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Published 20 January 2004. doi:10.1083/jcb.200307101
The Rockefeller University Press, 0021-9525 $8.00
JCB, Volume 164, Number 2, 301-311
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Article

Physical and functional interaction of the active zone proteins, CAST, RIM1, and Bassoon, in neurotransmitter release



Etsuko Takao-Rikitsu1, Sumiko Mochida2, Eiji Inoue1, Maki Deguchi-Tawarada1, Marie Inoue1, Toshihisa Ohtsuka1, and Yoshimi Takai3

1 KAN Research Institute, Kyoto 600-8815, Japan
2 Department of Physiology, Tokyo Medical University, Shinjuku 160-8402, Japan
3 Department of Molecular Biology and Biochemistry, Osaka University Graduate School of Medicine/Faculty of Medicine, Suita 565-0871, Japan

Address correspondence to Toshihisa Ohtsuka, KAN Research Institute, Kyoto Research Park, Shimogyo-ku, Kyoto 600-8815, Japan. Tel.: 81-75-315-7587. Fax: 81-75-325-5130. email: t-ohtsuka{at}kan.gr.jp

We have recently isolated a novel cytomatrix at the active zone (CAZ)–associated protein, CAST, and found it directly binds another CAZ protein RIM1 and indirectly binds Munc13-1 through RIM1; RIM1 and Munc13-1 directly bind to each other and are implicated in priming of synaptic vesicles. Here, we show that all the CAZ proteins thus far known form a large molecular complex in the brain, including CAST, RIM1, Munc13-1, Bassoon, and Piccolo. RIM1 and Bassoon directly bind to the COOH terminus and central region of CAST, respectively, forming a ternary complex. Piccolo, which is structurally related to Bassoon, also binds to the Bassoon-binding region of CAST. Moreover, the microinjected RIM1- or Bassoon-binding region of CAST impairs synaptic transmission in cultured superior cervical ganglion neurons. Furthermore, the CAST-binding domain of RIM1 or Bassoon also impairs synaptic transmission in the cultured neurons. These results indicate that CAST serves as a key component of the CAZ structure and is involved in neurotransmitter release by binding these CAZ proteins.

Key Words: synapse; synaptic vesicle; priming; CAZ

Abbreviations used in this paper: BsnBD, Bassoon-binding domain; CasBD, CAST-binding domain; CAZ, cytomatrix at the active zone; EPSP, excitatory post-synaptic potential; His, hexahistidine; P2, crude membrane; PSD, post-synaptic density; RID, RIM-interacting domain; SCG, superior cervical ganglion; SM3, synaptic membrane 3.


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