Abstract
Low density lipoprotein receptor-related protein-1 (LRP-1) mediates the endocytosis of multiple plasma membrane proteins and thereby models the composition of the cell surface. LRP-1 also functions as a catabolic receptor for fibronectin, limiting fibronectin accumulation in association with cells. The goal of the present study was to determine whether LRP-1 regulates cell surface levels of the beta(1) integrin subunit. We hypothesized that LRP-1 may down-regulate cell surface beta(1) by promoting its internalization; however, unexpectedly, LRP-1 expression was associated with a substantial increase in cell surface beta(1) integrin in two separate cell lines, murine embryonic fibroblasts (MEFs) and CHO cells. The total amount of beta(1) integrin was unchanged because LRP-1-deficient cells retained increased amounts of beta(1) in the endoplasmic reticulum (ER). Expression of human LRP-1 in LRP-1-deficient MEFs reversed the shift in subcellular beta(1) integrin distribution. Metabolic labeling experiments demonstrated that the precursor form of newly synthesized beta(1) integrin (p105) is converted into mature beta(1) (p125) more slowly in LRP-1-deficient cells. Although low levels of cell surface beta(1) integrin, in LRP-1-deficient MEFs, were associated with decreased adhesion to fibronectin, the subcellular distribution of beta(1) integrin was most profoundly dependent on LRP-1 only after the cell cultures became confluent. A mutagen-treated CHO cell line, in which LRP-1 is expressed but retained in the secretory pathway, also demonstrated nearly complete ER retention of beta(1) integrin. These studies support a model in which LRP-1 either directly or indirectly promotes maturation of beta(1) integrin precursor and thereby increases the level of beta(1) integrin at the cell surface.
Highlights
The 1 integrin subunit associates with multiple ␣-subunits to form transmembrane adhesion receptors for extracellular matrix proteins, including collagen, fibronectin, vitronectin, and laminin [1]
We hypothesized that lipoprotein receptor-related protein-1 (LRP-1) may down-regulate cell surface 1 by promoting its internalization; unexpectedly, LRP-1 expression was associated with a substantial increase in cell surface 1 integrin in two separate cell lines, murine embryonic fibroblasts (MEFs) and CHO cells
LRP-1-deficient MEFs Demonstrate Decreased p125/Cell Surface 1 Integrin—1 integrin was compared by immunoblot analysis in LRP-1-deficient MEF-2 cells and in two LRP-1positive cell lines (MEF-1 and PEA10)
Summary
The 1 integrin subunit associates with multiple ␣-subunits to form transmembrane adhesion receptors for extracellular matrix proteins, including collagen, fibronectin, vitronectin, and laminin [1]. Once present at the cell surface, mature integrins anchor the plasma membrane to the actin cytoskeleton and promote cell signaling [2, 3]. The membrane-bound protein chaperone, calnexin, associates with 1 in the ER, promoting integrin assembly but inhibiting integrin transfer to the cell surface [12, 13]. In addition to soluble ligands, LRP-1 mediates the endocytosis of other plasma membrane proteins, including the urokinase receptor (uPAR), tissue factor, and amyloid precursor protein (APP), and thereby down-regulates the plasma membrane levels of these proteins [21,22,23,24,25]. Regulation of the concentration and activity of membrane proteins represents an indirect mechanism by which LRP-1 may control cell signaling. By down-regulating cell surface uPAR in murine embryonic
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