July 1, 2004 Journal article Closed Access

Shweta Tyagi; Milan Surjit; Shahid Jameel; Anindita Kar Roy; Sunil K. Lal

MARC21 XML Export

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    <subfield code="a">Hepatitis E virus (HEV), a plus-stranded RNA virus contains three open reading frames. Of these, ORF1 encodes the viral nonstructural polyprotein; ORF2 encodes the major capsid protein and ORF3 codes for a phosphoprotein of undefined function. Using the yeast two-hybrid system to screen a human cDNA liver library we have isolated, an N-terminal deleted protein, alpha(1) -microglobulin/bikunin precursor (AMBP) that specifically interacts with the ORF3 protein of HEV. Independently cloned, full-length AMBP was obtained and tested positive for interaction with ORF3 using a variety of in vivo and in vitro techniques. AMBP, a liver-specific precursor protein codes for two different unrelated proteins alpha(1)-microglobulin (alpha(1)m) and bikunin. alpha(1) m individually interacted with ORF3. The above findings were validated by COS-1 cell immunoprecipitation, His(6) pull-down experiments, and co-localization experiments followed by fluorescence resonance energy transfer analysis. Human liver cells showing co-localization of ORF3 with endogenously expressing alpha(1) m showed a distinct disappearance of the protein from the Golgi compartment, suggesting that ORF3 enhances the secretion of alpha(1)m out of the hepatocyte. Using drugs to block the secretory pathway, we showed that alpha m was not degraded in the presence of ORF3. Finally, (1)pulse labeling of alpha(1)m showed that its secretion was expedited out of the liver cell at faster rates in the presence of the ORF3 protein. Hence, ORF3 has a direct biological role in enhancing alpha(1)m export from the hepatocyte.</subfield>
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    <subfield code="a">Milan Surjit</subfield>
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    <subfield code="a">Anindita Kar Roy</subfield>
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    <subfield code="a">Sunil K. Lal</subfield>
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    <subfield code="a">10.1074/jbc.m402017200</subfield>
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    <subfield code="a">Cell Biology</subfield>
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    <subfield code="a">Shweta Tyagi</subfield>
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    <subfield code="a">The ORF3 Protein of Hepatitis E Virus Interacts with Liver-specific α1-Microglobulin and Its Precursor α1-Microglobulin/Bikunin Precursor (AMBP) and Expedites Their Export from the Hepatocyte</subfield>
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