原创力文档a putative diacidic motif in the sars-cov orf6 protein influences its subcellular localization and suppression of expression of co-transfected expression constructs冠的假定的diacidic主题orf6蛋白质影响表达的亚细胞定位和抑制co-transfected表达结构.pdfVIP
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a putative diacidic motif in the sars-cov orf6 protein influences its subcellular localization and suppression of expression of co-transfected expression constructs冠的假定的diacidic主题orf6蛋白质影响表达的亚细胞定位和抑制co-transfected表达结构
Gunalan et al. BMC Research Notes 2011, 4:446
/1756-0500/4/446
RESEARCH ARTICLE Open Access
A putative diacidic motif in the SARS-CoV ORF6
protein influences its subcellular localization and
suppression of expression of co-transfected
expression constructs
Vithiagaran Gunalan 1,2,3, Ali Mirazimi1,2 and Yee-Joo Tan4*
Abstract
Background: The ORF6 protein is one of the eight accessory proteins of the severe acute respiratory syndrome
coronavirus (SARS-CoV). Numerous properties of ORF6 have been documented and this study focuses on two of
these, namely, its ability to suppress the expression of co-transfected expression constructs and its subcellular
localization to vesicular structures.
Results: Using a transient transfection system, ORF6’s ability to suppress the expression of co-transfected
expression constructs was measured in a quantitative manner. While ORF6 does not have a global effect on
protein synthesis, quantitative real-time PCR revealed that it down-regulated the mRNA level of the co-transfected
myc-nsp8 gene. Furthermore, alanine substitution of a diacidic cluster motif (aa53-56) in the ORF6 gene caused a
reduction in the suppression of expression of co-transfected myc-nsp8 gene. Our previous study revealed that
ORF6 localized to vesicular structures in SARS-CoV infected Vero E6 cells. Here, ORF6 was observed to be localized
to similar vesicular structures in Vero E6 cells which have been transiently transfected with a mammalian
expression plasmid encoding for untagged ORF6. ORF6 showed partial colocalization with cellular proteins CD63
and Lamp1, suggesting that the vesicular structures may be a subpopulation of endosomal/lysosomal vesicles. The
alanine substitution of the diacidic cluster motif also altered the subcellular localization of the ORF6 protein,
indicating a potential relationship between t
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