Abstract
Apobec-1 is the catalytic subunit of a multicomponent editosome complex that mediates apolipoprotein B (apoB) mRNA editing. We isolated a novel apobec-1-interacting protein by yeast two-hybrid cloning and identified the protein as BAG-4. BAG-4, a chaperone-regulating protein, also known as SODD (silencer of death domains), is a member of the BAG family of proteins. In this report, we found that apobec-1 is localized in the perinucleolar compartment in HepG2 cells and rat liver MCR-RH7777 cells. BAG-4 binds to apobec-1 via its N-terminal region independent of the BAG domain. It is ubiquitously expressed with predominant occurrence in human pancreas, heart, brain, and placenta. Immunoprecipitation experiments confirmed that BAG-4 interacts with Hsc70/Hsp90 in HepG2 cells. BAG-4 tagged with green fluorescent protein (GFP) or FLAG was localized both in cytoplasm of mouse BNLCL.2 liver cells and human liver hepatoma HepG2 cells. After heat shock, GFP-BAG-4 co-localizes with Hsc70 in the nucleus in HepG2 cells, whereas GFP-BAG-4 mutants lacking the BAG domain remain perinuclear. BAG-4 has no effects on apoB mRNA editing in vitro. However, unlike other apobec-1 complementation factors studied to date, antisense knockdown of BAG-4 in BNLCL.2 cells and in MCR-RH7777 cells increases rather than decreases endogenous apoB mRNA editing. Overexpression of BAG-4 in MCR-RH7777 cells also suppresses apoB mRNA editing. ApoB-48 production also increases with antisense BAG-4 expression in MCR-RH7777 cells. We previously showed that apoB mRNA editing is an intranuclear event (Lau, P. P., Xiong, W. J., Zhu, H. J., Chen, S. H., and Chan, L. (1991) J. Biol. Chem. 266, 20550-20554). Thus, BAG-4 overexpression down-regulates apoB mRNA editing by shuttling apobec-1 from the intranuclear perinucleolar compartment to the cytoplasm. We propose that BAG-4 functions as a negative regulator for apobec-1-mediated apoB mRNA editing through its ability to suppress the Hsp/Hsc70 chaperone activity and thereby editosome formation and, as a consequence, prevents nuclear localization of the apobec-1 editosome.
Highlights
In the primary structure of apobec-1 that have been postulated to be important in its distribution in both the nucleus and the cytoplasm
We propose that BAG-4 functions as a negative regulator for apobec-1-mediated apolipoprotein B (apoB) mRNA editing through its ability to suppress the Hsp/Hsc70 chaperone activity and thereby editosome formation and, as a consequence, prevents nuclear localization of the apobec-1 editosome
We found that down-regulation of ABBP-2 abolished endogenous apoB mRNA editing in apobec-1-expressing HepG2 cells and mouse liver BNLCL.2 cells and that ABBP-2 works in concert with Hsc70 [5]
Summary
In the primary structure of apobec-1 that have been postulated to be important in its distribution in both the nucleus and the cytoplasm None of these experimentally meet the functional criteria of nuclear localization or nuclear export signals. Fifteen amino acids (Leu173–Leu187) within the C-terminal domain of apobec-1 were sufficient as a determinant for cytoplasmic distribution. These residues failed to demonstrate nuclear export function in a reporter assay. We identify BAG-4 as a regulator of apoB mRNA editing by its interaction with apobec-1 in association with the chaperone, Hsc. The antiapoptotic activities of BAG family proteins may be a result of their interactions with Hsc70/Hsp and/or binding to Bcl-2. We demonstrate how BAG-4 interacts with Hsc to modulate apoB mRNA editing
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