Publications [#112800] of G Vann V. Bennett

Papers Published

1. RJ Hu, S Moorthy, V Bennett, Expression of functional domains of beta G-spectrin disrupts epithelial morphology in cultured cells., The Journal of cell biology, UNITED STATES, vol. 128 no. 6 (March, 1995), pp. 1069-80, ISSN 0021-9525
   (last updated on 2003/02/18)

   Abstract:
   Spectrin is a major structural protein associated with the cytoplasmic surface of plasma membranes of many types of cells. To study the functions of spectrin, we transfected Caco-2 intestinal epithelial cells with a plasmid conferring neomycin resistance and encoding either actin-binding or ankyrin-binding domains of beta G-spectrin fused with beta-galactosidase. These polypeptides, in principle, could interfere with the interaction of spectrin with actin or ankyrin, as well as block normal assembly of alpha- and beta-spectrin subunits. Cells expressing the fusion proteins represented only a small fraction of neomycin-resistant cells, but they could be detected based on expression of beta-galactosidase. Cells expressing spectrin domains exhibited a progressive decrease in amounts of endogenous beta G-spectrin, although alpha-spectrin was still present. Beta G-spectrin-deficient cells lost epithelial cell morphology, became multinucleated, and eventually disappeared after 10-14 d in culture. Spectrin-associated membrane proteins, ankyrin and adducin, as well as the Na+,K(+)-ATPase, which binds to ankyrin, exhibited altered distributions in cells transfected with beta G-spectrin domains. E-cadherin and F-actin, in contrast to ankyrin, adducin, and the Na+,K(+)-ATPase, were expressed, and they exhibited unaltered distribution in beta G-spectrin-deficient cells. Cells transfected with the same plasmid encoding beta-galactosidase alone survived in culture as the major population of neomycin-resistant cells, and they exhibited no change in morphology or in the distribution of spectrin-associated membrane proteins. These results establish that beta G-spectrin is essential for the normal morphology of epithelial cells, as well as for their maintenance in monolayer culture.

   Keywords:
   Ankyrins • Cell Death • Cells, Cultured • Epithelial Cells* • Epithelium • Gene Transfer Techniques • Giant Cells • Humans • Intestines • Microfilament Proteins • Recombinant Fusion Proteins • Spectrin • biosynthesis* • genetics • metabolism • metabolism*