Transporting epithelial cells build apical microvilli to increase membrane surface area and enhance absorptive capacity. The intestinal brush border provides an elaborate example, with tightly packed microvilli that function in nutrient absorption and host defense. Although the brush border is essential for physiological homeostasis, its assembly is poorly understood. We found that brush border assembly is driven by the formation of Ca2+-dependent adhesion links between adjacent microvilli. Intermicrovillar links are composed of protocadherin-24 and mucin-like protocadherin, which target to microvillar tips and interact to form a trans heterophilic complex. The cytoplasmic domains of microvillar protocadherins interact with the scaffolding protein, harmonin, and myosin-7b, which promote localization to microvillar tips. Finally, a mouse model of Usher syndrome lacking harmonin exhibits microvillar protocadherin mislocalization and severe defects in brush border morphology. These data reveal an adhesion-based mechanism for brush border assembly and illuminate the basis of intestinal pathology in Usher syndrome patients.

The myelin sheath surrounding axons ensures that nerve impulses travel quickly and efficiently, allowing for the proper function of the vertebrate nervous system. We previously showed that the adhesion G-protein-coupled receptor (aGPCR) Gpr126 is essential for peripheral nervous system myelination, although the molecular mechanisms by which Gpr126 functions were incompletely understood. aGPCRs are a significantly understudied protein class, and it was unknown whether Gpr126 couples to G-proteins. Here, we analyze Dhh Cre ; Gpr126 fl/fl conditional mutants, and show that Gpr126 functions in Schwann cells (SCs) for radial sorting of axons and myelination. Furthermore, we demonstrate that elevation of cAMP levels or protein kinase A activation suppresses myelin defects in Gpr126 mouse mutants and that cAMP levels are reduced in conditional Gpr126 mutant peripheral nerve. Finally, we show that GPR126 directly increases cAMP by coupling to heterotrimeric G-proteins. Together, these data support a model in which Gpr126 functions in SCs for proper development and myelination and provide evidence that these functions are mediated via G-protein-signaling pathways.

The brush border domain at the apex of intestinal epithelial cells is the primary site of nutrient absorption in the intestinal tract and the primary surface of interaction with microbes that reside in the lumen. Because the brush border is positioned at such a critical physiological interface, we set out to create a comprehensive list of the proteins that reside in this domain using shotgun mass spectrometry. The resulting proteome contains 646 proteins with diverse functions. In addition to the expected collection of nutrient processing and transport components, we also identified molecules expected to function in the regulation of actin dynamics, membrane bending, and extracellular adhesion. These results provide a foundation for future studies aimed at defining the molecular mechanisms underpinning brush border assembly and function.

These data establish myosin-1d as a component of the brush border cytoskeleton that demonstrates microvillar tip localization.

Synovial sarcoma is a soft tissue cancer associated with a recurrent t(X:18) translocation that generates one of two fusion proteins, SYT-SSX1 or SYT-SSX2. In this study, we demonstrate that SYT-SSX2 is a unique oncogene. Rather than confer enhanced proliferation on its target cells, SYT-SSX2 instead causes a profound alteration of their architecture. This aberrant morphology included elongation of the cell body and formation of neurite-like extensions. We also observed that cells transduced with SYT-SSX2 often repulsed one another. Notably, cell repulsion is a known component of ephrin signaling. Further analysis of SYT-SSX2-infected cells revealed significant increases in the expression and activation of Eph/ephrin pathway components. On blockade of EphB2 signaling SYT-SSX2 infectants demonstrated significant reversion of the aberrant cytoskeletal phenotype. In addition, we discovered, in parallel, that SYT-SSX2 induced stabilization of the microtubule network accompanied by accumulation of detyrosinated Glu tubulin and nocodazole resistance. Glu tubulin regulation was independent of ephrin signaling. The clinical relevance of these studies was confirmed by abundant expression of both EphB2 and Glu tubulin in SYT-SSX2-positive synovial sarcoma tissues. These results indicate that SYT-SSX2 exerts part of its oncogenic effect by altering cytoskeletal architecture in an Eph-dependent manner and cytoskeletal stability through a concurrent and distinct pathway.