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The E5 protein of the human papillomavirus type 16 down-regulates HLA-I surface expression in calnexin-expressing but not in calnexin-deficient cells.

Gruener M, Bravo IG, Momburg F, Alonso A, Tomakidi P - Virol. J. (2007)

Bottom Line: The molecular mechanisms underlying this effect are so far unknown.In addition, we show that the M1 mutant is only able to marginally down-regulate HLA-I surface expression compared to the wild-type protein.On the basis of our results we conclude that formation of this complex is responsible for retention of HLA-I molecules in the ER of the cells.

View Article: PubMed Central - HTML - PubMed

Affiliation: Division of Cell Differentiation, German Cancer Research Center, Heidelberg, Germany. m.gruener@dkfz.de

ABSTRACT
The human papillomavirus type 16 E5 protein (HPV16 E5) down-regulates surface expression of HLA-I molecules. The molecular mechanisms underlying this effect are so far unknown. Here we show that HPV16 E5 down-regulates HLA-I surface expression in calnexin-containing but not in calnexin-deficient cells. Immunoprecipitation experiments reveal that calnexin and HPV16E5 can be co-precipitated and that this association depends on the presence of a wild-type first hydrophobic region of E5. When an E5 mutant (M1) in which the first putative transmembrane helix had been disrupted was used for the transfections calnexin-E5 co-precipitation was strongly impaired. In addition, we show that the M1 mutant is only able to marginally down-regulate HLA-I surface expression compared to the wild-type protein. Besides, we demonstrate that E5 forms a ternary complex with calnexin and the heavy chain of HLA-I, which is mediated by the first hydrophobic region of the E5 protein. On the basis of our results we conclude that formation of this complex is responsible for retention of HLA-I molecules in the ER of the cells.

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HPV16 E5, M2 and M3 mutants but not M1 mutant strongly co-localize with calnexin. HaCaT cells were transfected with A) AU1-tagged codon-optimised E5 or AU1- tagged codonoptimised E5 mutants M1 B), M2 C), and M3 D) and analysed after 24 h by confocal laser scanning microscopy using a monoclonal anti-AU1 and polyclonal anticalnexin antibodies.
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Figure 8: HPV16 E5, M2 and M3 mutants but not M1 mutant strongly co-localize with calnexin. HaCaT cells were transfected with A) AU1-tagged codon-optimised E5 or AU1- tagged codonoptimised E5 mutants M1 B), M2 C), and M3 D) and analysed after 24 h by confocal laser scanning microscopy using a monoclonal anti-AU1 and polyclonal anticalnexin antibodies.

Mentions: As shown in Fig. 8, calnexin colocalized with the E5 protein expressed from the codonoptimized gene (Fig. 8A), as well as with the M2 and M3 mutants (Fig. 8C and 8D). In contrast, the disruption of the first helix in mutant M1 results in a change in the subcellular localisation of the protein, yielding a disperse and punctuate subcellular distribution, where only a partial co-localization with calnexin (Fig. 8B). These results are consistent with those found in the immunoprecipitation experiments and further confirm that the interaction of HPV16 E5 and calnexin requires a native, non-modified first transmembrane domain of the viral protein.


The E5 protein of the human papillomavirus type 16 down-regulates HLA-I surface expression in calnexin-expressing but not in calnexin-deficient cells.

Gruener M, Bravo IG, Momburg F, Alonso A, Tomakidi P - Virol. J. (2007)

HPV16 E5, M2 and M3 mutants but not M1 mutant strongly co-localize with calnexin. HaCaT cells were transfected with A) AU1-tagged codon-optimised E5 or AU1- tagged codonoptimised E5 mutants M1 B), M2 C), and M3 D) and analysed after 24 h by confocal laser scanning microscopy using a monoclonal anti-AU1 and polyclonal anticalnexin antibodies.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2164959&req=5

Figure 8: HPV16 E5, M2 and M3 mutants but not M1 mutant strongly co-localize with calnexin. HaCaT cells were transfected with A) AU1-tagged codon-optimised E5 or AU1- tagged codonoptimised E5 mutants M1 B), M2 C), and M3 D) and analysed after 24 h by confocal laser scanning microscopy using a monoclonal anti-AU1 and polyclonal anticalnexin antibodies.
Mentions: As shown in Fig. 8, calnexin colocalized with the E5 protein expressed from the codonoptimized gene (Fig. 8A), as well as with the M2 and M3 mutants (Fig. 8C and 8D). In contrast, the disruption of the first helix in mutant M1 results in a change in the subcellular localisation of the protein, yielding a disperse and punctuate subcellular distribution, where only a partial co-localization with calnexin (Fig. 8B). These results are consistent with those found in the immunoprecipitation experiments and further confirm that the interaction of HPV16 E5 and calnexin requires a native, non-modified first transmembrane domain of the viral protein.

Bottom Line: The molecular mechanisms underlying this effect are so far unknown.In addition, we show that the M1 mutant is only able to marginally down-regulate HLA-I surface expression compared to the wild-type protein.On the basis of our results we conclude that formation of this complex is responsible for retention of HLA-I molecules in the ER of the cells.

View Article: PubMed Central - HTML - PubMed

Affiliation: Division of Cell Differentiation, German Cancer Research Center, Heidelberg, Germany. m.gruener@dkfz.de

ABSTRACT
The human papillomavirus type 16 E5 protein (HPV16 E5) down-regulates surface expression of HLA-I molecules. The molecular mechanisms underlying this effect are so far unknown. Here we show that HPV16 E5 down-regulates HLA-I surface expression in calnexin-containing but not in calnexin-deficient cells. Immunoprecipitation experiments reveal that calnexin and HPV16E5 can be co-precipitated and that this association depends on the presence of a wild-type first hydrophobic region of E5. When an E5 mutant (M1) in which the first putative transmembrane helix had been disrupted was used for the transfections calnexin-E5 co-precipitation was strongly impaired. In addition, we show that the M1 mutant is only able to marginally down-regulate HLA-I surface expression compared to the wild-type protein. Besides, we demonstrate that E5 forms a ternary complex with calnexin and the heavy chain of HLA-I, which is mediated by the first hydrophobic region of the E5 protein. On the basis of our results we conclude that formation of this complex is responsible for retention of HLA-I molecules in the ER of the cells.

Show MeSH
Related in: MedlinePlus