Mammalian ELAV-like neuronal RNA-binding proteins HuB and HuC promote neuronal development in both the central and the peripheral nervous systems

Hu proteins are mammalian embryonic lethal abnormal visual system (ELAV)-li ke neuronal RNA-binding proteins that contain three RNA recognition motifs, Although Drosophila ELAV is required for the correct differentiation and s urvival of neurons, the roles played by the Hu genes in the mammalian nervo us system remain largely unknown, To explore the in vivo functions of mouse Hu proteins, we overexpressed them in rat pheochromocytoma PC12 cells, whe re they induced neuronal phenotype in the absence of nerve growth factor, W e have characterized the functions of various forms of mHuB and mHuC bearin g point mutations or deletions. Mutants of mHuC that had amino acid exchang es in the RNP1 domain of the first or second RNA recognition motifs (RRMs) lost biologic activity as well as RNA-binding activity. In addition, the mu tants containing only the third RRM failed to induce the neuronal phenotype in PC12 cells and inhibited the biologic activity of cotransfected wild-ty pe mHuB and mHuC, thus acting as a dominant-negative form. However, these m utants could not suppress the nerve growth factor-induced differentiation o f PC12 cells. Further, we misexpressed wild-type and dominant-negative Hu i n E9.5 mouse embryos, by using electroporation into the neural tube at the level of the rhombencephalon. mHuB and mHuC induced the ectopic expression of neuronal markers, whereas the dominant-negative forms of mHuB and mHuC s uppressed the differentiation of central nervous system motor neurons, From these results, we suggest that Hu proteins are required for neuronal diffe rentiation in the mammalian nervous system.