Coat proteins play multiple roles in the life cycle of a membrane-bound transport intermediate, functioning in lipid bilayer remodeling, cargo selection and targeting to an acceptor compartment. The Coat Protein complex II (COPII) coat is known to act in each of these capacities, but recent work highlights the necessity for numerous accessory factors at all stages of transport carrier existence. Here, we review recent findings that highlight the roles of COPII and its regulators in the biogenesis of tubular COPII-coated carriers in mammalian cells that enable cargo transport between the endoplasmic reticulum and ER-Golgi intermediate compartments, the first step in a series of trafficking events that ultimately allows for the distribution of biosynthetic secretory cargoes throughout the entire endomembrane system. K E Y W O R D S COPII coat, early secretory pathway, endoplasmic reticulum, ER exit site, posttranslational modification, Sec16A, Tango1/cTAGE5, TFG

Integrins are a family of heterodimeric transmembrane receptors that mediate adhesion to the extracellular matrix (ECM). In addition to their role as adhesion receptors, integrins can act as “bidirectional signal transducers” that coordinate a large number of cellular activities in response to the extracellular environment and intracellular signaling events. This bidirectional signaling helps maintain tissue homeostasis. Dysregulated bidirectional signaling, however, could trigger the propagation of feedback loops that can lead to the establishment of a disease state such as glaucoma. Here we discuss the role of integrins and bidirectional signaling as they relate to the glaucomatous phenotype with special emphasis on the αvβ3 integrin. We present evidence that this particular integrin may have a significant impact on the pathogenesis of glaucoma.

PURPOSE. Fibronectin fibrillogenesis is an integrin-mediated process that may contribute to the pathogenesis of primary open-angle glaucoma (POAG). Here, we examined the effects of avb3 integrins on fibrillogenesis in immortalized TM-1 cells and human trabecular meshwork (HTM) cells. METHODS. TM-1 cells overexpressing wild-type b3 (WTb3) or constitutively active b3 (CAb3) integrin subunits were generated. Control cells were transduced with an empty vector (EV). Deoxycholic acid (DOC) extraction of monolayers, immunofluorescence microscopy, and Oncell western analyses were used to determine levels of fibronectin fibrillogenesis and fibronectin fibril composition (EDAþ and EDBþ fibronectins) and conformation. avb3 and a5b1 Integrin levels were determined using fluorescence-activated cell sorting (FACS). Cilengitide and an adenovirus vector expressing WTb3 or CAb3 integrin subunits were used to examine the role of avb3 integrin in HTM cells. The role of the canonical a5b1 integrinmediated pathway in fibrillogenesis was determined using the fibronectin-binding peptide FUD, the b1 integrin function-blocking antibody 13, and the Rho kinase (ROCK) inhibitor Y27632. RESULTS. Activation of avb3 integrin enhanced the assembly of fibronectin into DOC-insoluble fibrils in both TM-1 and HTM cells. The formation of fibronectin fibrils was dependent on a5b1 integrin and could be inhibited by FUD. However, fibrillogenesis was unaffected by Y27632. Fibrils assembled by CAb3 cells also contained high levels of EDAþ and EDBþ fibronectin and fibronectin that was stretched. CONCLUSIONS. avb3 Integrin signaling altered the deposition and structure of fibronectin fibrils using a b1 integrin/ROCK-independent mechanism. Thus, avb3 integrins could play a significant role in altering the function of fibronectin matrices in POAG.

SUMMARYLength-dependent axonopathy of the corticospinal tract causes lower limb spasticity and is characteristic of several neurological disorders, including hereditary spastic paraplegia (HSP) and amyotrophic lateral sclerosis. Mutations in Trk-fused gene (TFG) have been implicated in both diseases, but the pathomechanisms by which these alterations cause neuropathy remain unclear. Here, we biochemically and genetically define the impact of a mutation within the TFG coiled-coil domain, which underlies earlyonset forms of HSP. We find that the TFG (p.R106C) mutation alters compaction of TFG ring complexes, which play a critical role in the export of cargoes from the endoplasmic reticulum (ER). Using CRISPR-mediated genome editing, we engineered human stem cells that express the mutant form of TFG at endogenous levels and identified specific defects in secretion from the ER and axon fasciculation following neuronal differentiation. Together, our data highlight a key role for TFG-mediated protein transport in the pathogenesis of HSP.

Most metazoan organisms have evolved a mildly acidified and calcium diminished sorting hub in the early secretory pathway commonly referred to as the Endoplasmic Reticulum-Golgi intermediate compartment (ERGIC). These membranous vesicular-tubular clusters are found tightly juxtaposed to ER subdomains that are competent for the production of COPII-coated transport carriers. In contrast to many unicellular systems, metazoan COPII carriers largely transit just a few hundred nanometers to the ERGIC, prior to COPI-dependent transport on to the cis-Golgi. The mechanisms underlying formation and maintenance of ERGIC membranes are poorly defined. However, recent evidence suggests an important role for Trk-fused gene (TFG) in regulating the integrity of the ER/ERGIC interface. Moreover, in the absence of cytoskeletal elements to scaffold tracks on which COPII carriers might move, TFG appears to promote anterograde cargo transport by locally tethering COPII carriers adjacent to ERGIC membranes. This action, regulated in part by the intrinsically disordered domain of TFG, provides sufficient time for COPII coat disassembly prior to heterotypic membrane fusion and cargo delivery to the ERGIC.