Receptors for growth hormone and prolactin. Regulation and activation

Sammanfattning: Cytokine receptor activation is dependent on ligand-induced receptor dimerization or oligomerization and subsequent activation of several intracellular proteins. Activation of Janus kinases (Jaks) is probably the first intracellular event that occurs after receptor dimerization. Signaling through both the growth hormone receptor (GHR) and the prolactin receptor (PrlR) is believed to be dependent on activation of Jak2; however, in vitro studies indicate that other Jak proteins can be phosphorylated in response to GH and Prl. The amount of biologically active hormone in the circulation, the number of functional receptors in the target tissue and factors involved in intracellular signaling determine the response to a particular cytokine. The general aim of this thesis was to study mechanisms that might influence the effects of GH and Prl in target tissues, and to produce a reagent for use in a cell-free assay to analyze biologically active GH.The presence of all known Jak proteins was demonstrated in human liver, rat liver and rat adipose tissue. Co-precipitation studies showed that Jak1, Jak2 and Tyk2 were associated with the GHR in human liver, whereas Jak1 and Jak2 interacted with the GHR in rat liver and rat adipose tissue. Furthermore, the relative amount of GHR-associated Jak1 and Jak2 differed markedly between rat liver and rat adipose tissue. These results indicate that Jak proteins other than Jak2 might influence GH signaling, in a tissue- and species-specific manner.Conditions involving an increased catabolic rate are believed to be associated with acquired GH insensitivity. The effects of surgically induced catabolism on GH sensitivity were investigated in patients undergoing elective abdominal surgery. In adipose tissue and skeletal muscle, changes in GHR expression correlated with changes in insulin-like growth factor I (IGF-I) expression before surgery compared with 3 days after surgery. IGF-I gene expression did not change in response to surgery in skeletal muscle. In contrast, IGF-I mRNA levels increased in adipose tissue, suggesting tissue differences in GH sensitivity. The 5'-untranslated regions (UTRs) in GHR transcripts differed between skeletal muscle and adipose tissue, suggesting that alternative promoter regions may be activated in these tissues during catabolism. Although PrlRs have not been identified in adipose tissue, there are indications that Prl may exert important metabolic actions on this tissue. We have detected expression of the long (L-PrlR) form and two short forms (S2-PrlR and S3-PrlR) of the PrlR in mouse adipose tissue. L-PrlR expression was higher in adipose tissue from lactating mice compared with male, virgin and pregnant mice. Furthermore, L-PrlR mRNA expression was increased in adipose tissue in Prl-transgenic mice. Concentrations of cytokines are usually determined by antibody-based assays that do not distinguish between biologically active and inactive isoforms. Bioassays are laborious and not suitable for analyses of multiple samples. As a first step in the development of an assay for measurement of bioactive GH, we have constructed a soluble hybrid receptor consisting of the extracellular domain of the GHR and the intracellular domain of the receptor for epidermal growth factor, containing an intrinsic tyrosine kinase domain. The hybrid receptor had GH-binding capacity as well as GH-induced tyrosine kinase activity. This provides a novel tool in the development of assays for bioactive GH, and the same concept may also be used for measurement of other cytokines.

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