Jaunin P, Jaisser F, Beggah A T, Takeyasu K, Mangeat P, Rossier B C, Horisberger J D, Geering K
Institute of Pharmacology and Toxicology, University of Lausanne, Switzerland.
J Cell Biol. 1993 Dec;123(6 Pt 2):1751-9. doi: 10.1083/jcb.123.6.1751.
The ubiquitous Na,K- and the gastric H,K-pumps are heterodimeric plasma membrane proteins composed of an alpha and a beta subunit. The H,K-ATPase beta subunit (beta HK) can partially act as a surrogate for the Na,K-ATPase beta subunit (beta NK) in the formation of functional Na,K-pumps (Horisberger et al., 1991. J. Biol. Chem. 257:10338-10343). We have examined the role of the transmembrane and/or the ectodomain of beta NK in (a) its ER retention in the absence of concomitant synthesis of Na,K-ATPase alpha subunits (alpha NK) and (b) the functional expression of Na,K-pumps at the cell surface and their activation by external K+. We have constructed chimeric proteins between Xenopus beta NK and rabbit beta HK by exchanging their NH2-terminal plus transmembrane domain with their COOH-terminal ectodomain (beta NK/HK, beta HK/NK). We have expressed these constructs with or without coexpression of alpha NK in the Xenopus oocyte. In the absence of alpha NK, Xenopus beta NK and all chimera that contained the ectodomain of beta NK were retained in the ER while beta HK and all chimera with the ectodomain of beta HK could leave the ER suggesting that ER retention of unassembled Xenopus beta NK is mediated by a retention signal in the ectodomain. When coexpressed with alpha NK, only beta NK and beta NK/HK chimera assembled efficiently with alpha NK leading to similar high expression of functional Na,K-pumps at the cell surface that exhibited, however, a different apparent K+ affinity. beta HK or chimera with the transmembrane domain of beta HK assembled less efficiently with alpha NK leading to lower expression of functional Na,K-pumps with a different apparent K+ affinity. The data indicate that the transmembrane domain of beta NK is important for efficient assembly with alpha NK and that both the transmembrane and the ectodomain of beta subunits play a role in modulating the transport activity of Na,K-pumps.
普遍存在的钠钾泵和胃氢钾泵是由一个α亚基和一个β亚基组成的异源二聚体质膜蛋白。氢钾ATP酶β亚基(βHK)在功能性钠钾泵的形成过程中可部分替代钠钾ATP酶β亚基(βNK)(Horisberger等人,1991年。《生物化学杂志》257:10338 - 10343)。我们研究了βNK的跨膜结构域和/或胞外结构域在以下方面的作用:(a)在没有同时合成钠钾ATP酶α亚基(αNK)的情况下其在内质网中的滞留,以及(b)钠钾泵在细胞表面的功能性表达及其被细胞外钾离子激活的情况。我们通过交换非洲爪蟾βNK和兔βHK的氨基末端加跨膜结构域与它们的羧基末端胞外结构域,构建了嵌合蛋白(βNK/HK,βHK/NK)。我们在非洲爪蟾卵母细胞中表达了这些构建体,有或没有与αNK共表达。在没有αNK的情况下,非洲爪蟾βNK以及所有包含βNK胞外结构域的嵌合体都滞留在内质网中,而βHK以及所有具有βHK胞外结构域的嵌合体能够离开内质网,这表明未组装的非洲爪蟾βNK在内质网中的滞留是由胞外结构域中的滞留信号介导的。当与αNK共表达时,只有βNK和βNK/HK嵌合体能与αNK有效组装,导致在细胞表面功能性钠钾泵有相似的高表达,然而它们表现出不同的表观钾离子亲和力。βHK或具有βHK跨膜结构域的嵌合体与αNK的组装效率较低,导致功能性钠钾泵的表达较低且具有不同的表观钾离子亲和力。数据表明,βNK的跨膜结构域对于与αNK的有效组装很重要,并且β亚基的跨膜结构域和胞外结构域在调节钠钾泵的转运活性中都起作用。