An ectromelia virus profilin homolog interacts with cellular tropomyosin and viral A-type inclusion protein.

Abstract

BACKGROUND:

Profilins are critical to cytoskeletal dynamics in eukaryotes; however, little is known about their viral counterparts. In this study, a poxviral profilin homolog, ectromelia virus strain Moscow gene 141 (ECTV-PH), was investigated by a variety of experimental and bioinformatics techniques to characterize its interactions with cellular and viral proteins.

RESULTS:

Profilin-like proteins are encoded by all orthopoxviruses sequenced to date, and share over 90% amino acid (aa) identity. Sequence comparisons show highest similarity to mammalian type 1 profilins; however, a conserved 3 aa deletion in mammalian type 3 and poxviral profilins suggests that these homologs may be more closely related. Structural analysis shows that ECTV-PH can be successfully modelled onto both the profilin 1 crystal structure and profilin 3 homology model, though few of the surface residues thought to be required for binding actin, poly(L-proline), and PIP2 are conserved. Immunoprecipitation and mass spectrometry identified two proteins that interact with ECTV-PH within infected cells: alpha-tropomyosin, a 38 kDa cellular actin-binding protein, and the 84 kDa product of vaccinia virus strain Western Reserve (VACV-WR) 148, which is the truncated VACV counterpart of the orthopoxvirus A-type inclusion (ATI) protein. Western and far-western blots demonstrated that the interaction with alpha-tropomyosin is direct, and immunofluorescence experiments suggest that ECTV-PH and alpha-tropomyosin may colocalize to structures that resemble actin tails and cellular protrusions. Sequence comparisons of the poxviral ATI proteins show that although full-length orthologs are only present in cowpox and ectromelia viruses, an ~ 700 aa truncated ATI protein is conserved in over 90% of sequenced orthopoxviruses. Immunofluorescence studies indicate that ECTV-PH localizes to cytoplasmic inclusion bodies formed by both truncated and full-length versions of the viral ATI protein. Furthermore, colocalization of ECTV-PH and truncated ATI protein to protrusions from the cell surface was observed.

CONCLUSION:

These results suggest a role for ECTV-PH in intracellular transport of viral proteins or intercellular spread of the virus. Broader implications include better understanding of the virus-host relationship and mechanisms by which cells organize and control the actin cytoskeleton.

• Authors: Christine Butler-Cole, Mary J Wagner, Melissa Da Silva, Gordon D Brown, Robert D Burke, Chris Upton
• Journal: Virology journal (Virol J) Vol. 4 Pg. 76 (Jul 24 2007) ISSN: 1743-422X [Electronic] England
• PMID: 17650322 (Publication Type: Journal Article, Research Support, N.I.H., Extramural, Research Support, Non-U.S. Gov't)

Chemical References

• A-type inclusion protein, poxvirus
• Actins
• Profilins
• Tropomyosin
• Viral Proteins

Topics

• Actins (chemistry)
• Animals
• Blotting, Western
• Cell Line
• Chlorocebus aethiops
• Cytoplasm (chemistry)
• Ectromelia virus (genetics, physiology)
• Immunoprecipitation
• Inclusion Bodies, Viral (chemistry)
• Microscopy, Confocal
• Microscopy, Fluorescence
• Models, Molecular
• Phylogeny
• Profilins (genetics, metabolism)
• Protein Binding
• Protein Structure, Tertiary
• Sequence Homology, Amino Acid
• Tropomyosin (metabolism)
• Viral Proteins (metabolism)