PU.1 inhibits GATA-1 function and erythroid differentiation by blocking GATA-1 DNA binding.
The lineage-specific transcription factors GATA-1 and PU.1 can physically interact to inhibit each other's function, but the mechanism of repression of GATA-1 function by PU.1 has not been elucidated. Both the N terminus and the C terminus of PU.1 can physically interact with the C-terminal zinc finger of GATA-1. It is demonstrated that the PU.1 N terminus, but not the C terminus, is required for inhibiting GATA-1 function. Induced overexpression of PU.1 in K562 erythroleukemia cells blocks hemin-induced erythroid differentiation. In this system, PU.1 does not affect the expression of GATA-1 messenger RNA, protein, or nuclear localization. However, GATA-1 DNA binding decreases dramatically. By means of electrophoretic mobility shift assays with purified proteins, it is demonstrated that the N-terminal 70 amino acids of PU.1 can specifically block GATA-1 DNA binding. In addition, PU.1 had a similar effect in the G1ER cell line, in which the GATA-1 null erythroid cell line G1E has been transduced with a GATA-1-estrogen receptor fusion gene, which is directly dependent on induction of the GATA-1 fusion protein to effect erythroid maturation. Consistent with in vitro binding assays, overexpression of PU.1 blocked DNA binding of the GATA-1 fusion protein as well as GATA-1-mediated erythroid differentiation of these G1ER cells. These results demonstrate a novel mechanism by which function of a lineage-specific transcription factor is inhibited by another lineage-restricted factor through direct protein-protein interactions. These findings contribute to understanding how protein-protein interactions participate in hematopoietic differentiation and leukemogenesis. (Blood. 2000;96:2641-2648)
Zhang
P
P
Hematology/Oncology Division, Harvard Institute of Medicine, Harvard Medical School, Boston, MA 02115, USA.
Zhang
X
X
Iwama
A
A
Yu
C
C
Smith
K A
KA
Mueller
B U
BU
Narravula
S
S
Torbett
B E
BE
Orkin
S H
SH
Tenen
D G
DG
D001692:Biological Transport; D002454:Cell Differentiation / Q000502:physiology; D019070:Cell Lineage; D002467:Cell Nucleus / Q000378:metabolism; D004247:DNA / Q000378:metabolism*; D004273:DNA, Neoplasm / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000037:antagonists & inhibitors* / Q000378:metabolism; D015672:Erythroid Precursor Cells / Q000378:metabolism*; D050981:Erythroid-Specific DNA-Binding Factors; D004920:Erythropoiesis / Q000502:physiology*; D004958:Estradiol / Q000494:pharmacology; D050982:GATA1 Transcription Factor; D015973:Gene Expression Regulation, Leukemic / Q000502:physiology; D005813:Genes, Synthetic; D006801:Humans; D020014:K562 Cells / Q000378:metabolism; D009363:Neoplasm Proteins / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011518:Proto-Oncogene Proteins / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D012333:RNA, Messenger / Q000032:analysis; D012334:RNA, Neoplasm / Q000032:analysis; D011960:Receptors, Estrogen / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000502:physiology; D015534:Trans-Activators / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D014157:Transcription Factors / Q000037:antagonists & inhibitors* / Q000378:metabolism; D014162:Transfection; D000099232:Proto-Oncogene Protein Spi-1
D004247:DNA; D004273:DNA, Neoplasm; D004268:DNA-Binding Proteins; D050981:Erythroid-Specific DNA-Binding Factors; D004958:Estradiol; D050982:GATA1 Transcription Factor; D009363:Neoplasm Proteins; D011518:Proto-Oncogene Proteins; D012333:RNA, Messenger; D012334:RNA, Neoplasm; D011960:Receptors, Estrogen; D011993:Recombinant Fusion Proteins; D015534:Trans-Activators; D014157:Transcription Factors; D000099232:Proto-Oncogene Protein Spi-1; C494688:GATA1 protein, human