Phosphatidylinositol 4,5-bisphosphate (PIP2) stimulates the electrogenic Na/HCO3 cotransporter NBCe1-A expressed in Xenopus oocytes.
Phosphatidylinositol 4,5-bisphosphate (PIP2) stimulates the electrogenic Na/HCO3 cotransporter NBCe1-A expressed in Xenopus oocytes.
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磷脂酰肌醇 4,5-二磷酸 (PIP2) 刺激非洲爪蟾卵母细胞中表达的产电 Na/HCO3 协同转运蛋白 NBCe1-A。
DOI：
10.1073/pnas.0906303106
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发表时间：
2009
影响因子：
11.1
通讯作者：
Bevensee,MarkO
中科院分区：
文献类型：
--
作者：
Wu,Jianping;McNicholas,CarmelM;Bevensee,MarkO
碳酸氢盐转运体受信号分子/离子的调节，如蛋白激酶、三磷酸腺苷和钙离子。虽然磷脂如PIP2可以刺激Na-H交换活性，但对磷脂对碳酸氢盐转运体的调节知之甚少。我们用膜片钳技术研究了非洲爪哇大片细胞异源表达大鼠NBCe1-A的功能和调控。将大片细胞液侧暴露于5%二氧化碳/33 mMHCO3−溶液中，74%附着于吸管(−VP=−60 mV)的大片的平均内向电流为14pA，其中含有低Na+的名义上不含HCO3的−溶液。在没有Na+的情况下，电流减小了80-90%，在存在200μM Did的情况下，电流大约减小了75%，在注入H2O的卵母细胞的大片中没有电流。NbCe1-A电流表现出时间依赖性的衰减，在有或没有钒酸盐和F-−的情况下，通过去除镁离子来抑制总的磷酸酶活性。用5或10μ的MPIP2(DiC8)与HCO3-−混合使用，可在54%表达NbC但不注射H2O的大片卵母细胞上产生内向电流。PIP2诱导的电流依赖于HCO3−，且随着NBCe1-A浓度的增加而增大，在无Na+时减小62%，在存在200μM Did时减小90%。聚阳离子新霉素(2 5 0~5 0 0μM)使PIP2诱发的内向电流降低69%，精胺(10 0μM)使电流降低97%。精胺、多聚D-赖氨酸和新霉素均可使基线HCO3−诱导的内向电流减少高达85%。综上所述，PIP2刺激NBCe1-A活性，而肌醇磷脂是碳酸氢盐转运蛋白的调节者。
Bicarbonate transporters are regulated by signaling molecules/ions such as protein kinases, ATP, and Ca2+. While phospholipids such as PIP2can stimulate Na-H exchanger activity, little is known about phospholipid regulation of bicarbonate transporters. We used the patch-clamp technique to study the function and regulation of heterologously expressed rat NBCe1-A in excised macropatches fromXenopus laevisoocytes. Exposing the cytosolic side of inside-out macropatches to a 5% CO2/33 mM HCO3−solution elicited a mean inward current of 14 pA in 74% of macropatches attached to pipettes (−Vp= −60 mV) containing a low-Na+, nominally HCO3−-free solution. The current was 80–90% smaller in the absence of Na+, approximately 75% smaller in the presence of 200 μM DIDS, and absent in macropatches from H2O-injected oocytes. NBCe1-A currents exhibited time-dependent rundown that was inhibited by removing Mg2+in the presence or absence of vanadate and F−to reduce general phosphatase activity. Applying 5 or 10 μM PIP2(diC8) in the presence of HCO3−induced an inward current in 54% of macropatches from NBC-expressing, but not H2O-injected oocytes. PIP2-induced currents were HCO3−-dependent and somewhat larger following more NBCe1-A rundown, 62% smaller in the absence of Na+, and 90% smaller in the presence of 200 μM DIDS. The polycation neomycin (250–500 μM) reduced the PIP2-induced inward current by 69%; spermine (100 μM) reduced the current by 97%. Spermine, poly-D-lysine, and neomycin all reduced the baseline HCO3−-induced inward currents by as much as 85%. In summary, PIP2stimulates NBCe1-A activity, and phosphoinositides are regulators of bicarbonate transporters.