Prolactin receptor is associated with c-src kinase in rat liver.

We have demonstrated here that growth hormone (GH) stimulates the formation of the active GTP-bound form of both RalA and RalB in NIH-3T3 cells. Full activation of RalA and RalB by GH required the combined activity of c-Src and JAK2, both kinases activated by GH independent of the other. Activation of RalA and RalB by growth hormone did not require the activity of JAK2 per se. Ras was also activated by GH and was required for the GH-stimulated formation of GTP-bound RalA and RalB. Activation of RalA by GH subsequently resulted in increased phospholipase D activity and the formation of its metabolite, phosphatidic acid. GH-stimulated RalA-phospholipase D-dependent formation of phosphatidic acid was required for activation of p44/42 MAPK and subsequent Elk-1-mediated transcription stimulated by GH. Thus we report the identification of a JAK2-independent pathway regulating GH-stimulated p44/42 MAPK activity.Due to lack of intrinsic kinase activity, members of the cytokine receptor superfamily, including the growth hormone (GH) 1 receptor, recruit and activate non-receptor tyrosine kinases of the JAK family to relay their cellular signals (1). JAK2 has been reported to be the predominant JAK required for the initiation of GH signal transduction upon ligand binding to the GH receptor (2-4). To date, all identified downstream signaling pathways utilized by GH apparently require JAK2 activity (2-4). The only reported JAK2-independent effect of GH is Ca 2ϩ entry via L-type calcium channels (5), although this has been disputed (6, 7). However, it is likely that other, as yet uncharacterized, signal transduction pathways stimulated by GH are activated independent of JAK2 activity.The major groups of signaling molecules thus far identified to be activated by GH include the following: 1) other receptor (EGF receptor) (8) and non-receptor (c-Src, c-Fyn (9), and FAK (10)) kinases, although as in the case of the EGF receptor it may be used simply as an adapter protein; 2) members of the MAP kinase family including p44/42 MAP kinase (11, 12), p38 MAP kinase (13), and c-Jun N-terminal kinase/stress-activated protein kinase (9) and the respective downstream pathways; 3) members of the insulin receptor substrate (IRS) group including IRS-1, -2, and -3 which may act as docking proteins for further activation of signaling molecules including phosphatidylinositol 3-kinase (14); 4) small Ras-like GTPases (15); and 5) STAT family members including STATs 1, 3, 5a, and 5b (16, 17), which constitute one major mechanism for transcriptional regulation by GH.Ras is a member of the Ras-like GTPase family (18,19). This family is characterized by similarities in the effector domain which Ras utilizes to interact with downstream target molecules. The Ras-like GTPases play a critical role in multiple signaling pathways leading from various cell-surface receptors. The activation and inactivation of the Ras-like GTPases are controlled by conformational change because of a GTP-GDP binding cycle that is controlled by the following three different reg...

Section: Hgh Stimulation Of Nih-3t3 Cells Increases the Level Of Gtp-mentioning

confidence: 99%

Identification of a JAK2-independent Pathway Regulating Growth Hormone (GH)-stimulated p44/42 Mitogen-activated Protein Kinase Activity

Zhu¹,

Ling²,

Lobie³

2002

“…Identification of Csk among our positive clones suggests that molecules of the Src family (29) could be involved in GHR signaling. Two Src family molecules, c-Src and Fyn, have been reported to be involved in signal transduction of prolactin receptor (44,45) which exhibits strong structural and functional similarities with the GHR (46). The molecule Grb4, whose full-length cDNA has not yet been cloned (18), as well as the Shb adaptor (30), are newly identified molecules; their role in signaling is unclear.…”

Section: Figmentioning

confidence: 99%

Grb10 Identified as a Potential Regulator of Growth Hormone (GH) Signaling by Cloning of GH Receptor Target Proteins

Moutoussamy

Renaudie

Lago

et al. 1998

The cloning of receptor targets procedure, used so far to identify proteins associated with tyrosine kinase receptors was modified to clone SH2 proteins able to bind to the growth hormone receptor (GHR). The cytoplasmic region of GHR, a member of the cytokine receptor superfamily does not contain tyrosine kinase activity. It was thus phosphorylated in bacteria by the Elk tyrosine kinase and radiolabeled to screen a mouse expression library. With this probe, we identified Shc and the p85 subunit of phosphatidylinositol 3-kinase as direct targets of the receptor. The other proteins identified, Csk, Shb, Grb4, and Grb10 are new potential transducers for cytokine receptors. We show in Huh-7 hepatoma cells that Grb10 and GHR associate under GH stimulation. Co-transfections in 293 cells further show that Grb10 interacts with both the GHR and Jak2. Functional tests demonstrate that Grb10 inhibits transcription of two reporter genes containing, respectively, the serum response element of c-fos and the GH response element 2 of the Spi2.1 gene, whereas it has no effect on a reporter gene containing only Stat5 binding elements. Our results suggest that Grb10 is a new target for a member of the cytokine receptor family that down-regulates some GH signaling pathways downstream of Jak2 and independently of Stat5.

“…Although genetic studies have established a critical role for Stat5 in mouse mammary gland differentiation (6,7), corresponding genetic evidence is not yet available for Jak2, because Jak2 null mice die in utero (8,9) and conditional Jak2 null mice have not been established. Although Jak2 has been regarded as the principal tyrosine kinase activated by prolactin (3), the picture has been complicated by evidence that prolactin also can activate other tyrosine kinases, including Src family kinases (10,11), focal adhesion kinase (12), Tec kinase (13), and the ErbB-2 receptor tyrosine kinase (14). Experimental testing of the importance of Jak2 for prolactin-induced differentiation of mammary epithelial cells is therefore warranted.…”

mentioning

confidence: 99%

Role of Tyrosine Kinase Jak2 in Prolactin-induced Differentiation and Growth of Mammary Epithelial Cells

Xie

LeBaron

Nevalainen

et al. 2002

Genetic studies in mice have established a critical role for prolactin receptors and transcription factor Stat5 in mammary gland differentiation. However, the enzymatic coupling between prolactin receptors and Stat5 in this process has not been established. In addition to Jak2, several other tyrosine kinases reportedly also are associated with prolactin receptors and may phosphorylate Stat5. Because Jak2 null mice die in utero, we targeted Jak2 in an ex vivo model of prolactin-induced mammary epithelial cell differentiation to determine the role of Jak2 in regulation of cell differentiation and growth. Two independent targeting strategies were used to suppress Jak2 in immortalized HC11 mouse mammary epithelial cells: 1) stable expression of a specific Jak2 antisense construct and 2) adenoviral delivery of a dominant-negative Jak2 gene. We now demonstrate that Jak2 is essential for prolactin-induced differentiation and activation of Stat5 in normal mouse mammary epithelial cells. Furthermore, suppression of Jak2 in HC11 cells was associated with constitutive activation of oncoprotein Stat3 and a hyperproliferative phenotype characterized by increased mitotic rate, reduced apoptosis, and reduced contact inhibition. Collectively, our data suggest that Jak2 is differentiation-inducing and growth-inhibitory in normal mammary epithelial cells, observations that may shed new light on the role of the Jak2-Stat5 pathway in breast cancer.