PMC:7464876 / 31549-34291
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{"target":"http://pubannotation.org/docs/sourcedb/PMC/sourceid/7464876","sourcedb":"PMC","sourceid":"7464876","source_url":"https://www.ncbi.nlm.nih.gov/pmc/7464876","text":"3.4. HBV L Colocalizes with ERGIC-53 upon ER Export Block\nGiven that the Sec24A isoform is the essential COPII component linked to the export of both the viral and subviral HBV envelope, we next investigated the fate of the ER-arrested envelopes due to Sec24A KD. To study spherical SVP trafficking, HuH-7 cells were transiently transfected with an expression construct encoding only the S envelope gene (HBV.S) (Figure 4A). Notably, this is an established approach to study exclusively the fate of subviral HBV spheres [29]. Consistent with our previous IF analysis [29] and for recapitulation herein, HBV.S showed a diffuse granular staining pattern typical for viral surface proteins that are synthesized at the ER (Figure 4B). Moreover, it largely colocalized with ERGIC-53-stained structures, implicating that HBV.S is transported out of the ER to the ERGIC in control cells (Figure 4B). Upon Sec24A inactivation, the staining pattern of HBV.S strongly changed, as it now appeared in dot-like structures, reminiscent of ER-arrested aggregates, that no longer coincided with ERGIC-53 signals (Figure 4B). Notably, under these conditions, the distribution of ERGIC-53 also altered. In normal cells, this protein predominantly localizes to the ERGIC compartment, as displayed by the given name ERGIC-53. It is a high mannose-specific lectin that cycles cargoes between the ER and the cis-Golgi through COPII and COPI-dependent pathways (see also Figure 5A) [49,50]. Upon immunostaining, it appeared in juxtanuclear, tubulovesicular clusters in control cells, which shifted to a perinuclear, ER-like structure in Sec24A KD cells (Figure 4B). Hence, the ER export of both HBV.S and ERGIC-53 is apparently impaired in Sec24A-silenced cells.\nIn contrast, the HBV L protein, synthesized in HBV-replicating cells, could not be traced within the ERGIC, as no colocalization between L and ERGIC-53 was detectable in the control cells (Figure 4C). In further difference to spherical SVPs, the naturally occurring structure of L was preserved in Sec24A KD cells without any signs of aggregation. Quite unexpectedly, in this setting, ERGIC-53 intensively colocalized with L, implicating that the lectin is seemingly entrapped by the L-specific CS structure if arrested in the ER. Similar staining experiments were done in cells depleted for the related Sec24B isoform. However, this intervention did not affect the typical juxtanuclear distribution of ERGIC-53 and, consequently, did not result in ERGIC-53/L colocalization (Figure 4C). We infer from these results that ERGIC-53 may be a preferred cargo of Sec24A rather than of Sec24B. If arrested in the ER, ERGIC-53 tightly associated with the HBV L envelope protein either by chance or for action.","divisions":[{"label":"title","span":{"begin":0,"end":57}},{"label":"p","span":{"begin":58,"end":1739}}],"tracks":[{"project":"2_test","denotations":[{"id":"32806600-32017353-20296218","span":{"begin":521,"end":523},"obj":"32017353"},{"id":"32806600-32017353-20296219","span":{"begin":568,"end":570},"obj":"32017353"},{"id":"32806600-9788882-20296220","span":{"begin":1460,"end":1462},"obj":"9788882"},{"id":"32806600-19609866-20296221","span":{"begin":1463,"end":1465},"obj":"19609866"}],"attributes":[{"subj":"32806600-32017353-20296218","pred":"source","obj":"2_test"},{"subj":"32806600-32017353-20296219","pred":"source","obj":"2_test"},{"subj":"32806600-9788882-20296220","pred":"source","obj":"2_test"},{"subj":"32806600-19609866-20296221","pred":"source","obj":"2_test"}]}],"config":{"attribute types":[{"pred":"source","value type":"selection","values":[{"id":"2_test","color":"#93ecb4","default":true}]}]}}