Immunodetection of α-Actinin in Focal Adhesions Is Limited by Antibody Inaccessibility

Fredrick M. Pavalko, Galen Schneider, Keith Burridge, Soo Siang Lim

Research output: Contribution to journalArticle

22 Scopus citations

Abstract

In this study we demonstrate that α-actinin is a prominent component of the focal adhesions of nonmuscle cells but that the α-actinin in focal adhesions is largely inaccessible to staining with antibodies against α-actinin. Our results explain a controversy that has existed in the literature. Investigators who microinject α-actinin into nonmuscle cells have routinely observed significant incorporation of α-actinin into focal adhesions as well as stress fibers. Immunofluorescence and immunoelectron microscopy have, however, indicated that α-actinin is located farther from the membrane than either talin or vinculin. Immunofluorescence studies of smooth muscle dense plaques and myotendinous junctions have also yielded conflicting results regarding the presence or absence of α-actinin at these sites. Here, we confirm that α-actinin immunofluorescence of fibroblasts yields weak or absent staining of focal adhesions. We also demonstrate that microinjected α-actinin readily incorporates into focal adhesions. However, various antisera against either the cell's endogenous αactinin or against the microinjected chicken gizzard αactinin fail to stain focal adhesions despite the presence of microinjected α-actinin at these sites. Furthermore, disassembly of stress fibers induced by dibutyrl cAMP demonstrates that α-actinin persists in focal adhesions in the absence of associated stress fibers, suggesting that α-actinin's association with focal adhesions is independent of stress fibers.

Original languageEnglish (US)
Pages (from-to)534-540
Number of pages7
JournalExperimental Cell Research
Volume217
Issue number2
DOIs
StatePublished - Apr 1995

    Fingerprint

ASJC Scopus subject areas

  • Cell Biology

Cite this