Biology subjects

Keeshen, C. M.

Publications and source records attributed to Keeshen, C. M..

3 recordsLinked to original sources

bioRxiv2024

G protein-Coupled Receptor Associated Sorting ProteinGASP1 mediated trafficking of the Glucagon Like Peptide-1Receptor contributes to the development of tolerance toincretin drugs

Incretin mimetic drugs are in widespread use for the treatment of type 2 diabetes and obesity and more recently have been prescribed for weight loss in otherwise healthy individuals. These drugs are all agonists of the glucagon-like peptide 1 receptor (GLP-1R) and function by supplementing effects produced by the endogenous hormone agonist glucagon-like peptide 1 (GLP-1). The therapeutic benefits of these medications, including improved glucose control and weight loss, require continued usage and wane with time. The molecular mechanisms underlying this loss of effect to incretin drugs remain unknown. Following activation by agonist and signaling to G protein, the GLP-1R engages arrestins and is endocytosed. Here we investigated the role of G protein-coupled receptor associated sorting protein 1 (GASP1), a critical regulator of the post-endocytic trafficking of GLP-1R, on tolerance to GLP-1R agonist drug. We found that tolerance to incretin drug was prevented at the cellular, tissue and whole animal level in mice with a selective disruption of the GASP1 protein in beta cells of the pancreatic islet. These studies implicate post-endocytic sorting of the GLP-1R in the loss of effectiveness of incretin therapeutics with prolonged use. These findings also suggest a novel strategy to prevent tolerance by biasing incretin drugs for G protein and away from arrestin engagement.

cell biology↗

bioRxiv2022

CRISPR activation rescues abnormalities in SCN2A haploinsufficiency-associated autism spectrum disorder

The majority of autism spectrum disorder (ASD) risk genes are associated with ASD due to haploinsufficiency, where only one gene copy is functional. Here, using SCN2A haploinsufficiency, a major risk factor for ASD, we show that increasing the expression of the existing functional SCN2A allele with CRISPR activation (CRISPRa) can provide a viable therapeutic approach. We first demonstrate therapeutic potential by showing that restoring Scn2a expression in adolescent heterozygous Scn2a conditional knock-in mice rescues electrophysiological deficits associated with Scn2a haploinsufficiency. Next, using an rAAV-CRISPRa based treatment, we restore electrophysiological deficits in both Scn2a heterozygous mice and human stem-cell-derived neurons. Our results provide a novel therapeutic approach for numerous ASD-associated genes and demonstrate that rescue of Scn2a haploinsufficiency, even at adolescent stages, can ameliorate neurodevelopmental phenotypes.

neuroscience↗

bioRxiv2021

Paradoxical hyperexcitability from NaV1.2 sodium channel loss in neocortical pyramidal cells

Loss-of-function variants in the gene SCN2A, which encodes the sodium channel NaV1.2, are strongly associated with autism spectrum disorder and intellectual disability. An estimated 20-30% of children with these variants are co-morbid for epilepsy, with altered neuronal activity originating in neocortex, a region where NaV1.2 channels are expressed predominantly in excitatory pyramidal cells. This is paradoxical, as sodium channel loss in excitatory cells would be expected to dampen neocortical activity rather than promote seizure. Here, we examined pyramidal neurons lacking NaV1.2 channels and found that they were intrinsically hyperexcitable, firing high-frequency bursts of action potentials (APs) despite decrements in AP size and speed. Compartmental modeling and dynamic clamp recordings revealed that NaV1.2 loss prevented potassium channels from properly repolarizing neurons between APs, increasing overall excitability by allowing neurons to reach threshold for subsequent APs more rapidly. This cell-intrinsic mechanism may therefore account for why SCN2A loss-of-function can paradoxically promote seizure.

neuroscience↗