Localization of human T-cell lymphotropic virus type II Tax protein is dependent upon a nuclear localization determinant in the N-terminal region.
Human T-cell lymphotropic viruses (HTLV) types I and II are closely related oncogenic retroviruses that have been associated with lymphoproliferative and neurological disorders. The proviral genome encodes a trans-regulatory Tax protein that activates viral genes and upregulates various cellular genes involved in both cell growth and transformation. Tax proteins of HTLV-I (Tax-I) and HTLV-II (Tax-II) exhibit more than 77% aa homology and expression of either Tax-I or Tax-II is sufficient for immortalization of cultured T lymphocytes. Tax-I shuttles from the nucleus to the cytoplasm and accumulates within the nucleus, whereas Tax-II is found mainly in the cytoplasm. In the present study we have used recombinant vectors to analyze the size and structure of the nuclear localization domain within the Tax-II protein sequence. The Tax-II protein was expressed in HeLa cells either as the complete protein, or regions thereof, that were individually fused to the green fluorescent protein (GFP). Immunoblot analysis of the fused Tax-II products confirmed their expression and size. Fluorescence microscopy studies indicated that the complete Tax-II as well as N-truncated forms presented a punctuate cytoplasmic distribution and that a nuclear localization determinant is confined to within the first 60 aa of Tax-II. Accordingly, site directed mutagenesis and deletion of specific sequences within the first 60 aa showed that the nuclear determinant lies within the first 41 residues of Tax-II. These results point to a direct involvement of the amino-terminal residues of Tax-II protein in determining its nuclear functionality.
Turci
Marco
M
Department of Mother and Child, Biology and Genetics, University of Verona, Strada Le Grazie 8, 37134 Verona, Italy.
Romanelli
Maria Grazia
MG
Lorenzi
Pamela
P
Righi
Paola
P
Bertazzoni
Umberto
U
D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal / Q000378:metabolism; D015153:Blotting, Western; D002467:Cell Nucleus / Q000378:metabolism*; D003593:Cytoplasm / Q000378:metabolism; D005456:Fluorescent Dyes; D016356:Gene Products, tax / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D005822:Genetic Vectors; D049452:Green Fluorescent Proteins / Q000032:analysis; D006367:HeLa Cells; D015367:Human T-lymphotropic virus 2 / Q000378:metabolism*; D006801:Humans; D007211:Indoles; D008856:Microscopy, Fluorescence; D008969:Molecular Sequence Data; D019913:Nuclear Localization Signals; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D017384:Sequence Deletion; D017386:Sequence Homology, Amino Acid
D000911:Antibodies, Monoclonal; D005456:Fluorescent Dyes; D016356:Gene Products, tax; D007211:Indoles; D019913:Nuclear Localization Signals; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins; C007293:DAPI