A key transferase in the generation of mature N-linked carbohydrates is the medial Golgi enzyme N-acetylglucos-aminyltransferase I (EC 18.104.22.168; GlcNAc-TI) which is encoded by the Mgat-1 gene. Previous studies revealed two size classes of Mgat-1 mRNA that are differentially expressed in mouse tissues. Nearly all tissues possess a shorter form (2.9 kb), whereas brain has predominantly a longer Mgat-1 mRNA (33 kb). We now show, by in situ hybridization of horizontal sections of adult mouse brain, that Mgat-1 RNA levels vary markedly in different brain cell types with the greatest expression being in neuronal cells. The differential expression of the 2.9 kb and -33 kb Mgat-1 mRNAs is likely to be controlled by tissue-specific promoters since the size difference between the mRNAs was found to reside entirely in the 5' untranslated region of the 33 kb mRNA. Evidence for this was obtained by an RNase H strategy, reverse transcription-polymerase chain reaction (RT-PCR) and 5' anchored PCR. All of the 0.4 kb difference in size was localized upstream of the previously isolated cDNA sequence. Sequence information from this region was obtained from a mouse brain cDNA library by a PCR amplification strategy and a probe specific for the 33 kb mRNA was generated. This probe hybridized uniquely to the 33 kb Mgat-1 mRNA and Southern blot analysis showed that the new sequence is physically linked to the Mgat-1 gene. A tissue culture model which displays an increase in expression of the 3.3 kb Mgat-1 mRNA transcript during differentiation to neuronal ceDs has been developed in P19 embryonal carcinoma (EC) cells. The combined data suggest that 5' exons in the Mgat-1 gene are differentially utilized by tissue-specific promoters and that transcription factor(s) which specify production of the 33 kb Mgat-1 mRNA are induced by retinoic acid treatment of P19 EC cells.
- Neuronal cell-specific gene expression
- P19 embryonal carcinoma cells
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