Involvement of PI3K-Dependent Signal Transduction During the Integrated Control of Basal and Ligand-Biased Pituitary Cell Functions

• PI3K-Dependent Signalling and the Regulation of Pituitary Cell Functions

• Author / Creator

  Pemberton, Joshua G

• Within multicellular organisms, individual cells must communicate with one another in order to coordinate physiological functions. One of the ways this is achieved is through the production and release of chemical messengers, called hormones. Hormones alter the functions of their target cells by binding to receptors that are often present on the cell-surface to regulate the activity of intracellular messaging proteins that control cellular responses. This process is collectively referred to as signal transduction. The ability of receptors to engage distinct signal transduction mechanisms following hormone binding allows for the hormone-selective regulation of cells and intracellular functions. Recent studies reveal that closely related hormones can selectively activate a distinct subset of signal transduction responses through shared receptors in a process called biased signalling. Interestingly, multiple variants of a single hormone are found in almost all organisms; however, the extent to which naturally occurring hormone variants utilize biased signalling during the integrated control of cell physiology is poorly understood. To explore the possibility of signalling bias within natural hormone families, my thesis work characterizes the signal transduction mechanisms utilized by two endogenous variants of gonadotropin-releasing hormone (GnRH; called GnRH2 and GnRH3) using a pituitary cell primary culture system established in a classical vertebrate model of hormonal communication, the goldfish (Carassius auratus). In general and across vertebrates, GnRHs reliably stimulate the release of luteinizing hormone (LH) and growth hormone (GH) from pituitary gonadotropes and somatotropes, respectively, to coordinate reproduction and somatic growth. Using hormone release and cellular hormone content as physiologically relevant endpoints, results demonstrate that different natural variants of GnRH utilize overlapping, and yet distinct, intracellular signal transduction mechanisms to control diverse aspects of pituitary cell function. In particular, I identify signal transduction mediated by the phosphoinositide 3-kinase (PI3K) superfamily as important for the integrated regulation of hormone release and production. These studies include the first evidence of biased intracellular regulation of the classical receptor-coupled class I PI3K isoforms, as well as demonstrate heterogeneity in the transduction systems involved downstream of PI3K-dependent signalling within goldfish gonadotropes and somatotropes. Lastly, results indicate that the highly-conserved mitogenic Raf-MEK-ERK cascade, as well as common PI3K-dependent signal transduction elements, likely interact to selectively affect GnRH-stimulated as well as basal hormone release and total availability in a cell type-, time-, and GnRH-dependent manner. Overall, the results presented in this thesis represent the first comprehensive examination of PI3K-dependent signalling in any pituitary cell type, including the only characterization of isoform-selective involvement of PI3Ks during regulated pituitary hormone exocytosis. The experiments performed also identify class I PI3Ks as evolutionarily conserved signalling targets of the G protein-coupled receptor superfamily and outline, for the first time, the versatility of PI3Ks as an intracellular platform for mediating biased cellular responses. Taken together, data presented in this thesis provide several novel findings that illustrate that the biased actions of individual GnRH variants leads to differences in their ability to control acute and long-term hormone release responses, as well as facilitate the selective regulation of cellular hormone availability. More generally, these studies represent the first steps in understanding how the complexity of biased signalling contributes to the endocrine control of reproduction and growth by selectively controlling pituitary cell functions. Furthermore, by using a basal vertebrate model to study biased signalling by natural hormone variants at the intracellular level, my doctoral research has provided important evolutionary insights into the molecular mechanisms that couple receptor activation to biased intracellular signal transduction responses and reveal how this phenomenon can regulate cell functions that can ultimately impact whole-organism physiology.

• Subjects / Keywords

  • Signal Transduction
  • Carassius auratus
  • Neuroendocrinology
  • G Protein-Coupled Receptor
  • Phosphoinositide 3-Kinase
  • Gonadotropin-Releasing Hormone
  • Pituitary

• Graduation date

  Fall 2015

• Type of Item

  Thesis

• Degree

  Doctor of Philosophy

• DOI

  https://doi.org/10.7939/R36D5PR78

• License

  This thesis is made available by the University of Alberta Libraries with permission of the copyright owner solely for non-commercial purposes. This thesis, or any portion thereof, may not otherwise be copied or reproduced without the written consent of the copyright owner, except to the extent permitted by Canadian copyright law.