Laminin-6 Is Activated by Proteolytic Processing and Regulates Cellular Adhesion and Migration Differently from Laminin-5

Abstract Laminin-6 (LN6) and laminin-5 (LN5), which share the common integrin-binding domain in the laminin α3 chain, are thought to cooperatively regulate cellular functions, but the former has poorly been characterized. Human fibrosarcoma HT1080 cells expressing an exogenous α3 chain were found to secrete LN6 with the full-length α3 chain and a smaller amount of its processed form lacking the carboxyl-terminal G4-5 domain, besides mature LN5 without G4-5 (mat-LN5). We prepared the unprocessed LN6 and mat-LN5, as well as LN6 mutants without G4-5 (LN6ΔG4-5) or G5 (LN6ΔG5). These laminins supported attachment of HT1080 cells and human keratinocytes (HaCaT) through integrins α3β1 and/or α6β1. LN6ΔG4-5, LN6ΔG5, and mat-LN5 promoted rapid cell spreading, whereas LN6 did hardly. A purified G4-5 fragment of the laminin α3 chain supported cell attachment through interaction with heparan sulfate proteoglycans and promoted cell spreading in combination with mat-LN5 or LN6ΔG4-5. These results imply that the G4-5 domain within the LN6 molecule suppresses cell adhesion, while the released G4-5 promotes it. The presence of G5 rather than the heparin-binding domain G4 was responsible for the impaired cell spreading activity of LN6. However, the unprocessed LN6 promoted cell spreading in the presence of mat-LN5. Unlike mat-LN5, both LN6ΔG4-5 and LN6 did weakly or did not stimulate cell motility. These findings demonstrate that LN6 and LN5 have distinct biological activities, but they may cooperatively support cell adhesion. The proteolytic processing of the α3 chain seems to regulate the physiological functions of LN6.