Biology subjects

Myers, C. G.

Publications and source records attributed to Myers, C. G..

4 recordsLinked to original sources

bioRxiv2024

Small molecule modulation of insulin receptor-insulin like growth factor-1 receptor heterodimers in human endothelial cells

ObjectivesThe insulin receptor (IR) and insulin like growth factor-1 receptor (IGF-1R) are heterodimers consisting of two extracellular -subunits and two transmembrane {beta} -subunits. Insulin {beta} and insulin like growth factor-1 {beta} hemi-receptors can heterodimerize to form hybrids composed of one IR {beta} and one IGF-1R {beta}. The function of hybrids in the endothelium is unclear. We sought insight by developing a small molecule capable of reducing hybrid formation in endothelial cells. MethodsWe performed a high-throughput small molecule screening, based on a homology model of hybrid structure. Endothelial cells were studied using western blotting and qPCR to determine the effects of small molecules that reduced hybrid formation. ResultsOur studies unveil a first-in-class quinoline-containing heterocyclic small molecule that reduces hybrids by >50% in human umbilical vein endothelial cells (HUVECs) with no effects on insulin or insulin like growth factor-1 receptors. This small molecule reduced expression of the negative regulatory p85 subunit of phosphatidylinositol 3-kinase, increased basal phosphorylation of the downstream target Akt and enhanced insulin/insulin-like growth factor-1 and shear stress-induced serine phosphorylation of Akt. In primary saphenous vein endothelial cells (SVEC) from patients with type 2 diabetes mellitus undergoing coronary artery bypass (CABG) surgery, hybrid receptor expression was greater than in patients without type 2 diabetes mellitus. The small molecule significantly reduced hybrid expression in SVEC from patients with type 2 diabetes mellitus. ConclusionsWe identified a small molecule that decreases the formation of IR: IGF-1R hybrid receptors in human endothelial cells, without significant impact on the overall expression of IR or IGF-1R. In HUVECs, reduction of IR: IGF-1R hybrid receptors leads to an increase in insulin-induced serine phosphorylation of the critical downstream signalling kinase, Akt. The underpinning mechanism appears, at least in part to involve the attenuation of the adverse effect of IR: IGF-1R hybrid receptors on PI3-kinase signalling. HighlightsO_LIWe have discovered a small molecule (HI) that inhibits insulin receptor/IGF-1 receptor hybrid formation. C_LIO_LIHI reveals previously unrecognised actions of insulin receptor/IGF-1 receptor hybrids distinct to insulin and IGF-1 receptors in endothelial cells. C_LIO_LITreatment of endothelial cells with HI enhances activity of the downstream signalling kinase Akt due to inhibitory regulation via PI3-K. C_LI

cell biology↗

bioRxiv2021

A small molecule reveals role of insulin receptor-insulin like growth factor-1 receptor heterodimers

The insulin receptor and insulin like growth factor-1 receptor are heterodimers consisting of two extracellular -subunits and two transmembrane {beta}-subunits. IR {beta} and IGF1R {beta} hemi-receptors can heterodimerize to form hybrids composed of one IR {beta} and one IGF1R {beta}. Widely distributed in mammalian tissues, in contrast to IR and IGF1R the physiological function of hybrids is unclear. To identify tool compounds that inhibit hybrid formation we performed a high-throughput small molecule screen based on a homology model of hybrid structure. Our studies unveil a first in class quinoline-containing heterocyclic small molecule that reduces hybrids by >50% in human umbilical vein endothelial cells with no effect on IR or IGF1R expression. Downstream of IR and IGF1R our small molecule led to reduced expression of the negative regulatory p85 subunit of phosphatidylinositol 3-kinase, an increase in phosphorylation of its downstream target Akt and enhanced insulin and shear-induced phosphorylation of Akt. We show that hybrids have a role in human endothelial cell physiology distinct from IR and IGF1R.

cell biology↗

bioRxiv2019

VE-cadherin endocytosis controls vascular integrity and patterning during development

Tissue morphogenesis requires dynamic intercellular contacts that are subsequently stabilized as tissues mature. The mechanisms governing these competing adhesive properties are not fully understood. Using gain- and loss-of-function approaches, we tested the role of p120-catenin (p120) and VE-cadherin (VE-cad) endocytosis in vascular development using mouse mutants that exhibit increased (VE-cadGGG/GGG) or decreased (VE-cadDEE/DEE) internalization. VE-cadGGG/GGG mutant mice exhibited reduced VE-cad-p120 binding, reduced VE-cad levels, microvascular hemorrhaging, and decreased survival. By contrast, VE-cadDEE/DEE mutants exhibited normal vascular permeability but displayed microvascular patterning defects. Interestingly, VE-cadDEE/DEE mutant mice did not require endothelial p120, demonstrating that p120 is dispensable in the context of a stabilized cadherin. In vitro, VE-cadDEE mutant cells displayed defects in polarization and cell migration that were rescued by uncoupling VE-cadDEE from actin. These results indicate that cadherin endocytosis coordinates cell polarity and migration cues through actin remodeling. Collectively, our results indicate that regulated cadherin endocytosis is essential for both dynamic cell movements and establishment of stable tissue architecture.\n\nSummary StatementThis study uses mouse genetic and in vitro approaches to demonstrate that cadherin endocytosis is critical for the formation of blood vessels during development by promoting actin-dependent collective cell migration, whereas the inhibition of this endocytosis by p120 binding is essential for vessel stabilization.

cell biology↗

bioRxiv2019

Mesoscale Modeling of a Nucleosome-Binding Antibody (PL2-6): Mono- vs. Bivalent Chromatin Complexes

Visualizing chromatin adjacent to the nuclear envelope (denoted \"epichromatin\") by in vitro immunostaining with a bivalent nucleosome-binding antibody (termed monoclonal antibody PL2-6) has suggested a distinct and conserved chromatin structure. Moreover, different staining patterns for chromatin complexed with the monovalent \"Fab\" fragment of PL2-6, compared to the bivalent form, point to distinct binding interactions. To help interpret antibody/chromatin interactions and these differential binding modes, we incorporate coarse-grained PL2-6 antibody modeling into our mesoscale chromatin model and analyze interactions and fiber structures for the antibody/chromatin complexes in open and condensed chromatin, with and without linker histone H1 (LH). Despite minimal and transient interactions at physiological salt, we capture differential binding for monomer and dimer antibody forms to open fibers, with much more intense interactions in the bivalent antibody/chromatin complex. For these open \"zigzag\" fiber morphologies, differences result from antibody competition for peptide tail contacts with internal chromatin fiber components (nucleosome core and linker DNA). Antibody competition results in dramatic conformational and energetic differences among monovalent, bivalent, and free chromatin systems in the parental linker DNA / tail interactions. These differences in binding modes and changes in internal fiber structure, driven by conformational entropy gains, help interpret the differential staining patterns for the monovalent versus bivalent antibody/chromatin complexes. More generally, such dynamic interactions which depend on the complex internal structure and self-interactions of the chromatin fiber have broader implications to other systems that bind to chromatin, such as linker histones and remodeling proteins.\n\nSTATEMENT OF SIGNIFICANCEUsing mesoscale modeling, we help interpret differential binding modes for antibody/chromatin interactions to elucidate the structural details of \"epichromatin\" (chromatin adjacent to the nuclear envelope), which had been visualized to produce different staining patterns for monovalent and bivalent forms of the PL2-6 antibody. To our knowledge, this is the first application of such a coarse-grained computational antibody model to probe chromatin structure and mechanisms of antibody/chromatin binding. Our work emphasizes how antibody units compete with native internal chromatin fiber units (histone tails, nucleosome core, and linker DNA) for fiber-stabilizing interactions and thereby drive differential antibody binding for open zigzag chromatin fibers. Such competition, which dynamically alters internal chromatin structure upon binding, could be relevant to other chromatin binding mechanisms such as those involving linker histones or chromatin remodeling proteins.

biophysics↗