Ligation of integrin alpha(5)beta(1) is required for internalization of vitronectin by integrin alpha(v)beta(3)

Remodeling of the matrix by tumor cells is necessary for tumor invasion. We have shown previously that malignant astrocytomas, in contrast to normal astrocytes, synthesize vitronectin and express integrins alpha(v) beta(3) and alpha(v) beta(5). The activity states of these two integrins are differentially controlled. Thus, we investigated the regulation of the activity of integrins alpha(v) beta(3) and alpha(v) beta(5) with regard to their role in vitronectin internalization in U-251MG astrocytoma cell monolayers adherent to fibronectin, collagen, or laminin in serum-free conditions. Binding of [I-125]vitronectin occurred in a specific, saturable manner that was partially inhibitable by monoclonal antibodies (mAbs) specific for integrins alpha(v) beta(3) or alpha(v) beta(5). Specific, lysosomally-mediated degradation of [I-125]vitronectin was detectable at 1 h and increased over the 24 h assay period. The cell substrate affected the rate of turnover of [I-125]vitronectin, which was 3.0 ng/min for cells plated on fibronectin but 0.35 ng/min for cells plated on collagen. Furthermore, although mAbs specific for either integrin alpha(v) beta(3) or alpha(v) beta(5) inhibited degradation (30%; combined effect 70%) of [I-125]vitronectin by cells plated on fibronectin, only mAb anti-alpha(v) beta(5) inhibited degradation (70-90%) by cells plated on collagen or laminin. To determine the requirement for integrin alpha(5) beta(1) ligation in order for integrin alpha(v) beta(3) to internalize its ligand, cells were plated on mAbs anti-integrin alpha(5) or anti-integrin alpha(3). When plated on mAb anti-alpha(5), mAbs anti-alpha(v) beta(3), and anti-alpha(v) beta(5) both inhibited degradation. However, when plated on mAb anti-alpha(3), mAb anti-alpha(v) beta(3) had no effect whereas mAb anti-alpha(v) beta(5) inhibited degradation. These data indicate that a signal from integrin alpha(5) beta(1) is necessary for integrain alpha(v) beta(3) to internalize vitronectin, whereas integrin alpha(v) beta(5) constitutively internalizes vitronectin.