Genetics of extracellular matrix remodeling during organ growth using the Caenorhabditis elegans pharynx model.

The organs of animal embryos are typically covered with an extracellular matrix (ECM) that must be carefully remodeled as these organs enlarge during post-embryonic growth; otherwise, their shape and functions may be compromised. We previously described the twisting of the Caenorhabditis elegans pharynx (here called the Twp phenotype) as a quantitative mutant phenotype that worsens as that organ enlarges during growth. Mutations previously known to cause pharyngeal twist affect membrane proteins with large extracellular domains (DIG-1 and SAX-7), as well as a C. elegans septin (UNC-61). Here we show that two novel alleles of the C. elegans papilin gene, mig-6(et4) and mig-6(sa580), can also cause the Twp phenotype. We also show that overexpression of the ADAMTS protease gene mig-17 can suppress the pharyngeal twist in mig-6 mutants and identify several alleles of other ECM-related genes that can cause or influence the Twp phenotype, including alleles of fibulin (fbl-1), perlecan (unc-52), collagens (cle-1, dpy-7), laminins (lam-1, lam-3), one ADAM protease (sup-17), and one ADAMTS protease (adt-1). The Twp phenotype in C. elegans is easily monitored using light microscopy, is quantitative via measurements of the torsion angle, and reveals that ECM components, metalloproteinases, and ECM attachment molecules are important for this organ to retain its correct shape during post-embryonic growth. The Twp phenotype is therefore a promising experimental system to study ECM remodeling and diseases.

C. elegans mig-6 encodes papilin isoforms that affect distinct aspects of DTC migration, and interacts genetically with mig-17 and collagen IV.

The gonad arms of C. elegans hermaphrodites acquire invariant shapes by guided migrations of distal tip cells (DTCs), which occur in three phases that differ in the direction and basement membrane substrata used for movement. We found that mig-6 encodes long (MIG-6L) and short (MIG-6S) isoforms of the extracellular matrix protein papilin, each required for distinct aspects of DTC migration. Both MIG-6 isoforms have a predicted N-terminal papilin cassette, lagrin repeats and C-terminal Kunitz-type serine proteinase inhibitory domains. We show that mutations affecting MIG-6L specifically and cell-autonomously decrease the rate of post-embryonic DTC migration, mimicking a post-embryonic collagen IV deficit. We also show that MIG-6S has two separable functions - one in embryogenesis and one in the second phase of DTC migration. Genetic data suggest that MIG-6S functions in the same pathway as the MIG-17/ADAMTS metalloproteinase for guiding phase 2 DTC migrations, and MIG-17 is abnormally localized in mig-6 class-s mutants. Genetic data also suggest that MIG-6S and non-fibrillar network collagen IV play antagonistic roles to ensure normal phase 2 DTC guidance.

The N-glycolyl form of mouse sialyl Lewis X is recognized by selectins but not by HECA-452 and FH6 antibodies that were raised against human cells.

E-, P- and L-selectins critically function in lymphocyte recirculation and recruiting leukocytes to inflammatory sites. MECA-79 antibody inhibits L-selectin-mediated lymphocyte adhesion in several species and does not require sialic acid in its epitope. Many other antibodies, however, recognize human selectin ligands expressing N-acetylneuraminic acid but not mouse selectin ligands expressing N-glycolylneuraminic acid, suggesting that difference in sialic acid in sialyl Lewis X leads to differential reactivity. We found that HECA-452 and FH6 monoclonal antibodies bind Chinese hamster ovary (CHO) cells expressing N-acetylneuraminyl Lewis X oligosaccharide but not its N-glycolyl form. Moreover, synthetic N-acetylneuraminyl Lewis X oligosaccharide but not its N-glycolyl oligosaccharide inhibited HECA-452 and FH6 binding. By contrast, E-, P- and L-selectin bound to CHO cells regardless of whether they express N-acetyl or N-glycolyl form of sialyl Lewis X, showing that selectins have a broader recognition capacity than HECA-452 and FH-6 anti-sialyl Lewis x antibodies.

UNC-52/perlecan affects gonadal leader cell migrations in C. elegans hermaphrodites through alterations in growth factor signaling.

The unc-52 gene of Claenorhabditis elegans encodes a homologue of the basement membrane heparan sulfate proteoglycan perlecan. Viable alleles reduce the abundance of UNC-52 in late larval stages and increase the frequency of distal tip cell (DTC) migration defects caused by mutations disrupting the UNC-6/netrin guidance system. These unc-52 alleles do not cause circumferential DTC migration defects in an otherwise wild-type genetic background. The effects of unc-52 mutations on DTC migrations are distinct from effects on myofilament organization and can be partially suppressed by mutations in several genes encoding growth factor-like molecules, including EGL-17/FGF, UNC-129/TGF-beta, DBL-1/TGF-beta, and EGL-20/WNT. We propose that UNC-52 serves dual roles in C. elegans larval development in the maintenance of muscle structure and the regulation of growth factor-like signaling pathways.