Characterization of the Cohen syndrome protein COH1 as a membrane-associated scaffold protein for intracellular transport processes in development and maintenance of neuronal cells

• Applicant: Dr. Wenke Seifert
• Subject Area: Molecular Biology and Physiology of Neurons and Glial Cells
• Term: from 2012 to 2017
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 223309248

Intracellular transport controls a wide spectrum of biological processes such as brain development and maintenance. Core determinants of regular brain function are neuronal differentiation and integration into an efficient network. Studying inherited neurological disorders may help to define molecular mechanisms underlying these processes. The autosomal recessive Cohen syndrome is mainly characterized by non-progressive mental retardation in combination with postnatal microcephaly and is caused by mutations in the COH1 (VPS13B) gene. COH1 encodes a protein of 3997 amino acids (450 kDa) length with partial sequence homology to yeast Vps13p. Recently, we established COH1 as Golgi-associated matrix protein regulating Golgi morphology and Golgi-associated membrane tubulation activity. Our preliminary data show that Coh1 is a positive regulator of neuritogenesis, suggesting that Cohen syndrome is caused by defective terminal neuronal differentiation and integration. To further investigate the molecular mechanisms controlled by COH1, we performed initial co-immunoprecipitations showing that COH1 similar to yeast Vps13p interacts with the Golgi-associated small GTPase RAB6 and the guanine nucleotide exchange factor subunit RIC1. This result implicates a functional network composed of COH1, RAB6 and the heteromeric complex RIC1/RGP1 in mammalian cells and suggests that disturbed Golgi-associated antero- and retrograde intracellular transport processes underlie defective terminal neuronal differentiation and integration in Cohen syndrome.The proposed project aims to identify further functional targets of COH1 to complete our understanding of the hitherto assumed COH1-RIC1/RGP1-RAB6 protein complex using different biochemical approaches, such as yeast-two-hybrid screens, GST-pull down and co-immunoprecipitations. Functionally, we will complement these biochemical analyses by assessment of Golgi-associated transport events and its impact on neurite extension and final synaptic integration, using different genetic and cell biological manipulations to perturb the function of COH1, RAB6, RIC1, and RGP1 and advanced imaging techniques. Finally, we aim to establish a conditional knockout mouse model to corroborate our in vitro studies with in vivo analyses of Coh1. Research within the proposed project is expected to provide profound insights into the molecular machinery that is controlled by COH1 to facilitate Golgi function and intracellular transport. Combined biochemical, genetic and cell biological approaches ensure the integration of our research to formulate a pathomechanistic model of Cohen syndrome. At the same time, our results will significantly contribute to the general comprehension of Golgi-associated transport processes for terminal neuronal differentiation and integration.

• DFG Programme: Research Grants