Tareilus E, Roux M, Qin N, Olcese R, Zhou J, Stefani E, Birnbaumer L
Department of Anesthesiology, University of California at Los Angeles 90095-1778, USA.
Proc Natl Acad Sci U S A. 1997 Mar 4;94(5):1703-8. doi: 10.1073/pnas.94.5.1703.
Two closely related beta subunit mRNAs (xo28 and xo32) were identified in Xenopus oocytes by molecular cloning. One or both appear to be expressed as active proteins, because: (i) injection of Xenopus beta antisense oligonucleotides, but not of sense or unrelated oligonucleotides, significantly reduced endogenous oocyte voltage-gated Ca2+ channel (VGCC) currents and obliterated VGCC currents that arise after injection of mammalian alpha1 cRNAs (alpha(1C) and alpha(1E)); (ii) coinjection of a Xenopus beta antisense oligonucleotide and excess rat beta cRNA rescued expression of alpha1 Ca2+ channel currents; and (iii) coinjection of mammalian alpha1 cRNA with cRNA encoding either of the two Xenopus beta subunits facilitated both activation and inactivation of Ca2+ channel currents by voltage, as happens with most mammalian beta subunits. The Xenopus beta subunit cDNAs (beta3xo cDNAs) predict proteins of 484 aa that differ in only 22 aa and resemble most closely the sequence of the mammalian type 3 beta subunit. We propose that "alpha1 alone" channels are in fact tightly associated alpha1beta3xo channels, and that effects of exogenous beta subunits are due to formation of higher-order [alpha1beta]beta(n) complexes with an unknown contribution of beta3xo. It is thus possible that functional mammalian VGCCs, rather than having subunit composition alpha1beta, are [alpha1beta]beta(n) complexes that associate with alpha2delta and, as appropriate, other tissue-specific accessory proteins. In support of this hypothesis, we discovered that the last 277-aa of alpha(1E) have a beta subunit binding domain. This beta binding domain is distinct from the previously known interaction domain located between repeats I and II of calcium channel alpha1 subunits.
通过分子克隆在非洲爪蟾卵母细胞中鉴定出两个密切相关的β亚基mRNA（xo28和xo32）。其中一个或两者似乎都表达为活性蛋白，原因如下：(i) 注射非洲爪蟾β反义寡核苷酸而非正义或无关寡核苷酸，可显著降低内源性卵母细胞电压门控Ca2+通道（VGCC）电流，并消除注射哺乳动物α1 cRNAs（α(1C)和α(1E)）后产生的VGCC电流；(ii) 共同注射非洲爪蟾β反义寡核苷酸和过量大鼠β cRNA可挽救α1 Ca2+通道电流的表达；(iii) 将哺乳动物α1 cRNA与编码两种非洲爪蟾β亚基之一的cRNA共同注射，可促进Ca2+通道电流的电压激活和失活，这与大多数哺乳动物β亚基的情况相同。非洲爪蟾β亚基cDNA（β3xo cDNA）预测的蛋白质为484个氨基酸，仅在22个氨基酸上有所不同，与哺乳动物3型β亚基的序列最为相似。我们提出，“仅α1”通道实际上是紧密相关的α1β3xo通道，外源性β亚基的作用是由于形成了高阶[α1β]β(n)复合物，其中β3xo的贡献未知。因此，功能性哺乳动物VGCCs可能不是具有α1β亚基组成，而是与α2δ以及适当的其他组织特异性辅助蛋白相关的[α1β]β(n)复合物。为支持这一假设，我们发现α(1E)的最后277个氨基酸具有一个β亚基结合结构域。这个β结合结构域与先前已知的位于钙通道α1亚基重复序列I和II之间的相互作用结构域不同。
