Truncation of Sp1 transcription factor by myeloblastin in undifferentiated HL60 cells.
When HL60 cells are exposed to 1,25-dihydroxyvitamin D3 (1,25D3), they undergo changes approximating the phenotype of the monocyte. Little is known, however, about the regulation and the mechanisms of this transition. It was previously noted that DNA binding by the Sp1 transcription factor in nuclear extracts of HL60 cells is profoundly altered when these cells are induced to differentiate by 1,25D3. In the present study, we show that in untreated HL60 cells only a truncated, approximately 30-kDa Sp1 fragment, encompassing the C-terminal region, binds to the GC element-containing DNA. Full-length 105-kDa Sp1 protein cannot be detected in these cells, although reverse transriptase-polymerase chain reaction reveals the presence of both 5' and 3' ends of Sp1 mRNA. Following treatment with 10(7) M 1,25D3 for 96 hr or in cells made resistant to 1,25D3 or to 1-beta-D-arabinocytosine, the Sp1 protein can be demonstrated. After an exposure to purified myeloblastin, a serine protease, purified recombinant Sp1 protein and extracts of 1,25D3-treated cells show a pattern of DNA binding similar to the pattern seen using extracts of untreated HL60 cells, indicating that the Sp1 protein is a target for myeloblastin. Because myeloblastin is present in naive HL60 cells and is downregulated during their differentiation, inhibition of proteolysis of these transcription factors seems to provide a mechanism through which differentiating HL60 cells can acquire a new repertoire of gene expression, perhaps for the maintenance of the differentiated phenotype.
Rao
J
J
Department of Pathology and Laboratory Medicine, UMDNJ-New Jersey Medical School, Newark 07103, USA.
Zhang
F
F
Donnelly
R J
RJ
Spector
N L
NL
Studzinski
G P
GP
D001667:Binding, Competitive; D002117:Calcitriol / Q000494:pharmacology*; D002454:Cell Differentiation / Q000502:physiology; D004268:DNA-Binding Proteins / Q000032:analysis / Q000737:chemistry / Q000235:genetics; D015536:Down-Regulation / Q000502:physiology; D015972:Gene Expression Regulation, Neoplastic / Q000235:genetics; D018922:HL-60 Cells; D006801:Humans; D053146:Myeloblastin; D009687:Nuclear Proteins / Q000032:analysis; D012333:RNA, Messenger / Q000032:analysis; D011994:Recombinant Proteins / Q000235:genetics; D012697:Serine Endopeptidases / Q000378:metabolism*; D016329:Sp1 Transcription Factor / Q000235:genetics / Q000378:metabolism*; D051705:Sp3 Transcription Factor; D013696:Temperature; D014157:Transcription Factors / Q000032:analysis
D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; D012333:RNA, Messenger; D011994:Recombinant Proteins; D016329:Sp1 Transcription Factor; D014157:Transcription Factors; D051705:Sp3 Transcription Factor; D012697:Serine Endopeptidases; D053146:Myeloblastin; D002117:Calcitriol