Developmental distribution of collagen IV isoforms and relevance to ocular diseases.
ABSTRACT Type IV collagens are the most abundant proteins in basement membranes. Distinct genes encode each of six isoforms, alpha1(IV) through alpha6(IV), which assemble into one of three characteristic heterotrimers. Disease-causing mutations in each of the six genes are identified in humans or mice and frequently include diverse ocular pathogenesis that encompass common congenital and progressive blinding diseases, such as optic nerve hypoplasia, glaucoma, and retinal degeneration. Understanding where and when collagen IV molecules are expressed is important because it defines limits for the location and timing of primary pathogenesis. Although localization of collagen IV isoforms in developed human eyes is known, the spatial and temporal distribution of type IV collagens throughout ocular development has not been determined in humans or in mice. Here, we use isoform-specific monoclonal antibodies to systematically reveal the localization of all six collagen IV isoforms in developing mouse eyes. We found that alpha1(IV) and alpha2(IV) always co-localized and were ubiquitously expressed throughout development. alpha3(IV) and alpha4(IV) also always co-localized but in a much more spatially and temporally specific manner than alpha1(IV) and alpha2(IV). alpha5(IV) co-localized both with alpha3(IV)/alpha4(IV), and with alpha6(IV), consistent with alpha5(IV) involvement in two distinct heterotrimers. alpha5(IV) was present in all basement membranes except those of the vasculature. alpha6(IV) was not detected in vasculature or in Bruch's membrane, indicating that alpha5(IV) in Bruch's membrane is part of the alpha3alpha4alpha5 heterotrimer. This comprehensive analysis defines the spatial and temporal distribution of type IV collagen isoforms in the developing eye, and will contribute to understanding the mechanisms underlying collagen IV-related ocular diseases that collectively lead to blindness in millions of people worldwide.
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ABSTRACT: ER-to-Golgi transport of proteins destined for the extracellular space or intracellular compartments depends on the COPII vesicle coat and is constitutive in all translationally active cells. Nevertheless, there is emerging evidence that this process is regulated on a cell- and tissue-specific basis, which means that components of the COPII coat will be of differential importance to certain cell types. The COPII coat consists of an inner layer, Sec23/24 and an outer shell, Sec13/31. We have shown previously that knock-down of Sec13 results in concomitant loss of Sec31. In zebrafish and cultured human cells this leads to impaired trafficking of large cargo, namely procollagens, and is causative for defects in craniofacial and gut development. It is now widely accepted that the outer COPII coat is key to the architecture and stability of ER export vesicles containing large, unusual cargo proteins. Here, we investigate zebrafish eye development following Sec13 depletion. We find that photoreceptors degenerate or fail to develop from the onset. Impaired collagen trafficking from the retinal pigment epithelium and defects in overall retinal lamination also seen in Sec13-depleted zebrafish might have been caused by increased apoptosis and reduced topical proliferation in the retina. Our data show that the outer layer of the COPII coat is also necessary for the transport of large amounts of cargo proteins, in this case rhodopsin, rather than just large cargo as previously thought. Full text available here (open access): http://bio.biologists.org/content/early/2013/01/10/bio.20133251.abstractBiology open. 03/2013; 2(3):256-66.
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ABSTRACT: During development, wound repair and disease-related processes, such as cancer, normal, or neoplastic cell types traffic through the extracellular matrix (ECM), the complex composite of collagens, elastin, glycoproteins, proteoglycans, and glycosaminoglycans that dictate tissue architecture. Current evidence suggests that tissue-invasive processes may proceed by protease-dependent or protease-independent strategies whose selection is not only governed by the characteristics of the motile cell population, but also by the structural properties of the intervening ECM. Herein, we review the mechanisms by which ECM dimensionality, elasticity, crosslinking, and pore size impact patterns of cell invasion. This summary should prove useful when designing new experimental approaches for interrogating invasion programs as well as identifying potential cellular targets for next-generation therapeutics.Journal of Microscopy 09/2013; 251(3):250-60. · 1.63 Impact Factor
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ABSTRACT: Molecular transport through the basement membrane is important for a number of physiological functions, and dysregulation of basement membrane architecture can have serious pathological consequences. The structure-function relationships that govern molecular transport in basement membranes are not fully understood. The basement membrane from the lens capsule of the eye is a collagen IV-rich matrix that can easily be extracted and manipulated in vitro. As such, it provides a convenient model for studying the functional relationships that govern molecular transport in basement membranes. Here we investigate the effects of increased transmembrane pressure and solute electrical charge on the transport properties of the lens basement membrane (LBM) from the bovine eye. Pressure-permeability relationships in LBM transport were governed primarily by changes in diffusive and convective contributions to solute flux and not by pressure-dependent changes in intrinsic membrane properties. The solute electrical charge had a minimal but statistically significant effect on solute transport through the LBM that was opposite of the expected electrokinetic behavior. The observed transport characteristics of the LBM are discussed in the context of established membrane transport modeling and previous work on the effects of pressure and electrical charge in other basement membrane systems.Biophysical Journal 04/2013; 104(7):1476-84. · 3.67 Impact Factor