A bipartite signal regulates the faithful delivery of apical domain marker podocalyxin/Gp135

• Published in: Molecular biology of the cell Vol. 18; no. 5; pp. 1710 - 1722
• Main Authors: Yu, Chun-Ying, Chen, Jen-Yau, Lin, Yu-Yu, Shen, Kuo-Fang, Lin, Wei-Ling, Chien, Chung-Liang, ter Beest, Martin B A, Jou, Tzuu-Shuh
• Format: Journal Article
• Language: English
• Published: United States The American Society for Cell Biology 01.05.2007
• Subjects: Amino Acid Motifs; Animals; Base Sequence; Binding Sites; Carrier Proteins - antagonists & inhibitors; Carrier Proteins - genetics; Cell Polarity; Cells, Cultured; Cytoskeletal Proteins - metabolism; Dogs; Endocytosis; Glycosylation; Membrane Microdomains - metabolism; Models, Biological; Mutation; Protein Kinase C - metabolism; Protein Sorting Signals - genetics; Protein Structure, Tertiary; RNA, Small Interfering - genetics; Sialoglycoproteins - chemistry; Sialoglycoproteins - genetics; Sialoglycoproteins - metabolism; Signal Transduction
• ISSN: 1059-1524; 1939-4586
• DOI: 10.1091/mbc.e06-07-0629

Podocalyxin/Gp135 was recently demonstrated to participate in the formation of a preapical complex to set up initial polarity in MDCK cells, a function presumably depending on the apical targeting of Gp135. We show that correct apical sorting of Gp135 depends on a bipartite signal composed of an extracellular O-glycosylation-rich region and the intracellular PDZ domain-binding motif. The function of this PDZ-binding motif could be substituted with a fusion construct of Gp135 with Ezrin-binding phosphoprotein 50 (EBP50). In accordance with this observation, EBP50 binds to newly synthesized Gp135 at the Golgi apparatus and facilitates oligomerization and sorting of Gp135 into a clustering complex. A defective connection between Gp135 and EBP50 or EBP50 knockdown results in a delayed exit from the detergent-resistant microdomain, failure of oligomerization, and basolateral missorting of Gp135. Furthermore, the basolaterally missorted EBP50-binding defective mutant of Gp135 was rapidly retrieved via a PKC-dependent mechanism. According to these findings, we propose a model by which a highly negative charged transmembrane protein could be packed into an apical sorting platform with the aid of its cytoplasmic partner EBP50.