GLUT-1-independent infection of the glioblastoma/astroglioma U87 cells by the human T cell leukemia virus type 1.
The human glucose transporter protein 1 (GLUT-1) functions as a receptor for human T cell leukemia virus (HTLV). GLUT-1 is a twelve-transmembrane cell surface receptor with six extracellular (ECL) and seven intracellular domains. To analyze HTLV-1 cytotropism, we utilized polyclonal antibodies to a synthetic peptide corresponding to the large extracellular domain of GLUT-1. The antibodies caused significant blocking of envelope (Env)-mediated fusion and pseudotyped virus infection of HeLa cells but had no significant effect on infection of U87 cells. This differential effect correlated with the detection of high-level surface expression of GLUT-1 on HeLa cells and very weak staining of U87 cells. To investigate this in terms of viral cytotropism, we cloned GLUT-1 cDNA from U87 cells and isolated two different versions of cDNA clones: the wild-type sequence (encoding 492 residues) and a mutant cDNA with a 5-base pair deletion (GLUT-1Delta5) between nucleotides 1329 and 1333. The deletion, also detected in genomic DNA, resulted in a frame-shift and premature termination producing a truncated protein of 463 residues. Transfection of the wild-type GLUT-1 but not GLUT-1Delta5 cDNA into CHO cells resulted in efficient surface expression of the human GLUT-1. Co-expression of GLUT-1 with GLUT-1Delta5 produces a trans-inhibition by GLUT-1Delta5 of GLUT-1-mediated HTLV-1 envelope (Env)-mediated fusion. Co-immunoprecipitation experiments demonstrated physical interaction of the wild-type and mutant proteins. Northern blot and RT-PCR analyses demonstrated lower GLUT-1 RNA expression in U87 cells. We propose two mechanisms to account for the impaired cell surface expression of GLUT-1 on U87 cells: low GLUT-1 RNA expression and the formation of GLUT-1/GLUT-1Delta5 heterodimers that are retained intracellularly. Significant RNAi-mediated reduction of endogenous GLUT-1 expression impaired HTLV-1 Env-mediated fusion with HeLa cells but not with U87 cells. We propose a GLUT-1-independent mechanism of HTLV-1 infection of U87 cells. The results may have important implications for HTLV-1 neurotropism and pathogenesis.
Jin
Qingwen
Q
Department of Neurology, Nanjing Medical University, Nanjing, Jiangsu Province, 210029, China.
Agrawal
Lokesh
L
Vanhorn-Ali
Zainab
Z
Alkhatib
Ghalib
G
D000818:Animals; D001254:Astrocytoma / Q000473:pathology / Q000821:virology*; D016466:CHO Cells; D045744:Cell Line, Tumor; D006224:Cricetinae; D018076:DNA, Complementary; D016368:Frameshift Mutation; D005909:Glioblastoma / Q000473:pathology / Q000821:virology*; D051272:Glucose Transporter Type 1 / Q000032:analysis / Q000235:genetics / Q000276:immunology / Q000502:physiology*; D015490:HTLV-I Infections / Q000821:virology*; D006367:HeLa Cells; D015368:Human T-lymphotropic virus 1 / Q000502:physiology*; D006801:Humans
D018076:DNA, Complementary; D051272:Glucose Transporter Type 1