Severe acute respiratory syndrome coronavirus 3a protein activates the mitochondrial death pathway through p38 MAP kinase activation

Abstract

The molecular mechanisms governing severe acute respiratory syndrome coronavirus-induced pathology are not fully understood. Virus infection and some individual viral proteins, including the 3a protein, induce apoptosis. However, the cellular targets leading to 3a protein-mediated apoptosis have not been fully characterized. This study showed that the 3a protein modulates the mitochondrial death pathway in two possible ways. Activation of caspase-8 through extrinsic signal(s) caused Bid activation. In the intrinsic pathway, there was activation of caspase-9 and cytochrome c release from the mitochondria. This was the result of increased Bax oligomerization and higher levels of p53 in 3a protein-expressing cells, which depended on the activation of p38 MAP kinase (MAPK) in these cells. For p38 activation and apoptosis induction, the 3a cytoplasmic domain was sufficient. In direct Annexin V staining assays, the 3a protein-expressing cells showed increased apoptosis that was attenuated with the p38 MAPK inhibitor SB203580. A block in nuclear translocation of the STAT3 transcription factor in cells expressing the 3a protein was also observed. These results have been used to present a model of 3a-mediated apoptosis. INTRODUCTIONThe aetiological agent for severe acute respiratory syndrome (SARS) was identified as a novel coronavirus (SARS-CoV) (Peiris et al., 2003). SARS-CoV has a polyadenylated, positive-sense RNA genome of approximately 30 kb (Marra et al., 2003). In addition to the prototypic coronavirus genes, the SARS-CoV genome also contains nine unique open reading frames (ORFs) (Marra et al., 2003). Of these, orf3a is the largest and encodes a protein of 274 aa, variously called ORF3A (Ito et al., 2005), X1 (Rota et al., 2003) or U274 (Tan et al., 2004b). The 3a protein has been predicted to contain an N-terminal signal peptide followed by three transmembrane domains and a C-terminal cytoplasmic domain of approximately 150 aa (Zeng et al., 2004).The 3a protein is associated with virus particles produced following infection of Vero E6 or Caco-2 cells (Ito et al., 2005;Shen et al., 2005) and can assemble into virus-like particles when co-expressed with the membrane and envelope proteins in insect cells (Shen et al., 2005). In vitro studies have also shown the 3a protein to interact with the viral envelope, membrane and spike proteins (Tan et al., 2004b;Tan, 2005) and the cellular protein caveolin-1 (Padhan et al., 2007). A deletion of orf3a was shown to reduce virus titres, but not to eliminate virus replication (Yount et al., 2005), and convalescent sera from SARS patients have antibodies to the 3a protein (Tan et al., 2004a), suggesting that it is expressed during virus infection of the host. The 3a protein was shown to upregulate the expression of fibrinogen in A549 lung epithelial cells and to possess an ionchannel activity selective for monovalent cations (Lu et al., 2006). Ectopic expression of the 3a protein has been shown to induce apoptosis in Vero E6 cells through activation of caspase-8, chromatin c...

JNK and p38 mitogen-activated protein kinase pathways contribute to porcine epidemic diarrhea virus infection

“…Apoptotic cell death is known to be modulated by the p38 MAPK and JNK cellular stress pathways (Kyriakis and Avruch, 2001;Wada and Penninger, 2004). Several viruses can trigger activation of p38 MAPK and/or JNK, which mediates apoptosis via either intrinsic or extrinsic pathways, and thereby contributes to viral replication (Padhan et al, 2008;Sumbayev and Yasinska, 2006;Wang et al, 2004;. We previously demonstrated that PEDV promotes apoptotic cell death in vitro and in vivo to support viral infection and pathogenesis (Kim and Lee, 2014).…”

Accessory proteins of SARS-CoV and other coronaviruses

“…The long-term triggering of the PERK pathway can also induce virus-related apoptosis through the expression of its downstream mediators of ATF4 and CHOP, similar to IBV infection (Liao et al, 2013). Padhan et al have shown that protein 3a can lead to increased activation of the p38 MAP kinase pathway and induce the mitochondria to leak cytochrome c to induce apoptosis (Padhan et al, 2008).…”

Viroporins: structure, function and potential as antiviral targets

“…3a mediates the production of infectious viral progeny, potentially linked to cellular trafficking of the spike glycoprotein (Tan, 2005), but has also been proposed to comprise a structural component of the infectious virion (Shen et al, 2005). 3a is pro-apoptotic in a number of cell lines, which appears directly dependent upon channel function, and may be linked to the induction of an ER stress response (Chan et al, 2009;Freundt et al, 2010;Law et al, 2005;Minakshi et al, 2009;Padhan et al, 2008). Two studies have reported small-molecule inhibitors targeting 3a.…”