Rab44 isoforms similarly promote lysosomal exocytosis, but exhibit differential localization in mast cells.
Rab44 isoforms similarly promote lysosomal exocytosis, but exhibit differential localization in mast cells.
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DOI：
10.1002/2211-5463.13133
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发表时间：
2021-04
期刊：
影响因子：
2.6
通讯作者：
Tsukuba T
中科院分区：
文献类型：
--
作者：
Kadowaki T;Yamaguchi Y;Ogawa K;Tokuhisa M;Okamoto K;Tsukuba T
Rab44是一个大型的Rab GTPase，包含一个Rab GTPase结构域和一些附加的N端结构域。我们最近使用Rab44缺失小鼠来证明Rab44调节肥大细胞的颗粒胞吐和IgE介导的过敏反应。在小鼠肥大细胞中，Rab44以两种亚型表达，即长亚型和短亚型；然而，这两种异构体的特征仍然未知。在这里，我们研究了在大鼠嗜碱性白血病（RBL）‐2H3细胞中表达的人长Rab44亚型和两种小鼠长Rab44亚型及其突变体的分泌和定位。人长异构体和两种小鼠异构体的表达导致β -己糖氨酸酶分泌增加。共聚焦和定量分析表明，人和小鼠的长亚型主要定位于溶酶体，而小鼠的短亚型主要定位于内质网。LysoTracker实时成像显示，各种突变体改变了LysoTracker阳性囊泡的大小和数量。碘霉素处理部分改变了长异构体在质膜和细胞质溶胶上的定位，而对短异构体与溶酶体的共定位影响不大。在机制上，人和小鼠的Rab44蛋白都与v‐SNARE蛋白vesicle - associated membrane protein 8 （VAMP8）相互作用。因此，Rab44亚型同样促进溶酶体胞外分泌，但在肥大细胞中表现出不同的定位。我们研究了人和小鼠Rab44亚型及其突变体在功能和定位上的差异。在大鼠嗜碱性白血病- 2H3细胞中，人类和小鼠长和短亚型的表达增加了β -己糖氨酸酶的分泌。然而，人和小鼠的长亚型主要定位于溶酶体，而小鼠的短亚型主要定位于内质网。
Rab44 is a large Rab GTPase containing a Rab GTPase domain and some additional N‐terminal domains. We recently used Rab44‐deficient mice to demonstrate that Rab44 regulates granule exocytosis in mast cells and IgE‐mediated anaphylaxis. In mouse mast cells, Rab44 is expressed as two isoforms, namely, the long and short forms; however, the characteristics of these two isoforms remain unknown. Here, we investigated secretion and localization of the human long Rab44 isoform and the two mouse isoforms and their mutants expressed in rat basophilic leukemia (RBL)‐2H3 cells. Expression of the human long isoform and both mouse isoforms caused an increase in β‐hexosaminidase secretion. Confocal and quantitative analyses showed that both human and mouse long isoforms localized mainly to lysosomes while the mouse short isoform localized mainly to the ER. Live imaging with LysoTracker indicated that the size and number of LysoTracker‐positive vesicles were altered by the various mutants. Ionomycin treatment partially altered localization of both long isoforms to the plasma membrane and cytosol, whereas it had little effect on colocalization of the short isoform with lysosomes. Mechanistically, both human and mouse Rab44 proteins interacted with vesicle‐associated membrane protein 8 (VAMP8), a v‐SNARE protein. Therefore, Rab44 isoforms similarly promote lysosomal exocytosis, but exhibit differential localization in mast cells. We investigated the difference in function and localization of the human and mouse Rab44 isoforms and their mutants. Expression of the human and mouse long and short isoforms increased β‐hexosaminidase secretion in rat basophilic leukemia‐2H3 cells. However, both human and mouse long isoforms localized mainly to lysosomes while the mouse short isoform localized mainly to the ER.
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