Ching-Ping Chan

The liver-enriched transcription factor CREB-H is a growth suppressor protein underexpressed in hepatocellular carcinoma

We have previously characterized transcription factor LZIP to be a growth suppressor targeted by hepatitis C virus oncoprotein. In search of proteins closely related to LZIP, we have identified a liver-enriched transcription factor CREB-H. LZIP and CREB-H represent a new subfamily of bZIP factors. CREB-H activates transcription by binding to cAMP responsive element, box B, and ATF6-binding element. Interestingly, CREB-H has a putative transmembrane (TM) domain and it localizes ambiently to the endoplasmic reticulum. Proteolytic cleavage that removes the TM domain leads to nuclear translocation and activation of CREB-H. CREB-H activates the promoter of hepatic gluconeogenic enzyme phosphoenolpyruvate carboxykinase. This activation can be further stimulated by cAMP and protein kinase A. CREB-H transcript is exclusively abundant in adult liver. In contrast, the expression of CREB-H mRNA is aberrantly reduced in hepatoma tissues and cells. The enforced expression of CREB-H suppresses the proliferation of cultured hepatoma cells. Taken together, our findings suggest that the liver-enriched bZIP transcription factor CREB-H is a growth suppressor that plays a role in hepatic physiology and pathology.

Modulation of the Unfolded Protein Response by the Severe Acute Respiratory Syndrome Coronavirus Spike Protein

ABSTRACT Perturbation of the function of endoplasmic reticulum (ER) causes stress leading to the activation of cell signaling pathways known as the unfolded protein response (UPR). Severe acute respiratory syndrome (SARS) coronavirus (SARS-CoV) uses ER as a site for synthesis and processing of viral proteins. In this report, we demonstrate that infection with SARS-CoV induces the UPR in cultured cells. A comparison with M, E, and NSP6 proteins indicates that SARS-CoV spike (S) protein sufficiently induces transcriptional activation of several UPR effectors, including glucose-regulated protein 78 (GRP78), GRP94, and C/EBP homologous protein. A substantial amount of S protein accumulates in the ER. The expression of S protein exerts different effects on the three major signaling pathways of the UPR. Particularly, it induces GRP78/94 through PKR-like ER kinase but has no influence on activating transcription factor 6 or X box-binding protein 1. Taken together, our findings suggest that SARS-CoV S protein specifically modulates the UPR to facilitate viral replication.

LKB1 tumor suppressor and salt-inducible kinases negatively regulate human T-cell leukemia virus type 1 transcription

BackgroundHuman T-cell leukemia virus type 1 (HTLV-1) causes adult T-cell leukemia (ATL). Treatment options are limited and prophylactic agents are not available. We have previously demonstrated an essential role for CREB-regulating transcriptional coactivators (CRTCs) in HTLV-1 transcription.ResultsIn this study we report on the negative regulatory role of LKB1 tumor suppressor and salt-inducible kinases (SIKs) in the activation of HTLV-1 long terminal repeats (LTR) by the oncoprotein Tax. Activation of LKB1 and SIKs effectively blunted Tax activity in a phosphorylation-dependent manner, whereas compromising these kinases, but not AMP-dependent protein kinases, augmented Tax function. Activated LKB1 and SIKs associated with Tax and suppressed Tax-induced LTR activation by counteracting CRTCs and CREB. Enforced expression of LKB1 or SIK1 in cells transfected with HTLV-1 molecular clone pX1MT repressed proviral transcription. On the contrary, depletion of LKB1 in pX1MT-transfected cells and in HTLV-1-transformed T cells boosted the expression of Tax. Treatment of HTLV-1 transformed cells with metformin led to LKB1/SIK1 activation, reduction in Tax expression, and inhibition of cell proliferation.ConclusionsOur findings revealed a new function of LKB1 and SIKs as negative regulators of HTLV-1 transcription. Pharmaceutical activation of LKB1 and SIKs might be considered as a new strategy in anti-HTLV-1 and anti-ATL therapy.

Selective Activation of Type II Interferon Signaling by Zika Virus NS5 Protein

ABSTRACT Severe complications of Zika virus (ZIKV) infection might be caused by inflammation, but how ZIKV induces proinflammatory cytokines is not understood. In this study, we show opposite regulatory effects of the ZIKV NS5 protein on interferon (IFN) signaling. Whereas ZIKV and its NS5 protein were potent suppressors of type I and type III IFN signaling, they were found to activate type II IFN signaling. Inversely, IFN-γ augmented ZIKV replication. NS5 interacted with STAT2 and targeted it for ubiquitination and degradation, but it had no influence on STAT1 stability or nuclear translocation. The recruitment of STAT1-STAT2-IRF9 to IFN-β-stimulated genes was compromised when NS5 was expressed. Concurrently, the formation of STAT1-STAT1 homodimers and their recruitment to IFN-γ-stimulated genes, such as the gene encoding the proinflammatory cytokine CXCL10, were augmented. Silencing the expression of an IFN-γ receptor subunit or treatment of ZIKV-infected cells with a JAK2 inhibitor suppressed viral replication and viral induction of IFN-γ-stimulated genes. Taken together, our findings provide a new mechanism by which the ZIKV NS5 protein differentially regulates IFN signaling to facilitate viral replication and cause diseases. This activity might be shared by a group of viral IFN modulators. IMPORTANCE Mammalian cells produce three types of interferons to combat viral infection and to control host immune responses. To replicate and cause diseases, pathogenic viruses have developed different strategies to defeat the action of host interferons. Many viral proteins, including the Zika virus (ZIKV) NS5 protein, are known to be able to suppress the antiviral property of type I and type III interferons. Here we further show that the ZIKV NS5 protein can also boost the activity of type II interferon to induce cellular proteins that promote inflammation. This is mediated by the differential effect of the ZIKV NS5 protein on a pair of cellular transcription factors, STAT1 and STAT2. NS5 induces the degradation of STAT2 but promotes the formation of STAT1-STAT1 protein complexes, which activate genes controlled by type II interferon. A drug that specifically inhibits the IFN-γ receptor or STAT1 shows an anti-ZIKV effect and might also have anti-inflammatory activity.

Polymorphisms of CCR5 gene in a southern Chinese population and their effects on disease progression in HIV infections

Seven mutant alleles of the CCR5 gene were identified in southern Chinese, six of which were novel mutations. The most common was a heterozygous mutant allele (R223Q) found in 21 healthy donors and 20 HIV patients. No significant difference in the mutant frequency was observed between healthy donors (0.044) and HIV patients (0.041), but the frequency was significantly higher in asymptomatic (0.066) than in symptomatic HIV patients (0.019).

Comparative analysis of the activation of unfolded protein response by spike proteins of severe acute respiratory syndrome coronavirus and human coronavirus HKU1

BackgroundWhereas severe acute respiratory syndrome (SARS) coronavirus (SARS-CoV) is associated with severe disease, human coronavirus HKU1 (HCoV-HKU1) commonly circulates in the human populations causing generally milder illness. Spike (S) protein of SARS-CoV activates the unfolded protein response (UPR). It is not understood whether HCoV-HKU1 S protein has similar activity. In addition, the UPR-activating domain in SARS-CoV S protein remains to be identified.ResultsIn this study we compared S proteins of SARS-CoV and HCoV-HKU1 for their ability to activate the UPR. Both S proteins were found in the endoplasmic reticulum. Transmembrane serine protease TMPRSS2 catalyzed the cleavage of SARS-CoV S protein, but not the counterpart in HCoV-HKU1. Both S proteins showed a similar pattern of UPR-activating activity. Through PERK kinase they activated the transcription of UPR effector genes such as Grp78, Grp94 and CHOP. N-linked glycosylation was not required for the activation of the UPR by S proteins. S1 subunit of SARS-CoV but not its counterpart in HCoV-HKU1 was capable of activating the UPR. A central region (amino acids 201–400) of SARS-CoV S1 was required for this activity.ConclusionsSARS-CoV and HCoV-HKU1 S proteins use distinct UPR-activating domains to exert the same modulatory effects on UPR signaling.

Suppression of Type I Interferon Production by Human T-Cell Leukemia Virus Type 1 Oncoprotein Tax through Inhibition of IRF3 Phosphorylation

ABSTRACT Infection with human T-cell leukemia virus type 1 (HTLV-1) is associated with adult T-cell leukemia (ATL) and tropical spastic paraparesis. Type I interferons (IFNs) are key effectors of the innate antiviral response, and IFN-α combined with the nucleoside reverse transcriptase inhibitor zidovudine is considered the standard first-line therapy for ATL. HTLV-1 oncoprotein Tax is known to suppress innate IFN production and response but the underlying mechanisms remain to be fully established. In this study, we report on the suppression of type I IFN production by HTLV-1 Tax through interaction with and inhibition of TBK1 kinase that phosphorylates IRF3. Induced transcription of IFN-β was severely impaired in HTLV-1-transformed ATL cells and freshly infected T lymphocytes. The ability to suppress IRF3 activation was ascribed to Tax. The expression of Tax alone sufficiently repressed the induction of IFN production by RIG-I plus PACT, cGAMP synthase plus STING, TBK1, IKKε, IRF3, and IRF7, but not by IRF3-5D, a dominant-active phosphomimetic mutant. This suggests that Tax perturbs IFN production at the step of IRF3 phosphorylation. Tax mutants deficient for CREB or NF-κB activation were fully competent in the suppression of IFN production. Coimmunoprecipitation experiments confirmed the association of Tax with TBK1, IKKε, STING, and IRF3. In vitro kinase assay indicated an inhibitory effect of Tax on TBK1-mediated phosphorylation of IRF3. Taken together, our findings suggested a new mechanism by which HTLV-1 oncoprotein Tax circumvents the production of type I IFNs in infected cells. Our findings have implications in therapeutic intervention of ATL. IMPORTANCE Human T-cell leukemia virus type 1 (HTLV-1) is the cause of adult T-cell leukemia (ATL), an aggressive and fatal blood cancer, as well as another chronic disabling disease of the spinal cord. Treatments are unsatisfactory, and options are limited. A combination of antiviral cellular protein alpha interferon and zidovudine, which is an inhibitor of a viral enzyme called reverse transcriptase, has been recommended as the standard first-line therapy for ATL. Exactly how HTLV-1 interacts with the cellular machinery for interferon production and action is not well understood. Our work sheds light on the mechanism of action for the inhibition of interferon production by an HTLV-1 oncogenic protein called Tax. Our findings might help to improve interferon-based anti-HTLV-1 and anti-ATL therapy.

Group I p21-activated kinases facilitate Tax-mediated transcriptional activation of the human T-cell leukemia virus type 1 long terminal repeats

BackgroundHuman T-cell leukemia virus type 1 (HTLV-1) is the causative agent of adult T-cell leukemia and tropical spastic paraparesis. HTLV-1 encodes transactivator protein Tax that interacts with various cellular factors to modulate transcription and other biological functions. Additional cellular mediators of Tax-mediated transcriptional activation of HTLV-1 long terminal repeats (LTR) remain to be identified and characterized.ResultsIn this study, we investigated the regulatory role of group I p21-activated kinases (Paks) in Tax-induced LTR activation. Both wild-type and kinase-dead mutants of Pak3 were capable of potentiating the activity of Tax to activate LTR transcription. The effect of Paks on the LTR was attributed to the N-terminal regulatory domain and required the action of CREB, CREB-regulating transcriptional coactivators (CRTCs) and p300/CREB-binding protein. Paks physically associated with Tax and CRTCs. Paks were recruited to the LTR in the presence of Tax. siRNAs against either Pak1 or Pak3 prevented the interaction of Tax with CRTC1 and the recruitment of Tax to the LTR. These siRNAs also inhibited LTR-dependent transcription in HTLV-1-transformed MT4 cells and in cells transfected with an infectious clone of HTLV-1.ConclusionGroup I Paks augment Tax-mediated transcriptional activation of HTLV-1 LTR in a kinase-independent manner.

Severe acute respiratory syndrome coronavirus nucleocapsid protein does not modulate transcription of the human FGL2 gene.

Among the structural and nonstructural proteins of severe acute respiratory syndrome coronavirus (SARS-CoV), the nucleocapsid (N) protein plays pivotal roles in the biology and pathogenesis of viral infection. N protein is thought to dysregulate cell signalling and the transcription of cellular genes, including FGL2, which encodes a prothrombinase implicated in vascular thrombosis, fibrin deposition and pneumocyte necrosis. Here, we showed that N protein expressed in cultured human cells was predominantly found in the cytoplasm and was competent in repressing the transcriptional activity driven by interferon-stimulated response elements. However, the expression of N protein did not influence the transcription from the FGL2 promoter. More importantly, N protein did not modulate the expression of FGL2 mRNA or protein in transfected or SARS-CoV-infected cells. Taken together, our findings did not support the model in which SARS-CoV N protein specifically modulates transcription of the FGL2 gene to cause fibrosis and vascular thrombosis.

Inhibition of AIM2 inflammasome activation by a novel transcript isoform of IFI16

Mouse p202 is a disease locus for lupus and a dominant‐negative inhibitor of AIM2 inflammasome activation. A human homolog of p202 has not been identified so far. Here, we report a novel transcript isoform of human IFI16‐designated IFI16‐β, which has a domain architecture similar to that of mouse p202. Like p202, IFI16‐β contains two HIN domains, but lacks the pyrin domain. IFI16‐β is ubiquitously expressed in various human tissues and cells. Its mRNA levels are also elevated in leukocytes of patients with lupus, virus‐infected cells, and cells treated with interferon‐β or phorbol ester. IFI16‐β co‐localizes with AIM2 in the cytoplasm, whereas IFI16‐α is predominantly found in the nucleus. IFI16‐β interacts with AIM2 to impede the formation of a functional AIM2‐ASC complex. In addition, IFI16‐β sequesters cytoplasmic dsDNA and renders it unavailable for AIM2 sensing. Enforced expression of IFI16‐β inhibits the activation of AIM2 inflammasome, whereas knockdown of IFI16‐β augments interleukin‐1β secretion triggered by dsDNA but not dsRNA. Thus, cytoplasm‐localized IFI16‐β is functionally equivalent to mouse p202 that exerts an inhibitory effect on AIM2 inflammasome.