Integration of the activation of the human hyaluronan synthase 2 gene promoter by common cofactors of the transcription factors retinoic acid receptor and nuclear factor kappaB.
Hyaluronan (HA) is a polysaccharide of the vertebrate extracellular matrix, produced by three related HA synthases (HASs) that influence numerous physiological processes. We screened the first 2250 bp of the HAS2 promoter for transcription factor response elements (REs) in silico and found 1 cluster of 2 retinoic acid (RA) REs, 3 discrete NF-kappaB factors, and 12 Sp1 REs. In parallel, we scanned nine overlapping promoter regions in HaCaT human immortalized keratinocytes using chromatin immunoprecipitation assays to identify binding of mediator, coactivator, and corepressor proteins and Sp1 transcription factor in response to all-trans-RA and tumor necrosis factor-alpha (TNF-alpha). We found that all-trans-RA modulated the binding of the RA receptor and several coregulators to the region containing the RARE cluster at position -1230. The importance of this region is supported in reporter gene assays by the all-trans-RA induction of the respective promoter region. Similarly, we showed by chromatin immunoprecipitation assays as well as by gel-shift assays with nuclear extracts that TNF-alpha induced NF-kappaB binding to regions at positions -380, -1420, and -1890, demonstrated its association with RNA polymerase II and cofactor proteins, and confirmed the functionality of the respective promoter regions in vivo. These findings partially explain the induction of HAS2 mRNA by all-trans-RA and TNF-alpha and provide an example how the action of different transcription factor families can use the same cofactors.
Saavalainen
Katri
K
Department of Biochemistry, University of Kuopio, FIN-70211 Kuopio, Finland.
Tammi
Markku I
MI
Bowen
Timothy
T
Schmitz
M Lienhard
ML
Carlberg
Carsten
C
D001483:Base Sequence; D001665:Binding Sites; D002460:Cell Line; D015971:Gene Expression Regulation, Enzymologic; D015894:Genome, Human / Q000235:genetics; D014453:Glucuronosyltransferase / Q000235:genetics* / Q000378:metabolism*; D006801:Humans; D000076002:Hyaluronan Synthases; D016328:NF-kappa B / Q000378:metabolism*; D011401:Promoter Regions, Genetic / Q000235:genetics*; D011485:Protein Binding; D012333:RNA, Messenger / Q000235:genetics; D018168:Receptors, Retinoic Acid / Q000378:metabolism*; D016329:Sp1 Transcription Factor / Q000378:metabolism; D014158:Transcription, Genetic / Q000187:drug effects / Q000235:genetics; D014212:Tretinoin / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology
D016328:NF-kappa B; D012333:RNA, Messenger; D018168:Receptors, Retinoic Acid; D016329:Sp1 Transcription Factor; D014409:Tumor Necrosis Factor-alpha; D014212:Tretinoin; D014453:Glucuronosyltransferase; C101754:HAS2 protein, human; D000076002:Hyaluronan Synthases