SPARC, a secreted protein associated with cellular proliferation, inhibits cell spreading in vitro and exhibits Ca+2-dependent binding to the extracellular matrix.

Abstract

Abstract. SPARC (Secreted Protein Acidic and Richin Cysteine) is a Ca÷2-binding glycoprotein that is differentially associated with morphogenesis, remodeling, cellular migration, and proliferation. We show here that exogenous SPARC, added to cells in culture, was associated with profound changes in cell shape, caused rapid, partial detachment of a confluent monolayer, and inhibited spreading of newly plated cells.Bovine aortic endothelial cells, exposed to 2-40/zg SPARC/ml per 2 x 106 cells, exhibited a rounded morphology in a dose-dependent manner but remained attached to plastic or collagen-coated surfaces. These round cells synthesized protein, uniformly excluded trypan blue, and grew in aggregates after replating in media without SPARC. SPARC caused rounding of bovine endothelial cells, fibroblasts, and smooth muscle cells; however, the cell lines F9, PYS-2, and 3T3 were not affected. The activity of native SPARC was inhibited by heat denaturation and prior incubation with anti-SPARC IgG. The effect of SPARC on endothelial cells appeared to be independent of the rounding phenomenon produced by the peptide GRGDSP. Immunofluorescence localization of SPARC on endothelial cells showed preferential distribution at the leading edges of membranous extensions.SPARC bound Ca ÷2 in both amino-and carboxylterminal (EF-hand) domains and required this cation for maintenance of native structure. Solid-phase binding assays indicated a preferential affinity of native SPARC for several proteins comprising the extracellular matrix, including types III and V collagen, and thrombospondin. This binding was saturable, Ca +2 dependent, and inhibited by anti-SPARC IgG. Endothelial cells also failed to spread on a substrate of native type III collagen complexed with SPARC. We propose that SPARC is an extracellular modulator of Ca ÷2 and cation-sensitive proteins or proteinases, which facilitates changes in cellular shape and disengagement of cells from the extracellular matrix.

SPARC is expressed by macroglia and microglia in the developing and mature nervous system

“…SPARC can be secreted to regulate local growth factor, ECM, and matrix metalloproteinase (MMP) activity, to modulate cell morphology and migration responses, angiogenesis, and cell proliferation and survival (Rempel et al, 2001;Francki et al, 2004;Barker et al, 2005a;Yan et al, 2005;Said et al, 2007a). In vitro, SPARC acts on many cell types as a deadhesive factor that induces cell rounding and inhibits cell migration (Sage et al, 1989b;Bradshaw and Sage, 2001), including on neuronal and glial cell lines (Ikemoto et al, 2000). However, SPARC is thought to act in vivo as a contextual regulator of cellular adhesion strength (Greenwood and Murphy-Ullrich, 1998), and is required for fibroblast migration during dermal wound healing (Basu et al, 2001), leukocyte infiltration after immune challenge (Kelly et al, 2007;Rempel et al, 2007), and tumor invasiveness (Rempel et al, 2001;Framson and Sage, 2004).…”

Frequent inactivation of SPARC by promoter hypermethylation in colon cancers

“…9,24 This suppression may be related to the rapid growth of tumors, since SPARC has an antiproliferative function through the modulation of cell cycle regulatory proteins or growth factors. 33 When we evaluated the relationship between SPARC suppression and clinicopathologic factors in 292 colorectal carcinomas, suppression of SPARC was not associated with age, sex, tumor site, MSI status or tumor stage. However, SPARC suppression was associated with decreased mucin production and with poor prognosis.…”

Ectopic Expression of SPARC inXenopusEmbryos Interferes with Tissue Morphogenesis: Identification of a Bioactive Sequence in the C-terminal EF Hand

“…Rat bone sialoprotein cDNA (a gift of Dr. Jaro Sodek, University of Toronto) was subcloned and transcribed similarly. Intact mouse SPARC protein was isolated from the conditioned medium of PYS-2 cells and was purified as previously described (Sage et al 1989). This protein preparation was shown to disrupt barrier function in cultured endothelial cells, as described by Goldblum et al (1994), and to inhibit endothelial and smooth muscle cell proliferation in vitro, according to Funk and Sage (1993).…”

“…Ectopic expression of SPARC produced comparable muscle defects in C. elegans (Schwarzbauer and Spencer 1993). Although the precise functions of SPARC during early embryonic development are not known, a variety of potential functions have been described for SPARC in vitro, e.g., inhibition of cell-substratum adhesion and cell spreading (Sage et al 1989), binding to and/or modulation of growth factors (Hasselaar and Sage 1992;Raines et al 1992), regulation of cell cycle progression Sage 1991, 1993), and augmentation of interendothelial cell permeability (Goldblum et al 1994; reviewed in Lane and Sage 1994). Furthermore, SPARC interacts with several ECM macromolecules in vitro (Tremble et al 1993;Kelm and Mann 1991).…”