Cryo-STEM Tomography Reveals the Outcome of Extracellular Matrix Assembly in the Absence of Extracellular Disulfide Bonding

The extracellular matrix (ECM) is a glycoprotein network that surrounds cells, providing them with mechanical support and regulating their growth, movement, and differentiation. Mutations that affect the structural organization of ECM cause myopathies, and altered ECM production and assembly may contribute to cancer metastasis. ECM is formed from precursor proteins secreted from cells along with enzymes that catalyze crucial steps in ECM formation or stabilization. One such enzyme is quiescin sulfhydryl oxidase 1 (QSOX1), which catalyzes disulfide bond formation and is secreted from fibroblast cells producing ECM. We are interested in the effects of QSOX1 on the ECM because QSOX1 is over-expressed in the stroma of aggressive breast cancers and may contribute to ECM remodeling in this context. To gain insight into the role of QSOX1 activity in ECM assembly, we use cryo-scanning transmission electron microscopy (cryo-STEM) tomography to visualize the 3D organization of major ECM components in situ and at nanometer-scale resolution in cultures of normal fibroblasts compared to fibroblasts lacking QSOX1. This study revealed disruption of ECM organization, and suggested altered ECM turnover, when QSOX1-mediated disulfide bonding was absent. The ECM components altered here in the absence of QSOX1 may be the possible enzymatic targets of over-expressed QSOX1 in cancer.