Heterotrimeric and small molecular mass GTP-binding proteins of rat brain neuronal and glial nuclei

Heterotrimeric and small molecular mass guanine nucleotide binding (GTP-bin ding) proteins were found in neuronal and glial nuclei isolated from rat br ain. Neuronal nuclei bound 0.213 +/- 0.055 pmoles of GTP/mu g protein (n = 8) and glial nuclei bound 0.145 +/- 0.038 pmoles of GTP/mu g protein (n = 8 ), The intrinsic GTPase activity of neuronal and glial nuclei was 0.0019 +/ - 0.0005 pmoles GTP hydrolyzed/min/mu g protein (n = 10) and 0.0022 +/- 0.0 006 pmoles GTP hydrolyzed/min/mu g protein (n 10), respectively Western blo t analysis was carried out using a peptide-specific antibody that recognize s a common sequence in the ar-subunit of the various heterotrimeric G-prote ins, The antibody revealed the presence of a polypeptide of molecular mass of 40 kDa only in neuronal nuclei. Small molecular mass GTP-binding protein s were detected by incubating nitrocellulose blots with [alpha-P-32]GTP, Th e results demonstrated the presence of 25-26 kDa GTP-binding proteins in bo th populations of nuclei. However, the binding of [alpha-P-32]GTP to neuron al nuclei was approximately 3-fold greater than to the glial nuclei, Furthe r analysis by two-dimensional polyacrylamide gel electrophoresis resolved t he neuronal nuclei 26 kDa protein into three forms (a-c) with the most acid ic form (c) being the major species. The neuronal 25 kDa protein was resolv ed into two forms that were present in approximately equal concentration. I n glial nuclei, only the 26 kDa (c) and a small amount of the 25 kDa protei ns were detected. However, both populations of nuclei contained the small m olecular mass GTP-binding protein, ran, Differential association of non-ran small molecular mass GTP-binding proteins and heterotrimeric G-proteins wi th neuronal nuclei suggests a potential role for these guanine nucleotide b inding proteins in the function of this cell type. (C) 1999 Wiley-Liss, Inc .