Roush D L, Gottardi C J, Naim H Y, Roth M G, Caplan M J
Department of Cellular and Molecular Physiology, Yale University School of Medicine, New Haven, Connecticut 06510, USA.
J Biol Chem. 1998 Oct 9;273(41):26862-9. doi: 10.1074/jbc.273.41.26862.
Tyrosine-dependent sequence motifs are implicated in sorting membrane proteins to the basolateral domain of Madin-Darby canine kidney (MDCK) cells. We find that these motifs are interpreted differentially in various polarized epithelial cell types. The H, K-ATPase beta subunit, which contains a tyrosine-based motif in its cytoplasmic tail, was expressed in MDCK and LLC-PK1 cells. This protein was restricted to the basolateral membrane in MDCK cells, but was localized to the apical membrane in LLC-PK1 cells. Similarly, HA-Y543, a construct in which a tyrosine-based motif was introduced into the cytoplasmic tail of influenza hemagglutinin, was sorted to the basolateral membrane of MDCK cells and retained at the apical membrane of LLC-PK1 cells. A chimera in which the cytoplasmic tail of the H,K-ATPase beta subunit protein was replaced with the analogous region of the Na,K-ATPase beta subunit polypeptide was localized to both surface domains of MDCK cells. Mutation of tyrosine-20 of the H,K-ATPase beta subunit cytoplasmic sequence to an alanine was sufficient to disrupt basolateral localization of this polypeptide. In contrast, these constructs all remain localized to the apical membrane in LLC-PK1 cells. The FcRII-B2 protein bears a di-leucine motif and is found at the basolateral membrane of both MDCK and LLC-PK1 cells. These results demonstrate that polarized epithelia are able to discriminate between different classes of specifically defined membrane protein sorting signals.
酪氨酸依赖的序列基序与膜蛋白分选至Madin-Darby犬肾（MDCK）细胞的基底外侧结构域有关。我们发现这些基序在各种极化上皮细胞类型中有着不同的解读。H，K-ATP酶β亚基在其胞质尾部含有一个基于酪氨酸的基序，在MDCK和LLC-PK1细胞中表达。该蛋白在MDCK细胞中局限于基底外侧膜，但在LLC-PK1细胞中定位于顶端膜。同样，HA-Y543（一种将基于酪氨酸的基序引入流感血凝素胞质尾部的构建体）被分选至MDCK细胞的基底外侧膜，并保留在LLC-PK1细胞的顶端膜。一种嵌合体，其中H，K-ATP酶β亚基蛋白的胞质尾部被Na，K-ATP酶β亚基多肽的类似区域取代，定位于MDCK细胞的两个表面结构域。将H，K-ATP酶β亚基胞质序列中的酪氨酸-20突变为丙氨酸足以破坏该多肽的基底外侧定位。相比之下，这些构建体在LLC-PK1细胞中均保留在顶端膜。FcRII-B2蛋白带有双亮氨酸基序，在MDCK和LLC-PK1细胞的基底外侧膜均有发现。这些结果表明，极化上皮细胞能够区分不同类别的特定定义的膜蛋白分选信号。
