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PMID: 11408592 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't

An intact dilysine-like motif in the carboxyl terminus of MAL is required for normal apical transport of the influenza virus hemagglutinin cargo protein in epithelial Madin-Darby canine kidney cells.

Molecular biology of the cell ·Vol. 12 ·No. 6 ·2001-06-00 ·Pages 1869-83

Puertollano R, Martínez-Menárguez JA, Batista A, Ballesta J, Alonso MA

Abstract

The MAL proteolipid, a component of the integral protein sorting machinery, has been demonstrated as being necessary for normal apical transport of the influenza virus hemagglutinin (HA) and the overall apical membrane proteins in Madin-Darby canine kidney (MDCK) cells. The MAL carboxy terminus ends with the sequence Arg-Trp-Lys-Ser-Ser (RWKSS), which resembles dilysine-based motifs involved in protein sorting. To investigate whether the RWKSS pentapeptide plays a role in modulating the distribution of MAL and/or its function in apical transport, we have expressed MAL proteins with distinct carboxy terminus in MDCK cells whose apical transport was impaired by depletion of endogenous MAL. Apical transport of HA was restored to normal levels by expression of MAL with an intact but not with modified carboxyl terminal sequences bearing mutations that impair the functioning of dilysine-based sorting signals, although all the MAL proteins analyzed incorporated efficiently into lipid rafts. Ultrastructural analysis indicated that compared with MAL bearing an intact RWKSS sequence, a mutant with lysine -3 substituted by serine showed a twofold increased presence in clathrin-coated cytoplasmic structures and a reduced expression on the plasma membrane. These results indicate that the carboxyl-terminal RWKSS sequence modulates the distribution of MAL in clathrin-coated elements and is necessary for HA transport to the apical surface.

MeSH Terms
Adaptor Protein Complex gamma Subunits Amino Acid Motifs Animals Antibodies, Monoclonal/metabolism Biological Transport Biotinylation Caveolin 1 Caveolins/metabolism Cell Line Cell Membrane/metabolism DNA/metabolism Detergents/pharmacology Dipeptides/chemistry Dogs Electrophoresis, Polyacrylamide Gel Endocytosis Endosomes/metabolism Epithelial Cells/metabolism Hemagglutinin Glycoproteins, Influenza Virus/metabolism Immunoblotting Kidney/metabolism Membrane Proteins/metabolism Membrane Transport Proteins Microscopy, Confocal Microscopy, Immunoelectron Mutation Myelin Proteins Myelin and Lymphocyte-Associated Proteolipid Proteins Protein Structure, Tertiary Proteolipids/chemistry Transfection Transferrin/metabolism
Chemicals
Adaptor Protein Complex gamma Subunits Antibodies, Monoclonal Caveolin 1 Caveolins Detergents Dipeptides Hemagglutinin Glycoproteins, Influenza Virus Membrane Proteins Membrane Transport Proteins Myelin Proteins Myelin and Lymphocyte-Associated Proteolipid Proteins Proteolipids Transferrin DNA lysyllysine
Authors & Affiliations
5 authors, click to expand affiliations / ORCID
Puertollano R
Centro de Biología Molecular "Severo Ochoa," Universidad Autónoma de Madrid, Spain.
Martínez-Menárguez J A
Batista A
Ballesta J
Alonso M A
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Article Info
Journal
Molecular biology of the cell
Abbr.
Mol Biol Cell
ISSN
1059-1524
DOI
10.1091/mbc.12.6.1869
Published
2001-06-00
Pages
1869-83
Language
English
Region
United States
NLM ID
9201390
PMCID
PMC37348
Subset
IM
External Links
PubMed source DOI: 10.1091/mbc.12.6.1869 PMC Full Text View journal details
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