Gitte Wolfsberg Haxholm:
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Class 1 Cytokine receptors are involved in important biological functions mediated through complex networks of intracellular signaling. However, the molecular details of how signaling is regulated are poorly understood. One of the primary reasons for this limited knowledge is the lack of structural information on the intracellular domains (ICDs) of these receptors. The overall aim of this study was to obtain an improved understanding of cytokine receptor signaling through structure-function studies on the prolactin receptor (PRLR).
The primary focus of this thesis was to structurally characterize the PRLR-ICD and the ICD of the related growth hormone receptor (GHR). We showed that both ICDs were intrinsically disordered throughout their entire lengths and that they associated with lipids characteristic of the inner plasma membrane leaflet through conserved motifs, implicating the membrane as an active component during signaling. Based on these results we proposed a model where cytokine receptor ICDs associate with the membrane to confine intracellular signaling and hence increase the efficiency of signaling. PRLR signaling is further regulated by protein-protein interactions and post-translational modifications (PTMs), but the molecular insight into these processes is very sparse. We initiated a project to map the phosphorylation pattern of the PRLR in cells and further identified the PRLR-ICD as a novel potential target for direct associations with a mitogen activated protein kinase (MAPK), leading to subsequent phosphorylation of the receptor.
A minor part of this thesis focused on the role of two residues located in the dimer-interface of the extracellular domains (ECDs) in PRLR signaling. We showed that a single residue had the capacity to regulate signaling through the MAPK pathway in vivo and suggested an in vitro correlation between the life-times of 1:2 hormone/PRLR-variant complexes and the stabilities of isolated PRLR-ECD variants with activation of the MAPK pathway.
Conclusively, the large extent of disorder uncovered in these ICDs calls for a rethinking of how to address future cellular as well as structural studies of cytokine receptors. The results presented in this thesis have the potential to inspire future studies on how specific associations and PTMs affect PRLR signaling.