To function properly, intracellular signaling pathways depend upon appropriate and unique subcellular locations of their constituent proteins. Research in the Wedegaertner lab focuses on understanding cellular mechanisms of G protein signaling. Heterotrimeric G proteins (abg) are well known for their function in linking G protein-coupled receptors (GPCRs) to a variety of intracellular responses. In the textbook view, G proteins carry out their function while associated with the cytoplasmic surface of a cell’s plasma membrane (PM). In contrast to PM-limited G protein signaling, it has become increasing clear that G proteins can have important cellular functions at diverse intracellular sites. A major focus of the lab is to understand mechanisms that regulate membrane binding and trafficking to distinct subcellular locations for G proteins, and other signaling proteins, and to understand novel signaling functions at different subcellular locations.
Signaling by G Protein beta-gamma subunits at the Golgi
One research area in the lab addresses a recently appreciated non-canonical G protein function: the role of Gbg in regulating Golgi function and integrity. The Golgi is an essential organelle in directing the transport of secretory and plasma membrane (PM) proteins, as well as transport to other organelles. It has been estimated that one-third of the human proteome traverses the secretory pathway – from synthesis at the endoplasmic reticulum (ER), through the ER and Golgi apparatus, and then ultimately transport via vesicles or larger transport carriers to the PM or to intracellular organelles. Proper transport of transmembrane proteins to the PM and proper secretion of proteins is universally essential for cell function. Moreover, numerous disease states result from dysregulation of protein transport. Using a variety of cell biology techniques, including the development of novel tools for inducible recruitment of G proteins or G protein regulators to specific subcellular organelles, our lab has defined a role for signaling by Gbg subunits at the Golgi in regulating protein transport. In addition, he Golgi is a dynamic organelle that undergoes reversible breakdown (i.e., fragmentation and dispersal) under normal physiological conditions in cells, as well as in pathophysiological conditions, such as infection, cancer and neurodegenerative disease. Thus, our work is also addressing an unexpected role for Gbg subunits in regulating changes in Golgi morphology.
Signaling by mutant G alpha q/11 subunits in uveal melanoma
Uveal melanoma is the most common cancer of the eye in adults. In up to 50% of cases, uveal melanoma metastasizes to the liver, and patient survival is extremely low; no effective therapies have been developed for metastatic uveal melanoma. Strikingly, more than 90% of uveal melanomas harbor mutually exclusive activating mutations in the closely related heterotrimeric G protein a subunits Gaq or Ga11. Mutation in Gaq or Ga11 is an early event in uveal melanoma and is thus considered a driver mutation for this cancer. Our focus is to understand how mutational activation disrupts subcellular localization and dysregulates signaling of Gaq and Ga11 and to understand the mechanisms that govern intracellular trafficking of mutationally activated Gaq and Ga11. By understanding the unique subcellular trafficking mechanisms of mutationally activated Gaq and Ga11 in uveal melanoma cells, we hope to define novel therapeutic targets for disrupting trafficking and signaling by these mutationally activated G proteins.
Mechanisms of localization and trafficking of GRKs
G protein-coupled receptor kinases (GRKs) reversibly bind to cellular membranes to phosphorylate agonist-activated GPCRs. Due to the critical nature of properly regulating GPCR signaling, changes in the steady-state level, function or localization of a particular GRK can tip the balance towards too much or too little signaling and thus contribute to numerous disease states, such as heart failure, hypertension, cancer, opiate addiction, and mental health disorders. In addition to their classical function of GPCR phosphorylation, it has become increasingly clear that GRKs have important non-canonical functions to phosphorylate and interact with other proteins at diverse locations in the cell. Our focus has been on understanding key determinants in the GRK4, 5 and 6 sub-family that govern subcellular localization of these proteins. Our most recent work has identified a role for dimerization of GRK5 and GRK6 in regulating their PM localization, and we have identified differences in subcellular trafficking itineraries of GRK5 and GRK6 due to differences in post-translational lipid modification. Our goal is to better understand the mechanisms that regulate localization and trafficking of GRKs and thereby reveal novel therapeutic targets.