Prolactin stimulates the JAK2 and focal adhesion kinase pathways in human breast carcinoma T47-D cells

Abstract

Treatment of T47-D human breast carcinoma cells with recombinant prolactin (rhPRL) induced a concentration- and time-dependent increase in the phosphotyrosine content of JAK2. rhPRL also stimulated JAK2 tyrosine phosphorylation more weakly in three other breast carcinoma lines, MCF-7, ZR-75-1 and MDA-MB-231. Furthermore it stimulated tyrosine phosphorylation of two isoforms of the transcriptional activator STAT5, STAT5a and STAT5b. Surprisingly, rhPRL treatment of T47-D cells also stimulated tyrosine phosphorylation of focal adhesion kinase (FAK), as determined by immunoprecipitation with anti-phosphotyrosine antibody followed by immunoblotting with a specific FAK antibody. The effect of rhPRL was rapid and concentration-dependent, being maximal at 5 ng/ml. At rhPRL concentrations above 25 ng/ml, FAK tyrosine phosphorylation declined but remained above control levels at 100 ng/ml. rhPRL also stimulated paxillin tyrosine phosphorylation in T47-D cells with similar concentration- and time-dependence. In a second human breast carcinoma cell line, MCF-7, rhPRL produced very similar effects on FAK and paxillin tyrosine phosphorylation. These findings identify a new protein tyrosine kinase pathway in the action of the lactogenic hormone rhPRL and represent the first report that a hormone acting through a member of the haemopoietin receptor superfamily can regulate the FAK/paxillin pathway.

Nek3 kinase regulates prolactin-mediated cytoskeletal reorganization and motility of breast cancer cells

“…In the present study, we identify a novel PRL-dependent interaction between the serine/threonine kinase Nek3 and paxillin (Figure 5a). Although paxillin has been shown to be tyrosine phosphorylated in response to PRL stimulation (Canbay et al, 1997;Montes et al, 2005), the data presented in this study are the first report of paxillin serine phosphorylation in response to PRL stimulation. Untransfected and control siRNA T47D transfected (Figure 5b and data not shown) showed a significant increase in paxillin serine phosphorylation in the presence of PRL.…”

“…PRL treatment of T47D cells has been shown to result in tyrosine phosphorylation of paxillin (Canbay et al, 1997;Acosta et al, 2003); however, there are no reports of changes in paxillin serine/threonine phosphorylation in response to PRL. To evaluate this, untransfected and control siRNAtransfected T47D cells were stimulated with PRL, and anti-paxillin immunoprecipitates were examined for serine phosphorylation (Figure 5b).…”

Regulation of Neutrophil Adhesion by Pituitary Growth Hormone Accompanies Tyrosine Phosphorylation of Jak2, p125FAK, and Paxillin

“…It is noteworthy to add that a small amount of constitutive tyrosine phosphorylation of Jak2 and p125 FAK was observed in neutrophils without any stimulation. In comparison with other experiments, the constitutive tyrosine phosphorylation of Jak2 and p125 FAK in our system might be due to the difference in cell types (23,27,45). Inhibition of neutrophil adhesion and tyrosine phosphorylation of Jak2, p125 FAK , and paxillin by genistein implies that GH-induced neutrophil adhesion is related to the pattern of tyrosine phosphorylation of these signaling molecules.…”

Human prolactin (hPRL) antagonists inhibit hPRL-activated signaling pathways involved in breast cancer cell proliferation

“…Recently, the group of Clevenger showed that hPRL is chemoattractant for human breast carcinomas, and may thereby facilitate breast cancer metastasis, which frequently arise from in lymph node, a site rich in Tlymphocytes capable of producing hPRL (Maus et al, 1999). Such a hPRL-induced cell mobility may involve the Focal Adhesion Kinase pathway, whose activation by hPRL has been shown in T-47D cells (Canbay et al, 1997). Since G129R-hPRL acts by interfering with functional receptor homodimerization (Go n et al, 1996b; Kinet et al, 1999), a step which appears to be a pre-requisite for most (if not all) hPRLR-triggered e ects, it is reasonable to anticipate that the analog will also antagonize these hPRL-induced phenomena, although this assumption awaits experimental evidence.…”

“…In mammary epithelial cells (tumor and normal), PRL induces phosphorylation (and activation) of the PRLR-associated tyrosine kinase JAK2, which in turn phosphorylates latent cytoplasmic transcription factors STAT5, STAT3 and STAT1 (DaSilva et al, 1996;Schaber et al, 1998). Other pathways are also activated by PRL in mammary epithelial cells, which involve the Ras/Raf/MAP kinase (Das and Vonderhaar, 1996a,b;Schaber et al, 1998), the Protein Kinase C (Waters and Rillema, 1989;Banerjee and Vonderhaar, 1992) and the Focal Adhesion Kinase-paxillin (Canbay et al, 1997) pathways (for a review, Llovera et al, 2000). Based among others on the use of speci®c kinase inhibitors, it has been shown that the JAK/STAT and MAPK pathways play a critical role in PRL-induced proliferation of mammary epithelial cells Vonderhaar, 1996a,b, 1997).…”