Braga Tiago, Ringvall Maria, Tveit Heidi, Abrink Magnus, Pejler Gunnar
Swedish University of Agricultural Sciences, Department of Anatomy, Physiology and Biochemistry, Uppsala, Sweden.
Mol Immunol. 2009 Jan;46(3):422-8. doi: 10.1016/j.molimm.2008.10.019. Epub 2008 Dec 6.
Mast cell granule maturation and storage of granule components has previously been shown to be critically dependent on serglycin (SG), a proteoglycan abundantly stored in mast cell secretory granules. The N-terminal portion of serglycin contains a conserved disulfide motif that is similar to motifs found in secretory granule compounds of neuroendocrine cells. Interference with such motifs of neuroendocrine cells with dithiothreitol (DTT) has previously been shown to cause cellular missorting. To investigate the implication for serglycin, serglycin(+/+) and serglycin(-/-) bone marrow derived mast cells (BMMCs) were treated with DTT followed by assessment of proteoglycan synthesis and secretory granule integrity. Treatment of serglycin(+/+) BMMCs with DTT almost completely abolished biosynthetic incorporation of (35)S-sulfate into proteoglycans, caused a dramatic reduction of granular staining with May Grünwald/Giemsa as well as disruption of granule dense core formation as shown by transmission electron microscopy. In addition, the storage of carboxypeptidase A, a major secretory granule compound, was markedly reduced following DTT treatment. In contrast, none of these effects were seen after treatment of SG(-/-) BMMCs with DTT, indicating that they were serglycin-specific. Notably, DTT treated serglycin(+/+) BMMCs showed similar morphology as did the serglycin(-/-) BMMCs. DTT treatment affected neither the viability of the BMMCs nor the mRNA levels for serglycin or carboxypeptidase A. Together, these data indicate that DTT causes dramatic, serglycin-specific effects on mast cell granule. These findings are thus in accordance with a role for the N-terminal disulfide motif in serglycin for regulation of mast cell secretory granule integrity.
肥大细胞颗粒成熟和颗粒成分的储存先前已被证明严重依赖于丝甘蛋白聚糖（SG），这是一种大量储存在肥大细胞分泌颗粒中的蛋白聚糖。丝甘蛋白聚糖的N端部分含有一个保守的二硫基序，类似于在神经内分泌细胞分泌颗粒化合物中发现的基序。先前已证明用二硫苏糖醇（DTT）干扰神经内分泌细胞的此类基序会导致细胞分选错误。为了研究对丝甘蛋白聚糖的影响，用DTT处理丝甘蛋白聚糖（+/+）和丝甘蛋白聚糖（-/-）骨髓来源的肥大细胞（BMMCs），然后评估蛋白聚糖合成和分泌颗粒完整性。用DTT处理丝甘蛋白聚糖（+/+）BMMCs几乎完全消除了（35）S-硫酸盐向蛋白聚糖中的生物合成掺入，导致May Grünwald/Giemsa染色的颗粒显著减少，以及透射电子显微镜显示的颗粒致密核心形成的破坏。此外，DTT处理后，主要分泌颗粒化合物羧肽酶A的储存明显减少。相比之下，用DTT处理SG（-/-）BMMCs后未观察到这些影响，表明它们是丝甘蛋白聚糖特异性的。值得注意的是，DTT处理的丝甘蛋白聚糖（+/+）BMMCs表现出与丝甘蛋白聚糖（-/-）BMMCs相似的形态。DTT处理既不影响BMMCs的活力，也不影响丝甘蛋白聚糖或羧肽酶A的mRNA水平。总之，这些数据表明DTT对肥大细胞颗粒产生显著的、丝甘蛋白聚糖特异性的影响。因此，这些发现与丝甘蛋白聚糖中N端二硫基序在调节肥大细胞分泌颗粒完整性中的作用一致。
