Distribution of caveolin-1 and connexin43 in normal and injured alveolar epithelial R3/1 cells.
Using the new alveolar epithelial type I-like cell line R3/1 derived from fetal rat lung, we studied the distribution of connexin43 and caveolin-1 under conditions of bleomycin-induced injury in vitro. We show that under normal as well as under conditions of injury, endogenous connexin43 does not directly interact with endogenous caveolin-1 as revealed by immunofluorescence, glutathione S-transferase/caveolin-1 "pull down" assay, and co-immunoprecipitation experiments. The assessment of Triton X-100 solubility revealed that caveolin-1 was abundant in detergent-resistant membrane fractions. This is consistent with the localization of caveolin-1 in the lipid rafts/caveolae. Similarly, phosphorylated connexin43 was preferably detected in the Triton-insoluble fraction. Using a sucrose gradient we demonstrated that the majority of phosphorylated connexin43 colocalizes with caveolin-1 in lipid rafts, whereas all other forms of connexin43 remain in the bulk of cellular membranes and cytosolic proteins. Triton solubility assessment of bleomycin-treated cells revealed no differences in the caveolin-1 and connexin43 distribution. A further interesting outcome of our study is the shift of caveolin-1 from the lipid raft/caveolae fractions to the non-caveolar fractions after bleomycin treatment indicating an intracellular retention of caveolin-1. This result suggests the possibility that the translocation of caveolin-1 could be an important event regulating the metabolism of alveolar epithelial lung cells after injury.
Barth
K
K
Institute of Anatomy, Medical Faculty Carl Gustav Carus, Dresden University of Technology, Fiedlerstrasse 42, 01307 Dresden, Germany.
Gentsch
M
M
Bläsche
R
R
Pfüller
A
A
Parshyna
I
I
Koslowski
R
R
Barth
G
G
Kasper
M
M
D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D001761:Bleomycin; D051242:Caveolin 1; D022461:Caveolins / Q000378:metabolism*; D002460:Cell Line; D018031:Connexin 43 / Q000378:metabolism*; D004847:Epithelial Cells / Q000378:metabolism* / Q000473:pathology; D005455:Fluorescent Antibody Technique; D021962:Membrane Microdomains / Q000378:metabolism; D011650:Pulmonary Alveoli / Q000378:metabolism* / Q000473:pathology; D051381:Rats
C495537:Cav1 protein, rat; D051242:Caveolin 1; D022461:Caveolins; D018031:Connexin 43; D001761:Bleomycin