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Overexpression of micro RNA ‐30a inhibits hepatitis B virus X protein‐induced autophagosome formation in hepatic cells
Author(s) -
Kumar Satendra,
Gupta Parul,
Khanal Sweta,
Shahi Aashirwad,
Kumar Pushpendra,
Sarin Shiv K.,
Venugopal Senthil K.
Publication year - 2015
Publication title -
the febs journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.981
H-Index - 204
eISSN - 1742-4658
pISSN - 1742-464X
DOI - 10.1111/febs.13209
Subject(s) - transfection , rna , microbiology and biotechnology , autophagosome , tunel assay , biology , apoptosis , messenger rna , autophagy , terminal deoxynucleotidyl transferase , cell culture , gene , biochemistry , genetics
Hepatitis B virus ( HBV ) enters the host and survives by using several mechanisms. One of the ways that HBV survives and replicates in the host cells is by inducing autophagy. Previous reports have shown that micro RNA (mi RNA )‐30a inhibits autophagosome formation in cancer cells. Hence, we hypothesized that overexpression of mi RNA ‐30a could inhibit HBV ‐induced autophagosome formation in hepatic cells. To study this, both HepG2 cells and HepG2.2.1.5 cells ( HBV ‐expressing stable cell line) were transfected with mi RNA ‐30a, and the cells were collected either for RNA isolation or protein isolation after 72 h of transfection. Beclin‐1 expression was significantly higher in untransfected HepG2.2.1.5 cells than in HepG2 cells. Western blots showed that mi RNA ‐30a overexpression resulted in a significant decrease in beclin‐1 expression (eight‐fold and four‐fold in HepG2 and HepG2.2.1.5 cells, respectively) and c‐myc expression, whereas the numbers of terminal deoxynucleotidyl transferase‐mediated dUTP nick end labeling ( TUNEL )‐positive cells were increased. In contrast, overexpression of HBV X protein ( HB x) in HepG2 cells resulted in the enhancement of beclin‐1 (six‐fold increase as compared with the empty vector‐transfected cells) and c‐myc expression, whereas the numbers of TUNEL ‐positive cells were reduced. To confirm these findings, HB x and mi RNA ‐30a were coexpressed in HepG2 cells, and the results showed significant inhibition of autophagosome formation and beclin‐1 and c‐myc expression, whereas apoptosis increased. These data demonstrate that HB x induces autophagosome formation via beclin‐1 expression, whereas mi RNA ‐30a overexpression could successfully inhibit the beclin‐1 expression induced by HB x, thereby modulating autophagosome formation in hepatic cells.