Membrane Localization of the Pertussis Toxin-Sensitive G-Protein Subunits αi-2and αi-3and Expression of a Metallothionein- αi-2Fusion Gene in LLC-PK1Cells

The renal epithelial cell line LLC-PK1has topographically distinct regulatory roles for the α subunits of pertussis toxin-sensitive guanine nucleotide regulatory proteins (αisubunit); these include the inhibition of adenylyl cyclase at the basolateral membrane and the stimulation of Na+channel activ...

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Bibliographic Details
Published inProceedings of the National Academy of Sciences - PNAS Vol. 87; no. 12; pp. 4635 - 4639
Main Authors Ercolani, Louis, Stow, Jennifer L., Boyle, Jane F., Holtzman, Eliezer J., Lin, Herbert, Grove, J. Russell, Ausiello, Dennis A.
Format Journal Article
LanguageEnglish
Published National Academy of Sciences of the United States of America 01.06.1990
Subjects
Antibodies
Cell lines
Cell membranes
Cells
Complementary DNA
Epithelial cells
Gels
LLC PK1 cells
Messenger RNA
RNA
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Summary:The renal epithelial cell line LLC-PK1has topographically distinct regulatory roles for the α subunits of pertussis toxin-sensitive guanine nucleotide regulatory proteins (αisubunit); these include the inhibition of adenylyl cyclase at the basolateral membrane and the stimulation of Na+channel activity at the apical membrane. We now report that LLC-PK1cells contain two members of the αiprotein family, αi - 2and αi - 3, which have distinct cellular locations consistent with their diverse functional roles. By using specific αiantibodies and immunofluorescence, the αi - 2subunit was found to be localized to the basolateral membrane, whereas the αi - 3subunit was concentrated in the Golgi and was also detectable at low levels on apical membranes in some cells. Induction of a chimeric mouse metallothionein 1-rat or canine αi - 2gene stably transfected into the LLC-PK1cells produced an increase in the content of the αi - 2subunit, which was targeted only to the basolateral membrane. These findings suggest that αisubunit specificity for effectors may be achieved in polarized renal epithelial cells by their geographic segregation to different cellular membranes. The LLC-PK1cell stably transfected with the metallothionein-αi - 2fusion gene will provide a model for the study of guanine nucleotide regulatory protein function in epithelia.
ISSN:0027-8424
1091-6490