Avian reovirus sigmaA localizes to the nucleolus and enters the nucleus by a nonclassical energy- and carrier-independent pathway.
Avian reovirus sigmaA is a double-stranded RNA (dsRNA)-binding protein that has been shown to stabilize viral core particles and to protect the virus against the antiviral action of interferon. To continue with the characterization of this viral protein, we have investigated its intracellular distribution in avian cells. Most sigmaA accumulates into cytoplasmic viral factories of infected cells, and yet a significant fraction was detected in the nucleolus. The protein also localizes in the nucleolus of transfected cells, suggesting that nucleolar targeting is not facilitated by the viral infection or by viral factors. Assays performed in both intact cells and digitonin-permeabilized cells demonstrate that sigmaA is able to enter the nucleus via a nucleoporin-dependent nondiffusional mechanism that does not require added cytosolic factors or energy input. These results indicate that sigmaA by itself is able to penetrate into the nucleus using a process that is mechanistically different from the classical nuclear localization signal/importin pathway. On the other hand, two sigmaA arginines that are necessary for dsRNA binding are also required for nucleolar localization, suggesting that dsRNA-binding and nucleolar targeting are intimately linked properties of the viral protein.
Vázquez-Iglesias
Lorena
L
Departamento de Bioquímica y Biología Molecular, Facultad de Farmacia, Universidad de Santiago de Compostela, Santiago de Compostela, Spain.
Lostalé-Seijo
Irene
I
Martínez-Costas
José
J
Benavente
Javier
J
D000818:Animals; D002460:Cell Line; D002466:Cell Nucleolus / Q000378:metabolism*; D002642:Chick Embryo; D003593:Cytoplasm / Q000378:metabolism; D003600:Cytosol / Q000378:metabolism; D004072:Digitonin / Q000494:pharmacology; D028884:Karyopherins / Q000378:metabolism; D008856:Microscopy, Fluorescence / Q000379:methods; D019913:Nuclear Localization Signals / Q000378:metabolism; D030101:Orthoreovirus, Avian / Q000378:metabolism*; D012330:RNA, Double-Stranded / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000378:metabolism* / Q000502:physiology*; D011994:Recombinant Proteins / Q000737:chemistry; D013347:Subcellular Fractions / Q000378:metabolism; D014758:Viral Core Proteins / Q000378:metabolism* / Q000502:physiology*
D028884:Karyopherins; D019913:Nuclear Localization Signals; D012330:RNA, Double-Stranded; D016601:RNA-Binding Proteins; D011994:Recombinant Proteins; D014758:Viral Core Proteins; C403292:sigmaA protein, avian reovirus; D004072:Digitonin