Proteoglycans in Zebrafish Development

Forward genetic screens have identified several zebrafish mutants related to glycosaminoglycan biosynthesis with developmental defects. Many important studies have also used morpholino knockdown of gene expression for functional studies, but off-target effects may be difficult to predict. However, the introduction of the CRISPR-Cas9 genome editing method has now made it possible to generate genetic knockouts overcoming this problem. We know that more heparan sulfate (HS) than chondroitin sulfate (CS) is produced during early zebrafish development while at later stages the dominance of CS is massive. Since a majority of the identified mutants affect production of both HS and CS, their respective roles are sometimes difficult to discern. The two glycosaminoglycans (GAGs) may also have overlapping and redundant functions, such as supporting growth factor signaling, adding to the picture. Craniofacial phenotypes are present in many of the mutants with defective organization and morphogenesis of chondrocytes and bone. Other phenotypes include impaired axon sorting, disturbed heart development, and absent or partly developed pectoral fins. Morpholino studies have also indicated specific functions in angiogenesis, vasculogenesis, and left-right patterning which await confirmation using genetic knockouts. The chapter focuses on HS and CS, but some results also concern hyaluronan and keratan sulfate.