EXPRESSION CLONING OF A HUMAN G(T3) - SYNTHASE G(D3) AND G(T3) ARE SYNTHESIZED BY A SINGLE ENZYME

Gangliosides of the C series such as G(T3) are polysialylated glycosph ingolipids whose synthesis is developmentally regulated. Here we repor t the expression cDNA cloning and characterization of G(T3) synthase t hat adds the second alpha-2,8-sialic acid to G(D3), NeuNAc alpha 2-->8 NeuNAc alpha 2-->3Gal beta 1-->4Glc-Cer, thus forming G(T3), NeuNAc al pha 2-->8NeuNAc alpha 2-->8NeuNAc alpha 2-->3Gal beta 1-->4Glc-->Cer. Unexpectedly, the cloned cDNA was found to be identical to the cDNA th at encodes G(D3) synthase. The newly identified enzyme was therefore n amed G(D3)/G(T3) synthase (G(D3)/G(T3)ST). G(D3)/G(T3)ST synthesized G (T3) most efficiently when G(M3), NeuNAc alpha 2-->3Gal beta 1-->4Glc- ->Cer, was incubated as an acceptor, indicating that G(D3)/G(T3)ST is a polysialyltransferase that can transfer more than one sialic acid re sidue via alpha-2,8 linkage to gangliosides. Moreover, a longer period of incubation of G(D3) with G(D3)/G(T3)ST produced a significant amou nt of G(T3) and higher polysialogangliosides. Among various cell lines expressing G(D3)/G(T3)ST, higher polysialogangliosides including G(T3 ) were detected only in cell lines where the amount of G(D3)/G(T3) mRN A is sufficiently high. The expression of G(D3)/G(T3)ST mRNA among hum an tissues is highly restricted to fetal and adult brains. The G(D3)/G (T3)ST gene was found to be located at chromosome 12, region p12. Take n together, these results indicate that C series polysialogangliosides are synthesized by a ganglioside-specific polysialyltransferase, G(D3 )/G(T3)ST, that is specifically expressed in neural tissues.