Epithelial Shaping by Diverse Apical Extracellular Matrices Requires the Nidogen Domain Protein DEX-1 in Caenorhabditis elegans

Abstract

The body's external surfaces and the insides of biological tubes, like the vascular system, are lined by a lipid-, glycoprotein-, and glycosaminoglycan-rich apical extracellular matrix (aECM). aECMs are the body's first line of defense against infectious agents and promote tissue integrity and morphogenesis, but are poorly described relative to basement membranes and stromal ECMs. While some aECM components, such as zona pellucida (ZP) domain proteins, have been identified, little is known regarding the overall composition of the aECM or the mechanisms by which different aECM components work together to shape epithelial tissues. In Caenorhabditis elegans, external epithelia develop in the context of an ill-defined ZP-containing aECM that precedes secretion of the collagenous cuticle. C. elegans has 43 genes that encode at least 65 unique ZP proteins, and we show that some of these comprise distinct precuticle aECMs in the embryo. Previously, the nidogen-and EGF-domain protein DEX-1 was shown to anchor dendrites to the C. elegans nose tip in concert with the ZP protein DYF-7. Here, we identified a new, strong loss-of-function allele of dex-1, cs201. dex-1 mutants die as L1 larvae and have a variety of tissue distortion phenotypes, including excretory defects, pharyngeal ingression, alae defects, and a short and fat body shape, that strongly resemble those of genes encoding ZP proteins. DEX-1 localizes to ZP-containing aECMs in the tissues that show defects in dex-1 mutants. Our studies suggest that DEX-1 is a component of multiple distinct embryonic aECMs that shape developing epithelia, and a potential partner of multiple ZP proteins.

AC. elegansZona Pellucida domain protein functions via its ZPc domain

“…Initial surveys of ZP proteins across phyla suggested that some contain ZPn domains without ZPc domains, and yet are still capable of polymerization (Jovine et al, 2006), while proteins with only ZPc domains were not thought to exist (Bokhove & Jovine, 2018). However, we found that C. elegans does have five predicted ZPc-only proteins (Cohen et al, 2019). In proteins like uromodulin that have polymerizing ZPn domains, ZPc domains are thought to inhibit the ZPn domain (Bokhove & Jovine, 2018).…”

“…The LET-653 ZP domain aligns well with other ZP proteins, but has several unusual features. The ZPn domain lacks a key tyrosine residue thought to be important for ZPn polymerization (Gill et al, 2016;Monne et al, 2008), and the linker region between the ZP subdomains is unusually long, a feature whose impact is unclear (Cohen et al, 2019) was not as strong as that seen with N-terminally tagged transgenes (compare to Figure 2F).…”

“…The LET-653 ZPc domain has additional cysteines compared to some ZP proteins. ZPc domains fall into several families, with 4, 6, or 8 cysteines (Bokhove & Jovine, 2018;Cohen et al, 2019). The LET-653 ZPc domain has 8, as does the Zona Pellucida ZP protein ZP3 (Bokhove & Jovine, 2018;Han et al, 2010).…”

“…Crystal structures reveal that ZPn and ZPc domains independently take on Immunoglobulin (Ig)-like folds held in place by disulfide bonds between paired cysteines (Han et al, 2010;Lin et al, 2011;Saito et al, 2017). ZPn domains typically have four evenly spaced cysteines, while ZPc domains can have four, six, or eight cysteines with some variability in spacing (Bokhove & Jovine, 2018;Cohen, Flatt, Schroeder, & Sundaram, 2019). ZPc modified to promote efficient secretion (Bokhove & Jovine, 2018;Fahrenkamp, Algarra, & Jovine, 2020).…”

“…ZP proteins are important structural components of cuticle and other aECMs in invertebrates, including Caenorhabditis elegans and Drosophila Plaza et al, 2010). C. elegans has an unusually large set of ZP proteins (Cohen et al, 2019), many of which affect shaping of specific epithelia (Forman-Rubinsky, Cohen, & Sundaram, 2017;Gill et al, 2016;Kelley et al, 2015;Low et al, 2019;Muriel et al, 2003;Sapio, Hilliard, Cermola, Favre, & Bazzicalupo, 2005;Sebastiano, Lassandro, & Bazzicalupo, 1991;Vuong-Brender, Suman, & Labouesse, 2017;Yu, Nguyen, Hall, & Chow, 2000). Furthermore, C. elegans is genetically tractable and is transparent, allowing live imaging of the aECM (Corsi, Wightman, & Chalfie, 2015).…”