Human immunodeficiency virus type 1 Tat prevents dephosphorylation of Sp1 by TCF-4 in astrocytes.
Previous examination of the effect of TCF-4 on transcription of the human immunodeficiency virus type 1 (HIV-1) promoter in human astrocytic cells found that TCF-4 affects the HIV-1 promoter through the GC-rich domain (nt -80 to nt -68). Here, the physical interaction and a functional consequence of TCF4-Sp1 contact were characterized. It was shown that expression of TCF-4 in U-87 MG (human astrocytic) cells decreased basal and Sp1-mediated transcription of the HIV-1 promoter. Results from a GST pull-down assay, as well as combined immunoprecipitation and Western blot analysis of protein extracts from U-87 MG cells, revealed an interaction of Sp1 with TCF-4. Using in vitro protein chromatography, the region of Sp1 that contacts TCF-4 was mapped to aa 266-350. It was also found that, in cell-free extracts, TCF-4 prevented dsDNA-dependent protein kinase (DNA-PK)-mediated Sp1 phosphorylation. Surprisingly, TCF-4 failed to decrease Sp1-mediated transcription of the HIV-1 long terminal repeat (LTR) and Sp1 phosphorylation in cells expressing HIV-1 Tat. Results from immunoprecipitation/Western blotting demonstrated that TCF-4 lost its ability to interact with Sp1, but not with Tat, in Tat-transfected cells. Taken together, these findings suggest that activity at the HIV-1 promoter is influenced by phosphorylation of Sp1, which is affected by Tat and DNA-PK. Interactions among TCF-4, Sp1 and/or Tat may determine the level of viral gene transcription in human astrocytic cells.
Rossi
Andrea
A
Laboratory of Biology, Don C. Gnocchi Foundation, IRCCS, 20148 Milan, Italy.|Department of Neuroscience & Center for Neurovirology, Temple University School of Medicine, 1900 N 12th Street (015-96), Philadelphia, PA 19122, USA.
Mukerjee
Ruma
R
Department of Neuroscience & Center for Neurovirology, Temple University School of Medicine, 1900 N 12th Street (015-96), Philadelphia, PA 19122, USA.
Ferrante
Pasquale
P
Laboratory of Biology, Don C. Gnocchi Foundation, IRCCS, 20148 Milan, Italy.
Khalili
Kamel
K
Department of Neuroscience & Center for Neurovirology, Temple University School of Medicine, 1900 N 12th Street (015-96), Philadelphia, PA 19122, USA.
Amini
Shohreh
S
Department of Biology, Temple University School of Medicine, 1900 N 12th Street (015-96), Philadelphia, PA 19122, USA.|Department of Neuroscience & Center for Neurovirology, Temple University School of Medicine, 1900 N 12th Street (015-96), Philadelphia, PA 19122, USA.
Sawaya
Bassel E
BE
Department of Neuroscience & Center for Neurovirology, Temple University School of Medicine, 1900 N 12th Street (015-96), Philadelphia, PA 19122, USA.
D001253:Astrocytes; D045744:Cell Line, Tumor; D015696:Gene Products, tat / Q000235:genetics / Q000378:metabolism*; D015756:Genes, tat; D016325:HIV Long Terminal Repeat / Q000235:genetics / Q000502:physiology; D015497:HIV-1 / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D010766:Phosphorylation; D016329:Sp1 Transcription Factor / Q000235:genetics / Q000378:metabolism*; D051820:TCF Transcription Factors / Q000235:genetics / Q000378:metabolism*; D058118:Transcription Factor 7-Like 2 Protein; D014162:Transfection; D054322:tat Gene Products, Human Immunodeficiency Virus
D015696:Gene Products, tat; D016329:Sp1 Transcription Factor; D051820:TCF Transcription Factors; C544894:TCF7L2 protein, human; D058118:Transcription Factor 7-Like 2 Protein; D054322:tat Gene Products, Human Immunodeficiency Virus