Identification and characterization of a synaptojanin 2 splice isoform predominantly expressed in nerve terminals

We have previously identified synaptojanin 1, a phosphoinositide phosphatas e predominantly expressed in the nervous system, and synaptojanin 2, a broa dly expressed isoform. Synaptojanin 1 is concentrated in nerve terminals, w here it has been implicated in synaptic vesicle recycling and actin functio n. Synaptojanin 2A is targeted to mitochondria via a PDZ domain-mediated in teraction. We have now characterized an alternatively spliced form of synap tojanin 2 that shares several properties with synaptojanin 1. This isoform, synaptojanin 2B, undergoes further alternative splicing to generate synapt ojanin 2B1 and 2B2. Both amphiphysin and endophilin, two partners synaptoja nin 1, bind synaptojanin 2B2, whereas only amphiphysin binds synaptojanin 2 B1. Sequence similar to the endophilin-binding site in synaptojanin 1 is pr esent only in synaptojanin 2B2, and this sequence was capable of affinity p urifying endophilin from rat brain. The Sac1 domain of synaptojanin 2 exhib ited phosphoinositide phosphatase activity very similar to that of the Sac1 domain of synaptojanin 1. Site-directed mutagenesis further illustrated it s functional similarity to the catalytic domain of Sac1 proteins. Antibodie s raised against the synaptojanin 2B-specific carboxyl-terminal region iden tified a 160-kDa protein in brain and testis. Immunofluorescence showed tha t synaptojanin 2B is localized at nerve terminals in brain and at the sperm atid manchette in testis. Active Rac1 GTPase affects the intracellular loca lization of synaptojanin 2, but not of synaptojanin 1. These results sugges t that synaptojanin 2B has a partially overlapping function with synaptojan in 1 in nerve terminals, with additional roles in neurons and other cells i ncluding dspermatids.