DNase I hypersensitive sites in the 5' flanking region of the human plasminogen activator inhibitor type 2 (PAI-2) gene are associated with basal and tumor necrosis factor-alpha-induced transcription in monocytes.
The plasminogen activator inhibitor type 2 (PAI-2) gene encodes a serine proteinase inhibitor (serpin) which is rapidly induced in response to the inflammatory cytokine, tumour necrosis factor-alpha (TNFalpha) in monocytes and macrophages. As an initial step towards understanding the molecular mechanisms underlying PAI-2 gene regulation in monocytes, we report here the analysis of the chromatin structure of 9.6 kb of 5' flanking region of the human PAI-2 gene for potential cis-acting regulatory regions using DNase I hypersensitivity mapping. Sites sensitive to DNase I were mapped in two monocytic cell lines representative of early monocytic differentiation; U937 cells, which synthesise low constitutive levels of PAI-2 that were induced following treatment with TNFalpha, and a MonoMac6 cell line which did not synthesise PAI-2 even after treatment with TNFalpha. Six DNase I hypersensitive sites (DHS) were identified; three upstream of the transcription initiation site (DH1, DH2, DH3) and three downstream of the transcription initiation site which were contained within intron A (DH4, DH5) and the exon 2/intron B junction (DH6). Among these, one distally located DH site (DH2) disappeared in both cell lines following treatment with TNFalpha. Two DH sites (DH1, DH6) were absent in PAI-2-producing U937 cells, but were present in MonoMac6 cells, which did not produce PAI-2, indicating the possible involvement of negative regulatory elements in the suppression of PAI-2 gene expression. The results demonstrate the involvement of chromatin structure in transcriptional responsiveness of the PAI-2 gene promoter and identify several loci which may be key control regions for PAI-2 gene transcription.
Queensland Cancer Fund Experimental Oncology Program, The Queensland Institute of Medical Research, Brisbane, Australia.
D
D
Mahony
B W
BW
Stringer
J L
JL
Dickinson
T M
TM
Antalis
D001483:Base Sequence; D002460:Cell Line; D004247:DNA / Q000378:metabolism; D003850:Deoxyribonuclease I / Q000378:metabolism*; D018922:HL-60 Cells; D006801:Humans; D008264:Macrophages / Q000187:drug effects; D008969:Molecular Sequence Data; D009000:Monocytes / Q000187:drug effects*; D017396:Plasminogen Activator Inhibitor 2 / Q000235:genetics*; D014158:Transcription, Genetic / Q000187:drug effects*; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*
D017396:Plasminogen Activator Inhibitor 2; D014409:Tumor Necrosis Factor-alpha; D004247:DNA; D003850:Deoxyribonuclease I