Journal
MOLECULAR AND CELLULAR NEUROSCIENCE
Volume 20, Issue 4, Pages 563-578Publisher
ACADEMIC PRESS INC ELSEVIER SCIENCE
DOI: 10.1006/mcne.2002.1146
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The MEGF1 (protein I with multiple EGF-like domains) gene, which was identified using motif-trap screening, encodes an extraordinarily large protein containing two EGF-like and 34 cadherin motifs. In situ hybridization analysis revealed that the MEGF1 gene was specifically expressed in granule cells of the cerebellum. Interestingly, in the developing cerebellum, granule cells in the inner external germinal layer and migrating granule cells expressed MEGF1 mRNA whereas proliferating cells in the outer external germinal layer did not express MEGF1 mRNA Expression levels in the internal granule cell layer peaked during the third postnatal week and remained considerably high in the adult cerebellum. MEGF1 protein was detected in only the cerebellum as a single 480-kDa band by immunoblot analyses using polyclonal antibodies against either the N-terminal or the C-terminal region of MEGF1 protein. Using light and electron microscopic immunocytochemistry, specific immunostaining of the MEGF1 protein was observed in the molecular layer of the cerebellum, suggesting that MEGF1 protein was localized in the parallel fibers of cerebellar granule cells. This was corroborated by results from experiments using primary dispersed cultures of cerebellar granule cells and cerebellar microexplant cultures. The homophilic interaction of MEGF1 proteins was confirmed with both a cell aggregation assay and an in vitro copurification assay. Based on these results, a novel function of the enormous protocadherins in cerebellar development, namely, the modulation of the extracellular space surrounding parallel fibers during development, was proposed.
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