Identification of a novel pathway for the in vivo regulation of fast anterograde axonal transport

Abstract

HnRNP A2 is an RNA trafficking protein that binds to a specific cis-acting RNA trafficking element (A2RE) in myelin basic protein RNA and other transported RNAs. A2RE/hnRNPA2 determinants mediate several different steps in RNA trafficking including granule assembly, transport to the plus ends of microtubules and translational activation. A yeast two hybrid screen designed to identify proteins that interact with hnRNP A2 selected a clone corresponding to the carboxyl terminal portion of TOG (tumor overexpressed gene), a microtubule-associated protein that regulates microtubule dynamics. Co-immunostaining of oligodendrocytes with antibody to hnRNPA2 and TOG revealed extensive colocalization of TOG with hnRNP A2 granules in the dendrites. A small population of hnRNP A2 granules lacked TOG and some regions of TOG staining lacked hnRNP A2. In oligodendrocytes injected with fluorescent A2RE RNA and stained for TOG, granules containing fluorescent RNA colocalized with TOG. Co-injection of anti-TOG antibody with fluorescent A2RE RNA decreased colocalization with TOG and increased transport of the injected RNA. These observations suggest that molecular interaction between hnRNP A2 and TOG serves to anchor A2RE mRNAs/hnRNPA2 granules at plus ends of microtubules. Acknowledgements: Supported by NIH NS19943 (EB) and NS15190 (JHC), and NMSS RG2843 (EB). CP05-02Expression of retrograde transport proteins during oligodendrocyte R. M. Gould, T. Bukher and E. Barbarese N.Y.S. Institute for Basic Research, 1050 Forest Hill Road, Staten Island, NY, USA; University of Connecticut Health Center, Farmington, CT, USA Using a combination of subcellular fractionation and suppression subtractive hybridization, we found that a high molecular weight dynein light intermediate chain-2 mRNA copurifies with myelin basic protein mRNA in rat brain myelin. We used a combination of subtraction efficiency and Northern blot analysis to show that other dynein (dynein heavy chain, dynein intermediate chains 1 and 2 and dynein light intermediate chain-1) and dynactin (arp-1, p150 glued , dynamitin, CLIP-170 and capB1) mRNAs do not copurify with these mRNAs in rat brain myelin. We used immunofluorescence and Western blotting to show that all of these proteins are expressed by cultured oligodendrocytes. These results suggest that of all the dynein/dynactin proteins synthesized in oligodendrocytes, only one, dynein light intermediate chain-2, is synthesized in the cell's processes (near sties of myelin sheath assembly). Consequently, the addition of dynein light intermediate chain-2 molecules to other dynein/dynactin proteins at sites of myelin sheath assembly may influence retrograde transport initiated at these distal sites. Acknowledgements: Supported by grant RG2944A5/1 from the National Multiple Sclerosis Society. CP05-03Differences in the retrograde axonal transport of nerve growth factor and dopamine beta-hydroxylase I. A. Hendry, M. W. Weible II and N. Ozsarac Division of Neuroscience, JCSMR, ANU, Canberra, ACT, AustraliaThere are two potential po...