Competitive interaction of seven in absentia homolog-1A and Ca2+/calmodulin with the cytoplasmic tail of group 1 metabotropic glutamate receptors.
BACKGROUND
Group 1 metabotropic glutamate receptors (mGluR1 and mGluR5) are coupled to inositol trisphosphate/Ca2+ signaling via G proteins and play an important role in excitatory synaptic transmission. To explore the regulation of group 1 mGluR function, we applied the yeast two-hybrid system using the intracellular carboxy-terminal domain of group 1 mGluRs (group 1 ct-mGluRs) and attempted to identify novel protein-protein interactions of group 1 mGluRs.


RESULTS
The two-hybrid screening revealed a specific interaction between group 1 ct-mGluRs and Siah-1A, the mammalian homolog of Drosophila seven in absentia which is involved in photoreceptor cell differentiation via the ubiquitin/proteasome-dependent mechanism. This interaction occurs within a homologous 27-28 amino acid stretch within group 1 ct-mGluRs and requires the latter two-thirds of Siah-1A. Following coexpression in COS-7 cells, myc-tagged Siah-1A was coimmunoprecipitated with the flag-tagged ct-mGluR1 by anti-flag antibody. Furthermore, in vitro binding revealed that Siah-1A and Ca2+/calmodulin (CaM) binding sites overlap, such that Siah-1A binding is competitively inhibited by CaM in a Ca2+-dependent manner.


CONCLUSIONS
The results demonstrate a direct interaction between group 1 mGluRs and Siah-1A and suggest a novel modulatory mechanism mediated by a competitive interaction between Ca2+/CaM and Siah-1A.
Ishikawa
K
K
Department of Biological Sciences, Kyoto University Faculty of Medicine, Yoshida, Sakyo-ku, Kyoto 606-8501, Japan.
Nash
S R
SR
Nishimune
A
A
Neki
A
A
Kaneko
S
S
Nakanishi
S
S
D000595:Amino Acid Sequence; D000818:Animals; D001667:Binding, Competitive / Q000187:drug effects; D001921:Brain / Q000378:metabolism; D019556:COS Cells; D002118:Calcium / Q000378:metabolism / Q000494:pharmacology; D002147:Calmodulin / Q000378:metabolism*; D004533:Egtazic Acid / Q000494:pharmacology; D008969:Molecular Sequence Data; D009154:Mutation; D009687:Nuclear Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011233:Precipitin Tests; D011485:Protein Binding / Q000187:drug effects; D051381:Rats; D018094:Receptors, Metabotropic Glutamate / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D044767:Ubiquitin-Protein Ligases; D015003:Yeasts / Q000235:genetics / Q000378:metabolism; D000098563:Seven in Absentia Proteins
D002147:Calmodulin; D009687:Nuclear Proteins; D018094:Receptors, Metabotropic Glutamate; D011993:Recombinant Fusion Proteins; C104077:metabotropic glutamate receptor type 1; D004533:Egtazic Acid; D044767:Ubiquitin-Protein Ligases; D000098563:Seven in Absentia Proteins; D002118:Calcium