Quantitative proteomics of caveolin-1-regulated proteins: characterization of polymerase i and transcript release factor/CAVIN-1 IN endothelial cells

Mol Cell Proteomics. 2010 Oct;9(10):2109-24. doi: 10.1074/mcp.M110.001289. Epub 2010 Jun 28.

Abstract

Caveolae are organelles abundant in the plasma membrane of many specialized cells including endothelial cells (ECs), epithelial cells, and adipocytes, and in these cells, caveolin-1 (Cav-1) is the major coat protein essential for the formation of caveolae. To identify proteins that require Cav-1 for stable incorporation into membrane raft domains, a quantitative proteomics analysis using isobaric tagging for relative and absolute quantification was performed on rafts isolated from wild-type and Cav-1-deficient mice. In three independent experiments, 117 proteins were consistently identified in membrane rafts with the largest differences in the levels of Cav-2 and in the caveola regulatory proteins Cavin-1 and Cavin-2. Because the lung is highly enriched in ECs, we validated and characterized the role of the newly described protein Cavin-1 in several cardiovascular tissues and in ECs. Cavin-1 was highly expressed in ECs lining blood vessels and in cultured ECs. Knockdown of Cavin-1 reduced the levels of Cav-1 and -2 and weakly influenced the formation of high molecular weight oligomers containing Cav-1 and -2. Cavin-1 silencing enhanced basal nitric oxide release from ECs but blocked proangiogenic phenotypes such as EC proliferation, migration, and morphogenesis in vitro. Thus, these data support an important role of Cavin-1 as a regulator of caveola function in ECs.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't
  • Validation Study

MeSH terms

  • Animals
  • Base Sequence
  • Blotting, Western
  • Caveolin 1 / genetics
  • Caveolin 1 / metabolism*
  • Cell Line
  • Cell Movement
  • Cell Proliferation
  • Chromatography, Ion Exchange
  • DNA Polymerase I / metabolism*
  • Gene Silencing
  • Humans
  • Mass Spectrometry
  • Mice
  • Mice, Knockout
  • Microscopy, Fluorescence
  • Nitric Oxide / metabolism
  • Proteomics*
  • RNA, Small Interfering

Substances

  • Cav1 protein, mouse
  • Caveolin 1
  • RNA, Small Interfering
  • Nitric Oxide
  • DNA Polymerase I