Unbiased proteomic analysis of proteins interacting with the HIV-1 5'LTR sequence: role of the transcription factor Meis.
To depict the largest picture of a core promoter interactome, we developed a one-step DNA-affinity capture method coupled with an improved mass spectrometry analysis process focused on the identification of low abundance proteins. As a proof of concept, this method was developed through the analysis of 230 bp contained in the 5'long terminal repeat (LTR) of the human immunodeficiency virus 1 (HIV-1). Beside many expected interactions, many new transcriptional regulators were identified, either transcription factors (TFs) or co-regulators, which interact directly or indirectly with the HIV-1 5'LTR. Among them, the homeodomain-containing TF myeloid ectopic viral integration site was confirmed to functionally interact with a specific binding site in the HIV-1 5'LTR and to act as a transcriptional repressor, probably through recruitment of the repressive Sin3A complex. This powerful and validated DNA-affinity approach could also be used as an efficient screening tool to identify a large set of proteins that physically interact, directly or indirectly, with a DNA sequence of interest. Combined with an in silico analysis of the DNA sequence of interest, this approach provides a powerful approach to select the interacting candidates to validate functionally by classical approaches.
Laboratory of Biochemistry and Cell Biology (URBC), NAmur Research Institute for LIfe Sciences, University of Namur, 61 rue de Bruxelles, 5000 Namur, Belgium.
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A
Tacheny
S
S
Michel
M
M
Dieu
L
L
Payen
T
T
Arnould
P
P
Renard
D001665:Binding Sites; D004268:DNA-Binding Proteins / Q000032:analysis / Q000302:isolation & purification; D016325:HIV Long Terminal Repeat; D015497:HIV-1 / Q000235:genetics*; D006367:HeLa Cells; D018398:Homeodomain Proteins / Q000378:metabolism* / Q000502:physiology; D006801:Humans; D013058:Mass Spectrometry; D000075064:Myeloid Ecotropic Viral Integration Site 1 Protein; D016328:NF-kappa B / Q000032:analysis; D009363:Neoplasm Proteins / Q000378:metabolism* / Q000502:physiology; D040901:Proteomics / Q000379:methods*; D012097:Repressor Proteins / Q000032:analysis; D014157:Transcription Factors / Q000378:metabolism*; D014158:Transcription, Genetic
D004268:DNA-Binding Proteins; D018398:Homeodomain Proteins; D000075064:Myeloid Ecotropic Viral Integration Site 1 Protein; D016328:NF-kappa B; D009363:Neoplasm Proteins; D012097:Repressor Proteins; D014157:Transcription Factors