Insulin resistance (IR) is a metabolic disorder characterized by impaired glucose uptake in response to insulin. The current paradigm for insulin signaling centers upon the insulin receptor (InsR) and its substrate IRS1; the latter is believed to be the chief conduit for post-receptor signaling. We recently demonstrated that GIV, a Guanidine Exchange Factor (GEF) for the trimeric G protein, Gαi, is a major hierarchical conduit for the metabolic insulin response. By virtue of its ability to directly bind the InsR, IRS1 and PI3K, GIV enhances the InsR-IRS1-Akt-AS160 (RabGAP) signaling cascade and cellular glucose uptake via its GEF function. Phosphoinhibition of GIV-GEF by the fatty-acid/PKCθ pathway inhibits the cascade and impairs glucose uptake. Here we show that GIV directly and constitutively binds the exocyst complex subunit Exo-70 and also associates with GLUT4-storage vesicles (GSVs) exclusively upon insulin stimulation. Without GIV or its GEF function, membrane association of Exo-70 as well as exocytosis of GSVs in response to insulin are impaired. Thus, GIV is an essential component within the insulin signaling cascade that couples upstream signal transducers within the InsR and G-Protein signaling cascade to downstream vesicular trafficking events within the exocytic pathway. These findings suggest a role of GIV in coordinating key signaling and trafficking events of metabolic insulin response.

Environmental cues are transmitted to the interior of the cell via a complex network of signaling hubs. Receptor tyrosine kinases (RTKs) and trimeric G proteins are two such major signaling hubs in eukaryotes. Conventionally, canonical signal transduction via trimeric G proteins is thought to be triggered exclusively by G protein-coupled receptors. Here we used molecular engineering to develop modular fluorescent biosensors that exploit the remarkable specificity of bimolecular recognition, i.e., of both G proteins and RTKs, and reveal the workings of a novel platform for activation of G proteins by RTKs in single living cells. Comprised of the unique modular makeup of guanidine exchange factor Gα-interacting vesicle-associated protein (GIV)/girdin, a guanidine exchange factor that links G proteins to a variety of RTKs, these biosensors provide direct evidence that RTK-GIV-Gαi ternary complexes are formed in living cells and that Gαi is transactivated within minutes after growth factor stimulation at the plasma membrane. Thus, GIV-derived biosensors provide a versatile strategy for visualizing, monitoring, and manipulating the dynamic association of Gαi with RTKs for noncanonical transactivation of G proteins in cells and illuminate a fundamental signaling event regulated by GIV during diverse cellular processes and pathophysiologic states.

In eukaryotes, receptor tyrosine kinases (RTKs) and trimeric G proteins are two major signaling hubs. Signal transduction via trimeric G proteins has long been believed to be triggered exclusively by G protein-coupled receptors (GPCRs). This paradigm has recently been challenged by several studies on a multimodular signal transducer, Gα-Interacting Vesicle associated protein (GIV/Girdin). We recently demonstrated that GIV's C terminus (CT) serves as a platform for dynamic association of ligand-activated RTKs with Gαi, and for noncanonical transactivation of G proteins. However, exogenous manipulation of this platform has remained beyond reach. Here we developed cell-permeable GIV-CT peptides by fusing a TAT-peptide transduction domain (TAT-PTD) to the minimal modular elements of GIV that are necessary and sufficient for activation of Gi downstream of RTKs, and used them to engineer signaling networks and alter cell behavior. In the presence of an intact GEF motif, TAT-GIV-CT peptides enhanced diverse processes in which GIV's GEF function has previously been implicated, e.g., 2D cell migration after scratch-wounding, invasion of cancer cells, and finally, myofibroblast activation and collagen production. Furthermore, topical application of TAT-GIV-CT peptides enhanced the complex, multireceptor-driven process of wound repair in mice in a GEF-dependent manner. Thus, TAT-GIV peptides provide a novel and versatile tool to manipulate Gαi activation downstream of growth factors in a diverse array of pathophysiologic conditions.

Insulin resistance (IR) is a metabolic disorder characterized by impaired insulin signaling and cellular glucose uptake. The current paradigm for insulin signaling centers upon the insulin receptor (InsR) and its substrate IRS1; the latter is believed to be the sole conduit for postreceptor signaling. Here we challenge that paradigm and show that GIV/Girdin, a guanidine exchange factor (GEF) for the trimeric G protein Gαi, is another major hierarchical conduit for the metabolic insulin response. By virtue of its ability to directly bind InsR, IRS1, and phosphoinositide 3-kinase, GIV serves as a key hub in the immediate postreceptor level, which coordinately enhances the metabolic insulin response and glucose uptake in myotubes via its GEF function. Site-directed mutagenesis or phosphoinhibition of GIV-GEF by the fatty acid/protein kinase C-theta pathway triggers IR. Insulin sensitizers reverse phosphoinhibition of GIV and reinstate insulin sensitivity. We also provide evidence for such reversible regulation of GIV-GEF in skeletal muscles from patients with IR. Thus GIV is an essential upstream component that couples InsR to G-protein signaling to enhance the metabolic insulin response, and impairment of such coupling triggers IR. We also provide evidence that GIV-GEF serves as therapeutic target for exogenous manipulation of physiological insulin response and reversal of IR in skeletal muscles.

Background and aims

Lipoprotein(a) [Lp(a)] is causally associated with aortic valve stenosis (AS) but Lp(a) testing among AS patients is not broadly incorporated into clinical practice. We evaluated trends in Lp(a) testing in an academic medical center.

Methods

Educational efforts and adding Lp(a) to the lipid panel on the electronic medical record (EMR) and pre-procedure order sets were used to increase awareness of Lp(a) as a risk factor in AS. Medical records at University of California San Diego Health (UCSDH) were analyzed from 2010 to 2020 to define the yearly frequency of first time Lp(a) testing in patients with diagnosis codes for AS or undergoing transcatheter aortic valve replacement (TAVR).

Results

Lp(a) testing for any indication increased over 5-fold from 2010 to 2020. A total of 3808 patients had a diagnosis of AS and 417 patients had TAVR. Lp(a) levels >30 mg/dL were present in 37% of AS and 35% of TAVR patients. The rates of Lp(a) testing in AS and TAVR were 14.0% and 65.7%, respectively. In AS, Lp(a) testing increased over time from 8.5% in 2010, peaking at 24.2% in 2017, and declining to 13.9% in 2020 (p < 0.001 for trend). Following implementation of EMR order-sets in 2016, Lp(a) testing in TAVR cases increased to a peak of 88.5% in 2018.

Conclusions

Elevated Lp(a) is prevalent in AS and TAVR patients. Implementation of educational efforts and practice pathways resulted in increased Lp(a) testing in patients with AS. This study represents a paradigm that may allow increased global awareness of Lp(a) as a risk factor for AS.