Computational and experimental characterization of the novel ECM glycoprotein SNED1 and prediction of its interactome

The extracellular matrix (ECM) is a complex meshwork of proteins and an essential component of multicellular life. We have recently reported the characterization of a novel ECM protein, SNED1, and showed that it promotes breast cancer metastasis and regulates craniofacial development. However, the mechanisms by which it does so remain unknown. ECM proteins exert their functions by binding to cell surface receptors and interacting with other ECM proteins, actions that we can predict using knowledge of protein's sequence, structure, and post-translational modifications. Here, we combined insilico and in-vitro approaches to characterize the physico-chemical properties of SNED1 and infer its putative functions. To do so, we established a mammalian cell system to produce and purify SNED1 and its N-terminal fragment, which contains a NIDO domain, and demonstrated experimentally SNED1's potential to be glycosylated, phosphorylated, and incorporated into an insoluble ECM. We also determined the secondary and tertiary structures of SNED1 and its N-terminal fragment and obtained a model for its NIDO domain. Using computational predictions, we identified 114 proteins as putative SNED1 interactors, including the ECM protein fibronectin. Pathway analysis of the predicted SNED1 interactome further revealed that it may contribute to signaling through cell surface receptors, such as integrins, and participate in the regulation of ECM organization and developmental processes. Last, using fluorescence microscopy, we showed that SNED1 forms microfibrils within the ECM and partially colocalizes with fibronectin. Altogether, we provide a wealth of information on an understudied yet important ECM protein with the potential to decipher its pathophysiological functions.

Automated summary

This study presents a novel ECM protein, SNED1, which promotes breast cancer metastasis and regulates craniofacial development. By combining computational predictions and experimental approaches, the physico-chemical properties of SNED1 were characterized, and its potential functions were inferred, shedding light on its pathophysiological roles.