For cancer progression and metastatic dissemination, cancer cells migrate and penetrate through extracellular tissues. Cancer invasion is frequently facilitated by proteolytic processing of components of the extracellular matrix (ECM). The cellular regions mediating proteolysis are diverse and depend upon the physical structure, composition, and dimensionality of the ECM contacted by the cell surface. Cancer cells migrating across 2D substrate contain proteolytic structures such as lamellipodia, invadopodia, and the trailing edge. Likewise, invasive mesenchymal migration through 3D fibrillar ECM, as monitored for HT1080 fibrosarcoma and MDA-MB-231 breast carcinoma cells by submicron-resolved imaging, is mediated by several types of proteolytic structures rich in filamentous actin, ss1 integrin, and MT1-MMP with distinct location and function. These comprise (i) anterior pseudopod bifurcataions and the nucleus corresponding to zones of local cell compression by constraining collagen fibers, (ii) lateral small spikes that protrude into the ECM and cause small spot-like proteolytic foci, and (iii) a strongly proteolytic trailing edge sliding along reorganized ECM fibers. Through their combined action these proteolytic surface structures cleave, remove, and realign ECM barriers, support rear end retraction, generate tube-like matrix defects and laterally widen existing tracks during 3D tissue invasion.