Neutrophil elastase cleavage of the gC1q domain impairs the EMILIN1-α4β1 integrin interaction, cell adhesion and anti-proliferative activity

The extracellular matrix glycoprotein EMILIN1 exerts a wide range of functions mainly associated with its gC1q domain. Besides providing functional significance for adhesion and migration, the direct interaction between alpha 4 beta 1 integrin and EMILIN1-gC1q regulates cell proliferation, transducing net anti-proliferative effects. We have previously demonstrated that EMILIN1 degradation by neutrophil elastase (NE) is a specific mechanism leading to the loss of functions disabling its regulatory properties. In this study we further analysed the proteolytic activity of NE, MMP-3, MMP-9, and MT1-MMP on EMILIN1 and found that MMP-3 and MT1- MMP partially cleaved EMILIN1 but without affecting the functional properties associated with the gC1q domain, whereas NE was able to fully impair the interaction of gC1q with the alpha 4 beta 1 integrin by cleaving this domain outside of the E933 integrin binding site. By a site direct mutagenesis approach we mapped the bond between S913 and R914 residues and selected the NE-resistant R914W mutant still able to interact with the alpha 4 beta 1 integrin after NE treatment. Functional studies showed that NE impaired the EMILIN1-alpha 4 beta 1 integrin interaction by cleaving the gC1q domain in a region crucial for its proper structural conformation, paving the way to better understand NE effects on EMILIN1-cell interaction in pathological context.