Sulphated proteoglycan is required for collecting duct growth and branching but not nephron formation during kidney development

Kidney epithelia have separate origins; collecting ducts develop by ureteric bud growth and arborisation, nephrons by induced mesenchyme-epithelium transition. Both express sulphated glycosaminoglycans (GAGs) which are strikingly upregulated during nephron differentiation. However, sodium chlorate, an inhibitor of GAG sulphation, and the GAG-degrading enzymes heparitinase plus chondroitinase, did not prevent nephron development. In contrast, ureteric bud growth and branching were reversibly inhibited by the above reagents, the inhibition correlating quantitatively with sulphated GAG deprivation caused by a range of chlorate concentrations. Growth and branching could be independently restored during GAG deprivation by hepatocyte growth factor and phorbol-12-myristate acetate (PMA) respectively. Together these signalling effectors stimulated both branch initiation and growth. Thus growth and morphogenesis of ureteric bud involve distinct signalling pathways both regulated by GAGs.

Biosynthesis of heparin. Relationship between the polymerization and sulphation processes

Incubation of a mouse mastocytoma microsomal fraction with UDP-[3H]GlcA and UDP-GlcNAc yielded proteoglycans containing non-sulphated polysaccharide chains. Similar incubations performed in the presence of sulphate donor 3′-phosphoadenosine 5′-phosphosulphate (PAPS) produced both sulphated and non-sulphated proteoglycans, which were separated by chromatography on DEAE-cellulose Analysis by gel chromatography of single polysaccharide chains, released from the proteoglycans by alkali treatment, showed that the non-sulphated chains produced during incubation for 5 min or 25 min, either in the absence or in the presence of PAPS, were of fairly small molecular size, with an average peak Mr of approx. 10 x 10(3)-15 x 10(3). In contrast, the sulphated chains exceeded Mr 100 x 10(3) Pulse-chase experiments suggested that sulphated chains were capable of further elongation. These results indicate that sulphation promotes, by so far unknown mechanisms, further chain elongation. Sulphated proteoglycan (retarded on DEAE-cellulose chromatography) isolated after similar incubation of the microsomal fraction for 1 min only was found to contain a mixture of sulphated and virtually non-sulphated polysaccharide chains. However, when [35S]PAPS was included in the incubations, some 35S was found to be associated, essentially as N-sulphate groups, also with the latter type of chains, preferentially the high-Mr fraction. These results are interpreted in terms of a biosynthetic model by which the heparin proteoglycan is generated through transient interactions of macromolecular intermediates with distinctly separate complexes of membranebound enzymes.