Transcription factor binding sites in the pol gene intragenic regulatory region of HIV-1 are important for virus infectivity.
We have previously identified in the pol gene of human immunodeficiency virus type 1 (HIV-1) a new positive transcriptional regulatory element (nt 4481-4982) containing recognition sites for nuclear proteins (sites B, C, D and a GC-box) [C. Van Lint, J. Ghysdael, P. Paras, Jr, A. Burny and E. Verdin (1994) J. Virol. 68, 2632-2648]. In this study, we have further physically characterized each binding site and have shown that the transcription factors Oct-1, Oct-2, PU.1, Sp1 and Sp3 interact in vitro with the pol region. Chromatin immunoprecipitation assays using HIV-infected cell lines demonstrated in the context of chromatin that Sp1, Sp3, Oct-1 and PU.1 are recruited to the HS7 region in vivo. For each site, we have identified mutations abolishing factor binding to their cognate DNA sequences without altering the underlying amino acid sequence of the integrase. By transient transfection assays, we have demonstrated the involvement of the pol binding sites in the transcriptional enhancing activity of the intragenic region. Our functional results with multimerized wild-type and mutated pol binding sites separately (i.e. in the absence of the other sites) have demonstrated that the PU.1, Sp1, Sp3 and Oct-1 transcription factors regulate the transcriptional activity of a heterologous promoter through their respective HS7 binding sites. Finally, we have investigated the physiological role of the HS7 binding sites in HIV-1 replication and have shown that these sites are important for viral infectivity.
Goffin
Véronique
V
Laboratoire de Virologie Moléculaire, Institut de Biologie et de Médecine Moléculaires (IBMM), Service de Chimie Biologique, Université Libre de Bruxelles Rue des Profs Jeener et Brachet 12, 6041 Gosselies, Belgium.
Demonté
Dominique
D
Vanhulle
Caroline
C
de Walque
Stéphane
S
de Launoit
Yvan
Y
Burny
Arsène
A
Collette
Yves
Y
Van Lint
Carine
C
D000818:Animals; D001665:Binding Sites; D002460:Cell Line; D004252:DNA Mutational Analysis; D004268:DNA-Binding Proteins / Q000378:metabolism; D015967:Gene Expression Regulation, Viral; D015684:Gene Products, pol / Q000235:genetics*; D015696:Gene Products, tat / Q000378:metabolism; D015497:HIV-1 / Q000235:genetics* / Q000502:physiology; D006801:Humans; D051379:Mice; D050811:Octamer Transcription Factor-1; D017354:Point Mutation; D011401:Promoter Regions, Genetic; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D020218:Response Elements; D016329:Sp1 Transcription Factor / Q000378:metabolism; D051705:Sp3 Transcription Factor; D013937:Thymidine Kinase / Q000235:genetics; D015534:Trans-Activators / Q000378:metabolism; D014157:Transcription Factors / Q000737:chemistry / Q000378:metabolism*; D015533:Transcriptional Activation; D014779:Virus Replication; D016335:Zinc Fingers; D054322:tat Gene Products, Human Immunodeficiency Virus; D000099232:Proto-Oncogene Protein Spi-1
D004268:DNA-Binding Proteins; D015684:Gene Products, pol; D015696:Gene Products, tat; D050811:Octamer Transcription Factor-1; D011518:Proto-Oncogene Proteins; D016329:Sp1 Transcription Factor; D051705:Sp3 Transcription Factor; D013937:Thymidine Kinase; D015534:Trans-Activators; D014157:Transcription Factors; D054322:tat Gene Products, Human Immunodeficiency Virus; D000099232:Proto-Oncogene Protein Spi-1; C494165:POU2F1 protein, human; C494166:Pou2f1 protein, mouse; C497121:SP3 protein, human; C497122:Sp3 protein, mouse