Integrin and dystroglycan compensate each other to mediate laminin-dependent basement membrane assembly and epiblast polarization

Abstract

During early embryogenesis, endodermal γ1-laminin expression is required for basement membrane (BM) assembly, promoting conversion of non-polar pluripotent cells into polarized epiblast. The influence of laminin-111 (Lm111) and its integrin and dystroglycan (DG) receptors on epiblast in embryoid bodies (EBs), a model for differentiation of the embryonic plate, was further investigated. Lm111 added to the medium of EBs initiated conversion of inner nonpolar cell to the polarized epiblast epithelium with an exterior-to-central basal-to-apical orientation. Microinjection of Lm111 into EB interiors resulted in an interior BM with complete inversion of cell polarity. Lm111 assembled a BM on integrin-β1 null EBs with induction of polarization at reduced efficiency. β-Integrin compensation was not detected in these nulls with integrin adaptor proteins failing to assemble. A dimer of laminin LG domains 4–5 (LZE3) engineered to strongly bind to α-dystroglycan almost completely inhibited laminin accumulation on integrin β1-null EBs, reducing BM and ablating cell polarization. When Lm111 was incubated with integrin-β1/dystroglycan double-knockout EBs, laminin failed to accumulate on the EBs, the EBs did not differentiate, and the EBs underwent apoptosis. Collectively the findings support the hypotheses that the locus of laminin cell surface assembly can determine the axis of epithelial polarity. This requires integrin- and/ or dystroglycan-dependent binding to laminin LG domains with the highest efficiency achieved when both receptors are present. Finally, EBs that cannot assemble a matrix undergo apoptosis.

α-integrins dictate distinct modes of type IV collagen recruitment to basement membranes

“…Due to the challenge of imaging of BMs in vivo, the lethal phenotypes of null mutations of many BM components, and the expanded gene families of BM receptors and matrix components in vertebrates, it has been difficult to establish the mechanisms that mediate type IV collagen recruitment to BM in vivo 19,33 . Studies in vitro suggest several mechanisms might recruit laminin to cell surfaces, which in turn mediates type IV collagen incorporation.…”

“…Work in cell culture has suggested that interactions with laminin-binding integrin and dystroglycan receptors as well as cell surface sulfated glycolipids might seed laminin polymerization to initiate BM assembly, leading to subsequent collagen recruitment 19,33 .…”

“…Work in Drosophila and mouse embryos have indicated type IV collagen recruitment to BMs requires the presence of laminin 9,17,18 . Cell culture work has suggested that laminin deposition onto cell surfaces is mediated through interactions with integrin and dystroglycan receptors, and with sulfated glycolipids 2,19,20 . The cell surface mechanisms that mediate laminin and collagen deposition in vivo, however, remain elusive.…”

Competence to epithelialise coincides with competence to differentiate in pluripotent cells

“…Pluripotent cells polarise before they commit to a primed state (Shahbazi et al, 2017). Polarisation is initiated by integrin-mediated adhesion to the laminin matrix secreted by the emerging extraembryonic endoderm (Bedzhov & Zernicka-Goetz, 2014;Li et al, 2016) and subsequently stabilised by PTEN (Meng et al, 2017) and Integrin-linked-kinase (Sakai, 2003). However, several open questions remain.…”

A scar-like lesion is apparent in basement membrane after wound repair in vivo

“…The first is laminin, a heterotrimeric protein that polymerizes into a two-dimensional scaffold. Laminin binds directly to cell-surface molecules such as integrins and dystroglycans, and these cell surface interactions increase the local concentration of laminin to promote polymerization at cell membranes [10], which appears to be the first step of making a new basement membrane. The second glycoprotein is collagen IV, a non-fibrillar collagen that assembles into a covalently reinforced sheet that gives basement membrane its mechanical stiffness.…”