Cloning and characterization of the mammalian brain-specific, Mg2+-dependent neutral sphingomyelinase

The enzymatic breakdown of sphingomyelin by sphingomyelinases is considered the major source of the second messenger ceramide, Studies on the contribu tion of the Various described acidic and neutral sphingomyelinases to the s ignaling pool of ceramide have been hampered by the lack of molecular data on the neutral sphingomyelinases (nSMases), We recently identified a mammal ian nSMase, an integral membrane protein with remote similarity to bacteria l sphingomyelinases. However, its ubiquitous expression pattern is in contr ast to previous findings that sphingomyelinase activity is found mainly in brain tissues. By using an improved database search method, combined with p hylogenetic analysis, we identified a second mammalian nSMase (nSMase2) wit h predominant expression in the brain. The sphingomyelinase activity of nSM ase2 has a neutral pH optimum, depends on Mg2+ ions, and is activated by un saturated fatty acids and phosphatidylserine. Immunofluorescence reveals a neuron-specific punctate perinuclear staining, which colocalizes with a Gol gi marker in a number of cell lines, The likely identity of nSMase2 with cc a1, a rat protein involved in contact inhibition of 3Y1 fibroblasts, sugges ts a role for this enzyme in cell cycle arrest, Both mammalian nSMases are members of a superfamily of Mg2+-dependent phosphohydrolases, which also co ntains nucleases, inositol phosphatases, and bacterial toxins.