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Loss of epithelial polarity is accompanied by differential association of proteins with intracellular membranes
Author(s) -
Fialka Irene,
Pasquali Christian,
Kurzbauer Robert,
Lottspeich Friedrich,
Huber Lukas A.
Publication year - 1999
Publication title -
electrophoresis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.666
H-Index - 158
eISSN - 1522-2683
pISSN - 0173-0835
DOI - 10.1002/(sici)1522-2683(19990201)20:2<331::aid-elps331>3.0.co;2-a
Subject(s) - epithelial polarity , cell polarity , microbiology and biotechnology , cytosol , transmembrane protein , membrane protein , biology , membrane , polarity (international relations) , intracellular , chemistry , cell , biochemistry , receptor , enzyme
Cellular membranes play an important role in the formation and maintenance of epithelial polarity, which is lost early during carcinogenesis. We set out to identify membrane proteins which are altered during loss of cell polarity in mammary epithelium. As a model system we used murine mammary epithelial cells expressing the conditional oncoprotein c‐JunER, which induces a reversible loss of polarity upon β‐estradiol‐driven activation [1]. When grown either in the absence or presence of hormone, these cells exhibit a polarized or unpolarized phenotype, respectively. Different membrane fractions of polarized or unpolarized cells were analyzed by two‐dimensional electrophoresis (2‐DE) and differentially expressed membrane proteins were identified. To distinguish between transmembrane orientation and peripheral attachment of these proteins, were performed extractions with carbonate at high pH or with Triton X‐114. In addition, cytosolic proteins of both states were analyzed to investigate their differential association with distinct membrane fractions. We found ten protein spots preferentially or exclusively in polarized cells and 17 other proteins as being upregulated during loss of polarity. Some of the peripheral membrane proteins were identified by microsequencing. The resident Golgi protein nucleobindin and fructose‐bisphosphate aldolase were preferentially associated with membranes of polarized cells, whereas αB crystallin was detected exclusively and in high amounts in unpolarized cells.