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首页> 外文期刊>Cytokine >Intracellular localization of the p35 subunit of murine IL-12.
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Intracellular localization of the p35 subunit of murine IL-12.

机译：小鼠IL-12 p35亚基的细胞内定位。

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摘要

Production of interleukin-12 (IL-12), a heterodimer of p35 and p40 subunits, is limited by p35 expression. A long and a short murine p35 mRNA potentially encoding proteins differing in pre-sequence size are produced. Increased pre-sequence size could convert a cleaved signal peptide to an uncleaved signal peptide, raising the possibility that a membrane-bound form of p35 is produced. The intracellular localization of the p35 encoded by each mRNA isoform was determined by constructing cDNAs containing the long or short p35 cDNA isoform fused in-frame to a cDNA encoding green fluorescent protein (GFP). After transfection of a CV-1 African green monkey kidney cell line with the constructs, confocal microscopy and immunoblotting of extracted microsomal membranes demonstrated that the p35-GFP fusion protein encoded by the long or short mRNA accumulates in the Golgi apparatus as an endoglycosidase H-sensitive glycosylated integral membrane protein. In contrast, a p40-GFP fusion protein accumulates in the Golgi apparatus as a soluble protein. Since assembly of the p35 and p40 subunits to form bioactive IL-12 occurs in the ER, release of membrane-tethered IL-12 by proteolytic cleavage in a late Golgi or post-Golgi compartment may represent an as yet unidentified level at which bioactive IL-12 secretion is regulated.
机译：p35和p40亚基的异二聚体白细胞介素12（IL-12）的生产受到p35表达的限制。产生了长和短的鼠p35 mRNA，它们可能编码前序列大小不同的蛋白质。增大的前序列大小可将切割的信号肽转化为未切割的信号肽，从而增加了产生膜结合形式的p35的可能性。通过构建包含与编码绿色荧光蛋白（GFP）的cDNA符合读框的长或短p35 cDNA同工型的cDNA，可以确定每个mRNA同工型编码的p35的胞内定位。用该构建体转染CV-1非洲绿猴肾细胞系后，共聚焦显微镜检查和提取的微粒体膜的免疫印迹表明，长或短mRNA编码的p35-GFP融合蛋白在高尔基体中积累为内切糖苷酶H-。敏感的糖基化整合膜蛋白。相反，p40-GFP融合蛋白作为可溶蛋白在高尔基体中积累。由于ER中发生p35和p40亚基的组装以形成具有生物活性的IL-12，因此在高尔基体晚期或高尔基体后部通过蛋白水解裂解释放膜束缚的IL-12可能代表了目前尚未确定的生物活性IL的水平。 -12分泌受到调节。

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