The EXT proteins and HS biosynthesis. (a) Schematic representations of the EXT1 and EXT2 putative tumor suppressor proteins (not drawn to scale). These endoplasmic reticulum–localized (ER-localized) type II transmembrane proteins have an N-terminal cytoplasmic tail, a single transmembrane domain, a stem region, and a long C-terminal lumenal tail. Amino acids (a.a.) mutated in HME patients are labeled and indicated by big stars, amino acids mutated in HS-deficient CHO cells (27) are indicated by small yellow stars, and N-linked glycosylation sites are represented by a pink Y. The proposed D-glucuronic acid glycosyltransferase (GlcA-TII) catalytic domain (27) is shaded in light blue. (b) A schematic representation of the HS biosynthesis pathway (recently reviewed in ref. 43). A tetrasaccharide unit, common to both HS and chondroitin sulfate (CS), is synthesized by sequential additions of xylose (Xyl), two galactose (Gal) residues, and a D-glucuronic acid (GlcA) residue, covalently linked to a serine residue on the HS proteoglycan (HSPG) core protein. HS biosynthesis is specifically initiated by the addition of an N-acetylglucosamine residue (GlcNAc), which is carried out by the glycosyltransferase (GlcNAc-TI) encoded by the EXTL2 gene. Next, the HS-polymerase (HS-Pol), a Golgi-localized hetero-oligomeric complex of which EXT1 and EXT2 are key components, elongates the nascent chain by adding alternating GlcA and GlcNAc residues.