Probes for HA

Hyaluronan (HA) is a glycosaminoglycan composed of repeating disaccharide units of glucuronic acid and N-acetylglucosamine. HA is an extremely long, unbranched polymer, which often exceeds 106 Da and sometimes reaches 107 Da. A feature that epitomizes HA is its rapid turnover: one-third of the total body HA is turned over daily. The current model of HA catabolism postulates that high-molecular weight HA in the extracellular space is first cleaved into smaller fragments by a hyaluronidase(s) that resides at the cell surface, followed by internalization of fragments and their degradation into monosaccharides in lysosomes. Over the last decade, considerable research has shown that the HYAL family of hyaluronidases plays significant roles in HA catabolism. Nonetheless, the identity of a hyaluronidase responsible for the initial step of HA cleavage on the cell surface remains to be determined, as biochemical and enzymological properties of HYAL proteins are not entirely consistent with those expected of cell surface hyaluronidases. Recent identification of transmembrane 2 (TMEM2) as a cell surface protein that possesses potent hyaluronidase activity suggests that it may be the "missing" cell surface hyaluronidase, and that novel models of HA catabolism should include this protein.

For many years, biologists have focused on the role of Pitx2, expressed on the left side of developing embryos, in governing organ laterality. Here, we identify a different pathway during left-right asymmetry initiated by the right side of the embryo. Surprisingly, this conserved mechanism is orchestrated by the extracellular glycosaminoglycan, hyaluronan(HA) and is independent of Pitx2 on the left. Whereas HA is normally synthesized bilaterally as a simple polysaccharide, we show that covalent modification of HA by the enzyme Tsg6 on the right triggers distinct cell behavior necessary to drive the conserved midgut rotation and to pattern gut vasculature. HA disruption in chicken and Tsg6−/− mice results in failure to initiate midgut rotation and perturbs vascular development predisposing to midgut volvulus. Our study leads us to revise the current symmetry-breaking paradigm in vertebrates and demonstrates how enzymatic modification of HA matrices can execute the blueprint of organ laterality.

Hyaluronan (HA) plays an important role in the development and maintenance of tissues, and its degradation is implicated in many pathologic conditions. We recently reported that HA-binding protein involved in HA depolymerization (HYBID/KIAA1199; encoded by CEMIP) is a key molecule in HA depolymerization, but its developmental and pathologic functions remain elusive. We generated Hybid-deficient mice using the Cre/locus of crossover in P1 (loxP) system and analyzed their phenotypes. Hybid-deficient mice were viable and fertile, but their adult long bones were shorter than those of wild-type animals. Hybid-deficient mice showed lengthening of hypertrophic zone in the growth plate until 4 weeks after birth. There were fewer capillaries and osteoclasts at the chondroosseous junction in the Hybid-deficient mice compared with the wild-type mice. In situ hybridization demonstrated that Hybid was expressed by hypertrophic chondrocytes at the chondroosseous junction. Cultured primary chondrocytes expressed higher levels of Hybid than did osteoblasts or osteoclasts, and the Hybid expression in the chondrocytes was up-regulated after maturation to hypertrophic chondrocytes. High-molecular-weight HA was accumulated in the lengthened hypertrophic zone in Hybid-deficient mice. In addition, high-molecular-weight HA significantly reduced cell growth and tube formation in vascular endothelial growth factor-stimulated or -nonstimulated endothelial cells. HA metabolism by HYBID is involved in endochondral ossification during postnatal development by modulation of angiogenesis and osteoclast recruitment at the chondroosseous junction.