Neuropharmacology

Biography

Biography: Ranjana Arya

Abstract

Biological basis of pathogenesis of a large number of genetic disorders is not known, particularly for those diseases which affect neuro-muscular system. UDP-GlcNAc 2-epimerase /ManNAc kinase (GNE) is a bifunctional enzyme (N-terminal epimerase and C-terminal Kinase domain) that catalyzes rate limiting step in sialic acid biosynthesis. Homozygous misssense mutations in either epimerase or kinase domain of GNE leads to slowly progressive autosomal recessive genetic neuromuscular disorder, Hereditary Inclusion Body Myopathy (HIBM)/Distal myopathy with rimmed vacuoles (DMRV). These GNE related myopathies are characterized by hyposialylation of glycoproteins in muscle cells of patients and primary defect in either N or O-linked glycosylation. However, it appears from some recent experiments including those from our laboratory that mutant GNE may also affect targets that are not directly related to sialic acid biosynthesis. In particular cytoskeletal network, sarcomere organization and apoptotic signaling are likely to be altered in muscle cells. In absence of clear understanding of the pathomechanism, no treatment is currently available to cure the disease. Our laboratory focuses on deciphering alternate roles of GNE in regulating cell functions with an aim to identify more effective drug targets. We have established a HEK293 cell based assay system where pathologically relevant mutations of GNE are overexpressed alongwith GNE knockdown using shRNA. The system is validated by reduced sialic acid content of the cell and restoration of sialylation after supplementation with 5 mM sialic acid. Using this system, GNE has been shown to affect cell adhesion property via hyposialylation of β-1 integrin. This leads to increased binding of cell to fibronectin and activation of FAK/Src/paxillin to promote focal adhesion assembly. The cytoskeletal network is altered due to misbalance in G-actin and F-actin levels in GNE deficient cell lines. Interestingly, mutation in GNE causes increased apoptosis via mitochondrial dysfunction. The ultrastructure of mitochondria was found to be altered in GNE mutant cells along with disruption of membrane potential. In addition, total reactive oxygen species (ROS) were found to be altered indicative of oxidative stress generation. Our study clearly provides a base for understanding pathomechanism of GNE myopathy and the opportunity of using cell-based assay for exploring pharmacological drug molecules.