Abstract
At synaptic junctions, pre- and postsynaptic membranes are connected by cell adhesion and have distinct structures for specialized functions. The presynaptic membranes have a machinery for fast neurotransmitter release, and the postsynaptic membranes have clusters of neurotransmitter receptors. The molecular mechanism of the assembly of synaptic junctions is not yet clear. Pioneering studies identified postsynaptic density (PSD)-95/SAP90 as a prototypic synaptic scaffolding protein to maintain the structure of synaptic junctions. PSD-95/SAP90 belongs to a family of membrane-associated guanylate kinases and binds N-methyl-D-aspartate receptors, potassium channels, and neuroligins through the PDZ domains and GKAP/SAPAP/DAP through the guanylate kinase (GK) domain. We performed here a yeast two-hybrid screening for SAPAP-interacting molecules and identified a novel protein that has an inverse structure of membrane-associated guanylate kinases with an NH2-terminal GK-like domain followed by two WW and five PDZ domains. It binds SAPAP through the GK-like domain and NMDA receptors and neuroligins through the PDZ domains. We named this protein S-SCAM (synaptic scaffolding molecule) because S-SCAM may assemble receptors and cell adhesion proteins at synaptic junctions.
Highlights
Effective neurotransmission requires the precise localization of the receptors for neurotransmitters opposite to the presynaptic active zone
The first and second PDZ domains of postsynaptic density (PSD)-95/SAP90 interact with NMDA receptors and Shaker type potassium channels [5, 6]
Molecules interacting with the guanylate kinase (GK) domain of PSD-95/SAP90 have been identified by three groups and named GKAP/SAPAP/DAP [25,26,27,28]
Summary
DNA Constructs—Various constructs were prepared using pBTM116, pVP16-3, pCMV5, pGex5X-3 (Amersham Pharmacia Biotech), pGexKG, and pClneo Myc. pClneo Myc was constructed by ligating the oligonucleotides ctagaccccccaacatggagcagaagcttatcagcgaggaggacctgctcgagg/aattcctcgagcaggtcctcctcgctgataagcttctgctccatgttggggggt into NheI/EcoRI sites of pClneo (Promega). pBTM116 SAPAP1–2 contains residues 397–568 of SAPAP1. DNA Constructs—Various constructs were prepared using pBTM116, pVP16-3, pCMV5, pGex5X-3 (Amersham Pharmacia Biotech), pGexKG, and pClneo Myc. pClneo Myc was constructed by ligating the oligonucleotides ctagaccccccaacatggagcagaagcttatcagcgaggaggacctgctcgagg/aattcctcgagcaggtcctcctcgctgataagcttctgctccatgttggggggt into NheI/EcoRI sites of pClneo (Promega). PBTM116 SAPAP1–2 contains residues 397–568 of SAPAP1.
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