A new mouse mutant with cleavage-resistant versican and isoform-specific versican mutants demonstrate that proteolysis at the Glu441-Ala442 peptide bond in the V1 isoform is essential for interdigital web regression
Matrix Biology Plus
Two inherent challenges in the mechanistic interpretation of protease-deficient phenotypes are defining the specific substrate cleavages whose reduction generates the phenotypes and determining whether the phenotypes result from loss of substrate function, substrate accumulation, or loss of a function(s) embodied in the substrate fragments. Hence, recapitulation of a protease-deficient phenotype by a cleavage-resistant substrate would stringently validate the importance of a proteolytic event
... proteolytic event and clarify the underlying mechanisms. Versican is a large proteoglycan required for development of the circulatory system and proper limb development, and is cleaved by ADAMTS proteases at the Glu 441 -Ala 442 peptide bond located in its alternatively spliced GAGb domain. Specific ADAMTS protease mutants have impaired interdigit web regression leading to soft tissue syndactyly that is associated with reduced versican proteolysis. Versikine, the N-terminal proteolytic fragment generated by this cleavage, restores interdigit apoptosis in ADAMTS mutant webs. Here, we report a new mouse transgene, Vcan AA , with validated mutations in the GAGb domain that specifically abolish this proteolytic event. Vcan AA/AA mice have partially penetrant hindlimb soft tissue syndactyly. However, Adamts20 inactivation in Vcan AA/AA mice leads to fully penetrant, more severe syndactyly affecting all limbs, suggesting that ADAMTS20 cleavage of versican at other sites or of other substrates is an additional requirement for web regression. Indeed, immunostaining with a neoepitope antibody against a cleavage site in the versican GAGa domain demonstrated reduced staining in the absence of ADAMTS20. Significantly, mice with deletion of Vcan exon 8, encoding the GAGb domain, consistently developed soft tissue syndactyly, whereas mice unable to include exon 7, encoding the GAGa domain in Vcan transcripts, consistently had fully separated digits. These findings suggest that versican is cleaved within each GAG-bearing domain during web regression, and affirms that proteolysis in the GAGb domain, via generation of versikine, has an essential role in interdigital web regression. Ó 2021 Published by Elsevier B.V. This is an open access article under the CC BY-NC-ND license (http://creativecommons. org/licenses/by-nc-nd/4.0/). Research Article Please cite this article as: S Nandadasa, CBurin des Roziers, G. et al., A new mouse mutant with cleavage-resistant versican and isoform-specific versican mutants demonstrate that proteolysis at the Glu 441 -Ala 442 peptide bond in the V1 isoform is essential for interdigital web regression , Materials Biology, https://doi.