G protein-coupled signal transduction systems

Hébert Lab, Department of Pharmacology and Therapeutics, McGill University


RESEARCH PROJECTS

Research in my lab is centered broadly around the theme of G protein-coupled signal transduction systems. These signalling systems are activated by agonists that bind to G protein-coupled receptors (GPCRs) leading to the regulation of effector proteins (e.g. enzymes and ion channels) by a transducer. We are interested in 1) basic mechanisms of how these signalling systems are wired, 2) novel signalling complexes and pathways associated with alternative subcellular localization of GPCRs and 3) the roles that these architectural features of signalling complex design might play in cardiac disease with a particular emphasis on congenital heart disease. More recently, my group has also become interested in how novel allosteric regulators of these receptors might be developed and tested, here with a focus on inflammatory mediators, their receptors and their roles in health and disease. All of the projects are currently funded by CIHR, the first three by individual grants as PI and the latter as part of a CIHR Team in GPCR allosteric regulation.

 
Sorting and assembly decisions in GPCR signalling complex assembly

 

1) Hard-wiring of 7TM-R signalling complexes. Some 7TMs regulate the activity of multiple effectors. The majority of drugs that target G protein-coupled signal transduction systems act at ligand binding sites. A therapeutic strategy may require regulating the activity of a specific effector, but drugs aimed at 7TMs coupled to multiple effectors obviously lack specificity in this regard, and produce undesirable side effects. Thus, a primary objective of our research is to identify peptide motifs involved in specific protein-protein interactions yielding a novel strategy for therapeutic intervention- the modulation of specific receptor complexes by disrupting the interactions which lead to their formation and trafficking or augmenting interactions that lead to signal transduction with peptidic or peptidomimetic compounds. This work has been continually funded by both CIHR and HSFQ since I began my independent career in 1996.


GPCRs and their signalling machinery can be targetted to the nuclear membrane
(taken from Neumann 2006, Cardiovasc. Res. 71:6-7)

 

2) Alternate subcellular destinations for 7TM-R signalling complexes. An increasing number of GPCRs have also been demonstrated to be targeted to the endomembrane locations such as the nucleus as have their associated signalling cascades, which is an active area of research in my lab. Thus, the signalling landscape inside the cell may be as complex as that arranged to detect and interpret signals from outside the cell. It is clear that different organelles must communicate in real-time on a variety of distinct time-scales using a number of intracrine signalling pathways. The difficulty in this research has been to tease out the effects of nuclear receptors from those in the cell surface. We have developed new ligands for GPCRs, that are caged in a way that allows them to enter the cell and only there be converted into the active form. With these molecules, we can evaluate the contribution of nuclear GPCRs to cellular signalling in the context of intact cells, tissues and living animals. Recent work has also shown that G subunits are also key regulators of cellular signalling events but also serve a broader role in organizing the assembly and trafficking of receptor-based complexes in intracellular compartments such as the ER and Golgi apparatus. Recently, a number of studies have indicated a direct nuclear impact for Gβγ subunits. This novel role for Gβγ subunits as transcriptional regulators- may be potentially independent of classical functions as mediators of GPCR signalling. We are examining connections between events driven by receptor-mediated signalling at the cell surface and nucleus and between these systems and potentially receptor-independent functions of Gβγ in the nucleus. We are also examining the roles on different Gβγ pairs using a genetic approach in C. elegans. This work is funded by a 5-year CIHR grant initially obtained in 2006 shortly after my arrival at McGill which I have just renewed until 2017. Also, the studies in C. elegans have also recently been funded by NSERC from 2012-2017.



Functional selectivity in GPCR dimers when different protomers
are occupied by ligand.


3) Allosteric Regulation of GPCRs.
I have been investigating basic mechanisms of GPCR regulation, developing functional assays amenable to scaling up and for use as screens for novel modulators of the interactions within GPCR signalling complexes. We have already identified a series of candidates for allosteric modulators of FP (the receptor for prostaglandin F2α). GPCRs are the targets of a large number of the drugs used to treat human disease. Our current understanding of how these proteins function indicates that they are extremely dynamic- with respect to how they interact with their natural activators and the drugs which target them as well as with their partners which effect changes in cell function downstream. As part of two large team grants (CIHR 2006-2011 and CQDM 2009-2012) we have developed a number of biosensors that can interrogate large sections of the signalling spectrum or signature of a given GPCR. Further, although a great deal of structural information has recently been obtained about these receptors, conformational information has not been combined in modern screening campaigns for new drugs. Screening approaches can incorporate structural considerations designed to understand receptor function, but are limited in the sense that they can only provide snapshots of these events, rather than “films” which capture the dynamic aspects of these receptors in the environment of a living cell. We are adding a robust element of dynamics to drug screening and drug discovery by engineering conformation-sensitive fluorescent reporters into numerous positions on the receptor. With these tools, we can evaluate both interactions with receptor-binding drugs and with receptor signalling partners in a new way, which captures conformational information, as a complement to the signaling signatures. This research is funded by personal grants from the CQDM and is the focus of several team grants from CIHR and CQDM.