Repression of c-Kit and Its Downstream Substrates by GATA-1 Inhibits Cell Proliferation during Erythroid Maturation

Abstract

Stem cell factor (SCF), erythropoietin (Epo), and GATA-1 play an essential role(s) in erythroid development. We examined how these proteins interact functionally in G1E cells, a GATA-1 ؊ erythroblast line that proliferates in an SCF-dependent fashion and, upon restoration of GATA-1 function, undergoes GATA-1 proliferation arrest and Epo-dependent terminal maturation. We show that SCF-induced cell cycle progression is mediated via activation of the Src kinase/c-Myc pathway. Restoration of GATA-1 activity induced G 1 cell cycle arrest coincident with repression of c-Kit and its downstream effectors Vav1, Rac1, and Akt. Sustained expression of each of these individual signaling components inhibited GATA-1-induced cell cycle arrest to various degrees but had no effects on the expression of GATA-1-regulated erythroid maturation markers. Chromatin immunoprecipitation analysis revealed that GATA-1 occupies a defined Kit gene regulatory element in vivo, suggesting a direct mechanism for gene repression. Hence, in addition to its well-established function as an activator of erythroid genes, GATA-1 also participates in a distinct genetic program that inhibits cell proliferation by repressing the expression of multiple components of the c-Kit signaling axis. Our findings reveal a novel aspect of molecular cross talk between essential transcriptional and cytokine signaling components of hematopoietic development.Receptor tyrosine kinases (RTKs) trigger a multitude of cellular events, including proliferation, survival, differentiation, and migration. These functions are modulated in hematopoietic stem and progenitor cells by the essential RTK c-Kit (8,11,43). The expression of c-Kit is downregulated as progenitors mature to their respective lineages, with the exception of mast cells, which rely on c-Kit for survival, proliferation, and function throughout their life span (20). Unrestrained c-Kit activity contributes to several neoplastic disorders, including gastrointestinal stromal tumors (GIST), mastocytosis, and leukemia (5,12,21,37,46,55). In GIST, somatic kinase-activating Kit mutations result in malignant transformation. In the hematopoietic system, similar activating Kit mutations occur in stem/ progenitor cells and mast cells, causing mastocytosis and acute myelogenous leukemia, respectively (45, 54).Mutant mice without c-Kit (dominant white spotting, or W, mutants) demonstrate severe deficiencies in erythroid development with reduced CFU-erythroid progenitors in the fetal liver and embryonic death from anemia at around day 16 of gestation (11, 57). Erythropoietin (Epo) receptor (Epo-R)-deficient mice demonstrate a similar decrease in CFU-erythroid progenitors and die of anemia between days 13 and 15 of gestation, suggesting that erythroid progenitors cannot survive, proliferate, or differentiate unless both the c-Kit and Epo-R signal transduction pathways are functional. Recent studies suggest that Epo and Epo-R interactions contribute to this process by preventing apoptosis through activation of the survival factor...

MicroRNAs 221 and 222 inhibit normal erythropoiesis and erythroleukemic cell growth via kit receptor down-modulation

“…Thus, (i) the final phase of E culture, coupled with erythroblast maturation and declining proliferation, is characterized by a drop of kit mRNA and protein levels indicating transcriptional repression of the kit gene. This process may be mediated by GATA-1 transcription factor, which exerts a suppressive action on kit gene promoter in late erythropoiesis (31). (ii) Addition of KL to E culture causes an enhanced proliferation, coupled with a sharp increase of kit mRNA and protein levels: this finding suggests that in this culture system, the unblocking of kit mRNA trans- lation via miR 221 and 222 is potentiated by activation of kit gene transcription.…”

Oncogenic pathways of AML1-ETO in acute myeloid leukemia: Multifaceted manipulation of marrow maturation

“…Thus AML1-ETO repression of GATA-1 may initially disrupt its positive autoregulation and thereby undermine its entire transcriptional program. Notably, a critical component of the GATA-1 transcriptional program consists of downregulating c-Kit expression [59]. Abnormally sustained c-Kit expression in GATA-1-deficient cells may then predispose these genetically unstable cells to positive selection for receptor-activating mutations, accounting for the high frequency of C-KIT mutation in t(8;21)-positive AMLs (Figure 1).…”

CDC6 expression is regulated by lineage-specific transcription factor GATA1

“…Initially identified as a protein that binds cis-regulatory elements of the β-globin gene, it has since been shown to regulate a vast number of specific erythro-megakaryocytic genes 13,14 or hematopoietic proto-oncogenes, such as c-MYC and c-MYB. 15,16 underlining the potential dominant-negative effect of GATA1s. GATA1-dependent increase in CDC6 expression could also be seen at the protein level (Fig.…”