A novel calmodulin-binding protein, belonging to the WD-repeat family, is localized in dendrites of a subset of CNS neurons.

A rat brain synaptosomal protein of 110,000 M(r) present in a fraction highly enriched in adenylyl cyclase activity was microsequenced (Castets, F., G. Baillat, S. Mirzoeva, K. Mabrouk, J. Garin, J. d'Alayer, and A. Monneron. 1994. Biochemistry. 33:5063-5069). Peptide sequences were used to clone a cDNA encoding a novel, 780-amino acid protein named striatin. Striatin is a member of the WD-repeat family (Neer, E.J., C.J. Schmidt, R. Nambudripad, and T.F. Smith. 1994. Nature (Lond.). 371:297-300), the first one known to bind calmodulin (CaM) in the presence of Ca++. Subcellular fractionation shows that striatin is a membrane-associated, Lubrol-soluble protein. As analyzed by Northern blots, in situ hybridization, and immunocytochemistry, striatin is localized in the central nervous system, where it is confined to a subset of neurons, many of which are associated with the motor system. In particular, striatin is conspicuous in the dorsal part of the striatum, as well as in motoneurons. Furthermore, striatin is essentially found in dendrites, but not in axons, and is most abundant in dendritic spines. We propose that striatin interacts, through its WD-repeat domain and in a CaM/Ca(++)-dependent manner, with one or several members of a surrounding cluster of molecules engaged in a Ca(++)-signaling pathway specific to excitatory synapses.

Molecular cloning and characterization of phocein, a protein found from the Golgi complex to dendritic spines.

Phocein is a widely expressed, highly conserved intracellular protein of 225 amino acids, the sequence of which has limited homology to the sigma subunits from clathrin adaptor complexes and contains an additional stretch bearing a putative SH3-binding domain. This sequence is evolutionarily very conserved (80% identity between Drosophila melanogaster and human). Phocein was discovered by a yeast two-hybrid screen using striatin as a bait. Striatin, SG2NA, and zinedin, the three mammalian members of the striatin family, are multimodular, WD-repeat, and calmodulin-binding proteins. The interaction of phocein with striatin, SG2NA, and zinedin was validated in vitro by coimmunoprecipitation and pull-down experiments. Fractionation of brain and HeLa cells showed that phocein is associated with membranes, as well as present in the cytosol where it behaves as a protein complex. The molecular interaction between SG2NA and phocein was confirmed by their in vivo colocalization, as observed in HeLa cells where antibodies directed against either phocein or SG2NA immunostained the Golgi complex. A 2-min brefeldin A treatment of HeLa cells induced the redistribution of both proteins. Immunocytochemical studies of adult rat brain sections showed that phocein reactivity, present in many types of neurons, is strictly somato-dendritic and extends down to spines, just as do striatin and SG2NA.

Zinedin, SG2NA, and striatin are calmodulin-binding, WD repeat proteins principally expressed in the brain.

Striatin is an intracellular protein characterized by four protein-protein interaction domains, a caveolin-binding motif, a coiled-coil structure, a calmodulin-binding domain, and a WD repeat domain, suggesting that it is a signaling or a scaffold protein. Down-regulation of striatin, which is expressed in a few subsets of neurons, impairs the growth of dendrites as well as rat locomotor activity (Bartoli, M., Ternaux, J. P., Forni, C., Portalier, P., Salin, P., Amalric, M., and Monneron, A. (1999) J. Neurobiol. 40, 234-243). Zinedin, a "novel" protein described here, and SG2NA share with striatin identical protein-protein interaction domains and the same overall domain structure. A phylogenetic analysis supports the hypothesis that they constitute a multigenic family deriving from an ancestral gene. DNA probes and antibodies raised against specific domains of each protein showed that zinedin is mainly expressed in the central nervous system, whereas SG2NA, of more widespread occurrence, is mainly expressed in the brain and muscle. All three proteins are both cytosolic and membrane-bound. All three bind calmodulin in the presence of Ca(2+). In rat brain, SG2NA and striatin are generally not found in the same neurons. Both localize to the soma and dendrites, suggesting that they share a similar type of addressing and closely related functions.

Cloning of human striatin cDNA (STRN), gene mapping to 2p22-p21, and preferential expression in brain.

Rat striatin, a recently discovered calmodulin-binding protein belonging to the WD repeat family, is expressed in neurons, mostly in the striatum and motor and olfactory systems. Striatin is localized in the somato-dendritic compartment of neurons, mainly in the spines. It may play a role in dendritic Ca2+ signaling. Here we report the cloning and sequencing of human striatin cDNA (HGMW-approved symbol STRN), the localization of the gene to chromosome 2p22-p21, and its preferential expression in brain. The human cDNA sequence is predicted to encode a 780-amino-acid protein possessing eight WD repeats. Striatin is highly conserved between rat and human with 96% identity and 98% similarity at the amino acid level. Since the Caenorabditis elegans genome also contains a closely related striatin coding sequence, the function of striatin is likely to be well conserved.

Gene structure, expression pattern, and biological activity of mouse killer cell activating receptor-associated protein (KARAP)/DAP-12.

Natural killer cell and T cell subsets express at their cell surface a repertoire of receptors for MHC class I molecules, the natural killer cell receptors (NKRs). NKRs are characterized by the existence of inhibitory and activating isoforms, which are encoded by highly homologous but separate genes present in the same locus. Inhibitory isoforms express an intracytoplasmic immunoreceptor tyrosine-based inhibition motif, whereas activating isoforms lack any immunoreceptor tyrosine-based inhibition motif but harbor a charged amino acid residue in their transmembrane domain. We previously characterized KARAP (killer cell activating receptor-associated protein), a novel disulfide-linked tyrosine-phosphorylated dimer that selectively associates with the activating NKR isoforms. We report here the identification of the mouse KARAP gene, its localization on chromosome 7 and its genomic organization in five exons. Point mutation and transfection studies revealed that KARAP is a novel signaling transmembrane subunit whose transduction function depends on the integrity of an intracytoplasmic immunoreceptor tyrosine-based activation motif. In contrast to previous members of the immunoreceptor tyrosine-based activation motif polypeptide family, KARAP is ubiquitously expressed on hematopoietic and nonhematopoietic cells, suggesting its association with a broad range of activating receptors in a variety of tissues.