Huizing et al. Page 24 A uthor Man uscr ipt A uthor Man uscr ipt Figure 2: Intracellular Free Neu5Ac Metabolism and Associated Genetic Disorders Intracellular free Neu5Ac metabolism comprises three processes: (A) Cytoplasmic free Neu5Ac biosynthesis is initiated with the conversion of UDP-N-acetyl A glucosamine (UDP-GlcNAc) in a few enzymatic steps to Neu5Ac, which is activated in the uthor Man nucleus to CMP-Neu5Ac and then transported back to the cytosol [31, 32, 40]. Cytosolic CMP-Neu5Ac is transported into the Golgi by SLC35A1 [129] where it serves as a substrate for sialyltransferases that sialylate nascent glycans [130]. Cytosolic CMP-Neu5Ac also strongly feedback-inhibits the first committed enzyme of sialic acid biosynthesis, UDP- uscr GlcNAc 2 epimerase, providing negative feedback regulation of de novo cytoplasmic ipt Neu5Ac synthesis [33, 34]. (B) Intralysosomal free Neu5Ac salvage occurs through recycling of glycans (glycoproteins, gangliosides) through endocytosis by the endo-lysosomal system, where lysosomal enzymes degrade the glycans into their individual building block molecules, including individual monosaccharides. Free Neu5Ac is released from glycans by neuraminidase enzymes [84, 86]. Neu5Ac is then transported from the lysosomal lumen into the cytosol by SLC17A5 [1]. A (C) The fate of salvaged free Neu5Ac in the cytoplasm is unclear. A portion may be excreted uthor Man from the cell, recycled in the Neu5Ac biosynthesis pathway for direct synthesis of CMP- Neu5Ac, or degraded/catabolized by N-acetylneuraminate pyruvate lyase (NPL) [38] into ManNAc and pyruvate. The ManNAc generated in the cytoplasm can either directly re-enter the Neu5Ac biosynthesis pathway or can be converted to N-acetylglycosamine (GlcNAc) for uscr entry in the hexosamine pathway [38]. ipt Several rare genetic disorders are associated with these pathways: (1) GNE myopathy (MIM#605820; ~950 reported cases [131]); (2) N-acetylneuraminic acid phosphate synthase Neurosci Lett. Author manuscript; available in PMC 2021 June 11.