Src kinases and not JAKs activate STATs during IL-3 induced myeloid cell proliferation

Abstract

Interaction of IL-3 with its receptor is known to activate STAT-3 via phosphorylation of Tyrosine 701, which facilitates its dimerization and translocation to the nucleus, leading to the transcription of its target genes. In this communication, we have investigated the nature of tyrosine kinases that mediate STAT-3 phosphorylation during IL-3-mediated activation of myeloid cell proliferation. Our results show that interaction of IL-3 with its receptor leads to the activation of c-Src kinase activity, which in turn facilitates the binding of c-Src to STAT-3. This association leads to the phosphorylation of STAT-3, allowing this transcription factor to translocate to the nucleus. Expression of a dominant negative mutant of src (AMSrc) in these cells results in a block to IL-3 mediated phosphorylation of STAT-3, and its ability to bind to DNA. On the other hand, expression of a dominant negative mutant of JAK2 (JAK2KE) had no eect on IL-3-mediated activation of STAT-3. Our results also show that AMSrc does not aect the phosphorylation of JAK2, suggesting that JAK and STAT phosphorylation events are mediated by two independent pathways. Inhibition of c-Src activation by AMSrc, which leads to a block to STAT-3 activation, results in a dramatic inhibition of cell proliferation mediated by IL-3. However, expression of AMSrc does not activate apoptotic pathways. In contrast, expression of JAK2KE results in accelerated apoptosis of 32Dcl3 cells grown in the absence of IL-3 with concomitant down-regulation of Erk-2 kinase activity. These results suggest that Src family kinases mediate the phosphorylation of STATs and play a critical role in signal transduction pathways associated with myeloid cell proliferation while JAK kinases mediate the activation of Erk-2 pathway which appears to provide antiapoptotic signals. Thus the activation of JAKs and STATs appear to be two independent but related events, which dictate two separate biological outcomes, the combination of which results in proliferation and survival of myeloid precursor cells.

Engagement of the B-cell antigen receptor activates STAT through Lyn in a Jak-independent pathway

“…In this study, we found G-CSF-mediated STAT phosphorylation is not Lyn-dependent by means of antisense JAK or AG490 inhibition experiments. Consistent with this, overexpression of dominant-negative JAK2 had no effect on Srcmediated phosphorylation of STATs (Chaturvedi et al, 1998). Combined with our results, this suggests that two independent pathways exist that mediate STAT activation, one dependent on the JAKs and the other dependent on Src kinases.…”

Regulation by Gi2 proteins of v-fms-induced proliferation and transformation via Src-kinase and STAT3

“…Several groups have reported that v-src and Src-kinase can activate STAT3 (Cao et al, 1996;Campbell et al, 1997;Chaturvedi et al, 1998). In addition, Marrero et al (1995) reported that activation of STAT3 can be induced or enhanced by GCRs, in particular by angiotensin II AT 1 receptor.…”

“…As STAT3 has been shown to regulate mitogenesis or/and prevention of cell apoptosis in various cell types (Fukada et al, 1996;Chaturvedi et al, 1998) and 85 kDa STAT3D proteins are overexpressed in a i2 -G204A and a i2 -G204A/v-fms cells, we investigated whether excessive STAT3D expression inhibits NIH3T3 cell proliferation. Cells stably transfected with STAT3D55C cDNA showed increased expression of 85 kDa STAT3D (Figure 8), but also, as observed previously (Kim and Baumann, 1997), a slightly decreased expression of 89 kDa STAT3.…”

“…The regulation of Src-kinase activity by G i2 proteins is speci®c: Fyn expression and activity were not altered in cells expressing a i2 -G204A, indicating that Fyn did not compensate for Src in signalling leading to cell proliferation. Indeed, the role played by Src in the regulation of cell proliferation is thought to be essential: it was reported that Src stimulates JAK1 and STAT3 (Cao et al, 1996 ;Campbell et al, 1997;Chaturvedi et al, 1998) and also that impaired ®broblast proliferation related to de®cient Src-kinase activity could be rescued by c-myc (Roche et al, 1995b). In our model, we found no evidence linking Src activity and c-myc expression; v-fms expression increased c-myc levels to similar levels in cells with active or inactive G i2 proteins (i.e., intact or de®cient Src-kinase activity).…”

“…STAT3 is activated by phosphorylation on tyrosine residues by a number of cytokines (including IL-3, IL-6, CSF-1) and STAT3 activation by IL-3 or IL-6 requires Src-kinase (Chaturvedi et al, 1998;Wang et al, 1999). Similarly, we found that v-fms induces constitutive tyrosine phosphorylation of STAT3 in cells with active G i2 and Src proteins, but not in cells with de®cient G i2 and/or Src proteins, indicating that v-fms requires Src to activate STAT3 and that Src-mediated activation of STAT3 may be the decisive event regulated by G i2 proteins in the transduction of CSF-1R signalling.…”