The SH3p4/Sh3p8/SH3p13 protein family: Binding partners for synaptojanin and dynamin via a Grb2-like Src homology 3 domain

Abstract

The GTPase dynamin I and the inositol 5-phosphatase synaptojanin are nerve terminal proteins implicated in synaptic vesicle recycling. Both proteins contain COOHterminal proline-rich domains that can interact with a variety of Src homology 3 (SH3) domains. A major physiological binding partner for dynamin I and synaptojanin in the nervous system is amphiphysin I, an SH3 domain-containing protein also concentrated in nerve terminals. We have used the proline-rich tail of synaptojanin to screen a rat brain library by the two-hybrid method to identify additional interacting partners of synaptojanin. Three related proteins containing SH3 domains that are closely related to the SH3 domains of Grb2 were isolated: SH3p4, SH3p8, and SH3p13. Further biochemical studies demonstrated that the SH3p4͞8͞13 proteins bind to both synaptojanin and dynamin I. The SH3p4͞8͞13 transcripts are differentially expressed in tissues: SH3p4 mRNA was detected only in brain, SH3p13 mRNA was present in brain and testis, and the SH3p8 transcript was detected at similar levels in multiple tissues. Members of the SH3p4͞8͞13 protein family were found to be concentrated in nerve terminals, and pools of synaptojanin and dynamin I were coprecipitated from brain extracts with antibodies recognizing SH3p4͞8͞13. These findings underscore the important role of SH3-mediated interactions in synaptic vesicle recycling.

Two synaptojanin 1 isoforms are recruited to clathrin-coated pits at different stages

“…Endophilin is thought to be a particularly important interactor of SJ1 and to play a major role in its recruitment to sites of endocytosis (2,15,19,20). In both flies and worms, mutations of endophilin and synaptojanin have similar phenotypes, and loss of endophilin results in destabilization and mislocalization of synaptojanin (21)(22)(23)(24).…”

“…Most SH3 domain-containing proteins that bind SJ1, including endophilin, also interact with dynamin (15,17,18). This biochemical link to dynamin, together with the accumulation of coated vesicles at synapses of SJ1 knockout mice, has led to the hypothesis that the main function of SJ1-mediated hydrolysis of PI(4,5)P 2 is to facilitate clathrin uncoating after fission (3,5,28).…”

“…The portion of the C-terminal region common to both SJ1-145 and SJ1-170 interacts with the SH3 domains of a variety of proteins implicated in endocytosis and signaling: endophilin, amphiphysin, syndapin/pacsin, intersectin, and many others (10,(14)(15)(16)(17)(18). Endophilin is thought to be a particularly important interactor of SJ1 and to play a major role in its recruitment to sites of endocytosis (2,15,19,20).…”

“…The latter has curvature-generating and -sensing properties and may participate in the generation and maturation of endocytic bud necks (25)(26)(27). Mammalian cells express three isoforms of endophilin, with endophilin 2 (Endo2) being the predominant isoform in nonneuronal cells (15).…”

Endophilin isoforms have distinct characteristics in interactions with N-type Ca2+ channels and dynamin I

“…Studies have shown that three types of endophilin exist in rats: endophilin I is expressed only in brain, endophilin II in multiple tissues, and endophilin III mainly in brain, testis and thymus [1][2][3] . Endophilins I and II also form dimers through a coiled-coil domain [4][5][6][7] .…”