NM23-H1 and NM23-H2 repress transcriptional activities of nuclease-hypersensitive elements in the platelet-derived growth factor-A promoter.
The platelet-derived growth factor (PDGF)-A promoter is regulated by a number of GC-rich regulatory elements that possess non-B-form DNA structures. Screening of a HeLa cDNA expression library with the C-rich strand of a PDGF-A silencer sequence (5'-S1 nuclease-hypersensitive site (SHS)) yielded three cDNA clones encoding NM23-H1, a protein implicated as a suppressor of metastasis in melanoma and breast carcinoma. Recombinant human NM23-H1 cleaved within the 3'-portions of both 5'-SHS strands in either single-stranded or duplex forms. In contrast, NM23-H2, known as a transcriptional activator with a DNA cleavage function, cleaved within the 5'-portions of both strands, revealing that NM23-H1 and NM23-H2 cleave at distinct sites of the 5'-SHS and by different mechanisms. NM23-H1 and NM23-H2 also cleaved within the PDGF-A basal promoter region, again exhibiting preferences for cleavage within the 5'- and 3'-portions of the element, respectively. Transient transfection analyses in HepG2 cells revealed that both NM23-H1 and -H2 repressed transcriptional activity driven by the PDGF-A basal promoter (-82 to +8). Activity of the negative regulatory region (-1853 to -883), which contains the 5'-SHS, was also inhibited modestly by NM23-H1 and NM23-H2. These studies demonstrate for the first time that NM23-H1 interacts both structurally and functionally with DNA. They also indicate a role for NM23 proteins in repressing transcription of a growth factor oncogene, providing a possible molecular mechanism to explain their metastasis-suppressing effects.
Ma
Deqin
D
Department of Molecular and Biomedical Pharmacology, University of Kentucky Medical Center, Lexington, Kentucky 40536, USA.
Xing
Zhenlan
Z
Liu
Bin
B
Pedigo
Nancy G
NG
Zimmer
Stephen G
SG
Bai
Zengliang
Z
Postel
Edith H
EH
Kaetzel
David M
DM
D000951:Antigens, Neoplasm / Q000378:metabolism; D014408:Biomarkers, Tumor; D015723:Gene Library; D006367:HeLa Cells; D006801:Humans; D020559:Monomeric GTP-Binding Proteins / Q000378:metabolism*; D054778:NM23 Nucleoside Diphosphate Kinases; D009701:Nucleoside-Diphosphate Kinase; D009838:Oligodeoxyribonucleotides / Q000378:metabolism; D010982:Platelet-Derived Growth Factor / Q000235:genetics*; D011401:Promoter Regions, Genetic; D011994:Recombinant Proteins / Q000378:metabolism; D014157:Transcription Factors / Q000378:metabolism*; D014158:Transcription, Genetic
D000951:Antigens, Neoplasm; D014408:Biomarkers, Tumor; D054778:NM23 Nucleoside Diphosphate Kinases; D009838:Oligodeoxyribonucleotides; D010982:Platelet-Derived Growth Factor; D011994:Recombinant Proteins; D014157:Transcription Factors; C075278:platelet-derived growth factor A; C517938:NME1 protein, human; D009701:Nucleoside-Diphosphate Kinase; D020559:Monomeric GTP-Binding Proteins