Lyn and Fgr Protein-tyrosine Kinases Prevent Apoptosis during Retinoic Acid-induced Granulocytic Differentiation of HL-60 Cells

Abstract

The human promyelocytic leukemia cell line HL-60 can be induced to differentiate toward neutrophils and subsequently die via apoptosis in vitro. In this paper, we investigated the roles of protein-tyrosine kinases (PTKs) in retinoic acid (RA)-induced granulocytic differentiation of HL-60 cells. Accompanying the RA-induced differentiation, activities of src family PTKs Lyn and Fgr became detected and reached a plateau 2 days after the stimulation. The immunoblotting using anti-phosphotyrosine antibody (PY-20) showed that the proteins of 56 and 53 kDa were predominantly tyrosine-phosphorylated at day 2. Adsorption and immunoprecipitation of the cell lysate by specific antibodies evidenced that these phosphotyrosine-containing proteins are Lyn and Fgr PTKs. The degree of both activities and tyrosine phosphorylation of these PTKs was reduced to be minimal at day 5 when the HL-60 cells start to die by apoptosis. The inhibitors of PTKs, herbimycin A and genistein, were demonstrated to cause premature cell death of HL-60 cells in the presence of RA. The death was the consequence of an apoptotic process. The Ra-treated HL-60 cells, when incubated with specific c-lyn or c-fgr antisense oligodeoxynucleotide, also underwent premature death at day 2. These data implicate that Lyn and Fgr PTKs prevent programmed cell death to promote granulocytic differentiation of HL-60 cells.

Signal transduction pathways that contribute to myeloid differentiation

“…The expression and activation of Lyn and Fgr have been found to be induced during ATRA-induced granulocytic differentiation of HL-60 cells. 190 Treatment with antisense oligonucleotides against lyn or fgr mRNA resulted in premature death of the differentiating cells. 190 Similarly, high levels of c-Src kinase expression and activity were seen in HL-60 and U937 cells undergoing PMA-induced monocytic differentiation.…”

“…190 Treatment with antisense oligonucleotides against lyn or fgr mRNA resulted in premature death of the differentiating cells. 190 Similarly, high levels of c-Src kinase expression and activity were seen in HL-60 and U937 cells undergoing PMA-induced monocytic differentiation. 191 Elevated expression of Fgr and Hck were also observed in AML leukemic blasts that were induced to differentiate along the monocytic or granulocytic lineages.…”

Interaction between growth arrest-DNA damage protein 34 and Src kinase Lyn negatively regulates genotoxic apoptosis

“…In addition to a role in DNA damage responses, Lyn plays a role in other processes such as glutamate receptor signaling in neurons (14), cytoskeletal rearrangement and motility in mast cells (15), cytokine responses in hematopoietic cells, and B lymphocyte receptor signaling. In hematopoietic cells, cytokine-mediated activation of Lyn stimulates proliferation (16)(17)(18) and inhibits apoptosis (19)(20)(21)(22). In B lymphocytes, Lyn negatively regulates B cell receptor signaling and promotes growth arrest, but not apoptosis (23,24).…”

Outgrowth of pseudopodial cables induced by all‐trans retinoic acid in micromere‐derived cells isolated from sea urchin embryos

“…In sea urchin eggs, the activity of protein tyrosine kinase is found to increase following fertilization (Ribot et al 1984;Satoh & Garbers 1985) and pulse treatment of eggs with genistein after fertilization results in the failure of gut formation in embryos (Kinsey 1995 (Giguère et al 1987;Petkovich et al 1987;Brand et al 1988;Krust et al 1989). In HL-60 cells, tRA induces the expression of fgr, syk and lyn, src family intracellular tyrosine kinases (Katagiri et al 1996) (Kuno et al 1994b) activates the receptor's protein tyrosine kinase and augments phosphorylation of the protein tyrosine residue of IRS-1 (for instance) in micromere-derived cells (Kuno & Yasumasu 1996), as it does in mammalian cells (Petruzzelli et al 1982(Petruzzelli et al , 1984Roth & Cassell 1982;Kasuga et al 1983 (Kuno & Yasumasu 1996) (Kuno et al 1994b …”