PMC:7464876 / 41839-43693
Annnotations
{"target":"http://pubannotation.org/docs/sourcedb/PMC/sourceid/7464876","sourcedb":"PMC","sourceid":"7464876","source_url":"https://www.ncbi.nlm.nih.gov/pmc/7464876","text":"Several studies have shown that HBV budding and egress require functions of the ESCRT complex, a membrane scission machinery operating at several cellular membranes [20,21,22,23,24]. To examine whether the ER-related, core-recruiting CS structure of L might have a link to ESCRT, we inspected the intracellular distribution of the Vps4 ATPase, the terminal ESCRT subunit [25], in HBV-replicating and control HuH-7 cells. To this aim, cells were transfected with an EGFP-tagged version of Vps4A together with either empty plasmid DNA or the HBV replicon and processed for IF analysis. In control cells, Vps4A was distributed throughout the cytoplasm, along with some nuclear localization (Figure 9A). Conversely, in HBV-positive cells, the distribution of Vps4A changed in such that it now additionally appeared in the CS structure reminiscent for HBV (Figure 9A). The apparent recruitment of Vps4A to the L/core complex suggested an ESCRT intervention occurring at the ER-related CS structure. To further investigate the spatial and temporal interconnections between HBV and the ESCRT cascade, we reasoned to study the N-glycan pattern of L in order to learn whether the HBV budding driving activity of ESCRT occurred before or after its acquisition of EndoH resistance. For ESCRT inactivation in HBV-replicating HuH-7 cells, we silenced the expression of EAP20, one subunit of the heterotetrameric ESCRT-II complex. Consistent with our previous report [22], this led to a substantial decline of extracellular virion amounts (Figure 9B). In parallel, cellular lysates were mock-treated or digested with EndoH and assayed by L-specific WB. Upon ESCRT-II inactivation, cell-associated L displayed the same EndoH sensitivity as observed in siCon-treated cells (Figure 9B), implicating that EAP20 is acting prior to L arrival at the cis/medial Golgi complex.","tracks":[{"project":"2_test","denotations":[{"id":"32806600-17553870-20296236","span":{"begin":166,"end":168},"obj":"17553870"},{"id":"32806600-17551004-20296237","span":{"begin":169,"end":171},"obj":"17551004"},{"id":"32806600-24614091-20296238","span":{"begin":172,"end":174},"obj":"24614091"},{"id":"32806600-26719264-20296239","span":{"begin":175,"end":177},"obj":"26719264"},{"id":"32806600-26431433-20296240","span":{"begin":178,"end":180},"obj":"26431433"},{"id":"32806600-24614091-20296241","span":{"begin":1454,"end":1456},"obj":"24614091"}],"attributes":[{"subj":"32806600-17553870-20296236","pred":"source","obj":"2_test"},{"subj":"32806600-17551004-20296237","pred":"source","obj":"2_test"},{"subj":"32806600-24614091-20296238","pred":"source","obj":"2_test"},{"subj":"32806600-26719264-20296239","pred":"source","obj":"2_test"},{"subj":"32806600-26431433-20296240","pred":"source","obj":"2_test"},{"subj":"32806600-24614091-20296241","pred":"source","obj":"2_test"}]}],"config":{"attribute types":[{"pred":"source","value type":"selection","values":[{"id":"2_test","color":"#ec93b2","default":true}]}]}}