation of reticulon 3(RTN3) protein at 48 h post infection of mtHSV in HeLa cells. Expression of Ras protein on PM and whole Ras protein in HeLa cells was downregulated by siFTa (inhibitor of a subunits of human farnesyltransferase with siRNA) and siRTN3(inhibitor of RTN3 with siRNA) respectively, and HeLa cells could be killed effectively by siFTa and siRTN3 at 48 h post transfection. So siFTa and siRTN3 effectively suppressed mtHSV infection of HeLa cells. Further, experiments were made to study the relationship between Ras and RTN3 using confocal colocalization and coimmunoprecipitation. The results exhibited that Ras could interact with RTN3 at endoplasmic reticulum. The data put forward that Ras/RTN3 is an important access to HeLa cells for mtHSV. The molecular interaction between Ras and RTN3 may further improve the understanding of the function of Ras and RTN3 in mtHSV infection. The results provide further theoretical evidence that mtHSV may be used as an oncolytic agent for cancer therapy.