GPI-anchored carbonic anhydrase IV displays both intra- and extracellular activity in cRNA-injected oocytes and in mouse neurons.
GPI-anchored carbonic anhydrase IV displays both intra- and extracellular activity in cRNA-injected oocytes and in mouse neurons.
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GPI 锚定碳酸酐酶 IV 在注射 cRNA 的卵母细胞和小鼠神经元中显示出细胞内和细胞外活性。
DOI：
10.1073/pnas.1221213110
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发表时间：
2013
影响因子：
11.1
通讯作者：
Deitmer,JoachimW
中科院分区：
文献类型：
--
作者：
Schneider,Hans-Peter;Alt,MarcoD;Klier,Michael;Spiess,Alena;Andes,FabianT;Waheed,Abdul;Sly,WilliamS;Becker,HolgerM;Deitmer,JoachimW
可溶性胞质碳酸酐酶（CA）参与哺乳动物细胞质的pH调节。膜结合的CA异构体-例如异构体IV、IX、XII、XIV和XV-也催化二氧化碳可逆转化为质子和碳酸氢盐，但在细胞膜的细胞外表面。当人CA亚型IV异源表达inXenopusoacytes，我们观察到，通过测量H+在细胞膜的外面和在细胞质中的离子选择性微电极，不仅细胞外催化CA活性，但也强大的细胞内活性。CA IV在卵母细胞中的表达通过免疫细胞化学证实，CA IV活性通过质谱测定。CA IV的细胞外和细胞内催化活性可以用苯甲酰胺（CA抑制剂，其相对缓慢的膜渗透性）来分析。在缺乏CA IV的突变小鼠的急性小脑切片中，CO2去除/添加后颗粒细胞的胞质H+移位显著慢于野生型小鼠。我们的研究结果表明，膜相关的CA IV有助于强大的催化活性胞内，这种活动参与调节H+动力学在胞质溶胶中，无论是在注射的卵母细胞和小鼠神经元。
Soluble cytosolic carbonic anhydrases (CAs) are well known to participate in pH regulation of the cytoplasm of mammalian cells. Membrane-bound CA isoforms—such as isoforms IV, IX, XII, XIV, and XV—also catalyze the reversible conversion of carbon dioxide to protons and bicarbonate, but at the extracellular face of the cell membrane. When human CA isoform IV was heterologously expressed inXenopusoocytes, we observed, by measuring H+at the outer face of the cell membrane and in the cytosol with ion-selective microelectrodes, not only extracellular catalytic CA activity but also robust intracellular activity. CA IV expression in oocytes was confirmed by immunocytochemistry, and CA IV activity measured by mass spectrometry. Extra- and intracellular catalytic activity of CA IV could be pharmacologically dissected using benzolamide, the CA inhibitor, which is relatively slowly membrane-permeable. In acute cerebellar slices of mutant mice lacking CA IV, cytosolic H+shifts of granule cells following CO2removal/addition were significantly slower than in wild-type mice. Our results suggest that membrane-associated CA IV contributes robust catalytic activity intracellularly, and that this activity participates in regulating H+dynamics in the cytosol, both in injected oocytes and in mouse neurons.