HCMV-encoded viral protein US12 promotes autophagy by inducing autophagy flux.

The human cytomegalovirus (HCMV)-encoded US12 gene family is a group of ten predicted seven-transmembrane domain proteins that are structurally similar to G-protein-coupled receptors or transmembrane Bax inhibitor-1 motif-containing proteins; however, the roles of US12 family proteins in virus-host interactions remain to be discovered. Here, we suggest a new function of the US12 protein in regulating cellular autophagy. US12 is predominantly located to the lysosome and interacts with the lysosomal membrane protein 2 (LAMP2). A liquid chromatography-mass spectrometry (MS)/MS-based targeted proteomics analysis shows that US12 is tightly correlated with autophagy. US12 induces autophagy via upregulating ULK1 phosphorylation and subsequent LC3-II conversion, thereby accelerating autophagic flux. Moreover, HeLa cells overexpressing US12 displays intense LC3-specific staining and autolysosome formation even under nutrient-sufficient conditions. Furthermore, the physical interaction of p62/SQSTM1 with US12 is involved in the resistance to the degradation of p62/SQSTM1 by autophagy, despite the induction of both autolysosome formation and autophagic flux. Although the effect of US12 expression in HCMV infection on autophagy remains undetermined, these findings provide new insights into the viral drivers of host autophagy during HCMV evolution and pathogenesis.

In vitro development of a hemangioblast from a human embryonic stem cell, SNUhES#3.

AIMS: Recent reports demonstrated that a hemangioblast population emerged during hematopoietic development in both mouse and human embryonic stem cell (hESC) differentiation cultures. MAIN METHODS: In this study, a new uncharacterized hESC line, SNUhES#3, was studied for its capacity to proliferate with STO cells and differentiate into hemangioblasts in co-culture with OP9 cells. KEY FINDINGS: We were able to obtain CD34(+)CD45(-) cells from SNUhES#3 cells after 12 days of in vitro culture, and this cell population could be maximized to 12.6% of the total. These cells, derived from SNUhES#3, showed the morphology of hematopoietic precursor cells and endothelial lineage cells with high efficiency. Reverse transcription polymerase chain reaction (RT-PCR) analysis showed that the hematopoietic markers CD34, GATA2, and LMO2 were co-expressed with the endothelial marker CD31 from day 8, whereas ES cell marker OCT4 no longer existed at an early stage. Moreover, we found that the efficacy of colony forming by SNUhES#3 cells is better than that of H9 cells. SIGNIFICANCE: These findings provide evidence that SNUhES#3 cells can be used as an established human ESC line, and co-culture with OP9 can induce SNUhES#3 cells to differentiate into hemangioblasts, the common precursors of the hematopoietic and endothelial lineages.

HS 1-associated protein X-1 is cleaved by caspase-3 during apoptosis.

Caspase-3 (CASP3) plays a key role in apoptosis. In this study, HAX-1 was identified as a new substrate of CASP3 during apoptosis. HAX-1 was cleaved by CASP3 during etoposide-(ETO) induced apoptosis, and this event was inhibited by a CASP3-specific inhibitor. The cleavage site of HAX-1, at Asp(127), was located using N-terminal amino acid sequencing of in vitro cleavage products of recombinant HAX-1. Overexpression of HAX-1 inhibited ETO-induced apoptotic cell death. It also inhibited CASP3 activity. Together, these results suggest that HAX-1, a substrate of CASP3, inhibits the apoptotic process by inhibiting CASP3 activity.