No abstract

Mammalian casein kinases I (CKI) belong to a family of serine/threonine protein kinases involved in diverse cellular processes including cell cycle progression, membrane trafficking, circadian rhythms, and Wnt signaling. Here we show that CKI␣ co-purifies with centaurin-␣ 1 in brain and that they interact in vitro and form a complex in cells. In addition, we show that the association is direct and occurs through the kinase domain of CKI within a loop comprising residues 217-233. These residues are well conserved in all members of the CKI family, and we show that centaurin-␣ 1 associates in vitro with all mammalian CKI isoforms. To date, CKI␣ represents the first protein partner identified for centaurin-␣ 1 . However, our data suggest that centaurin-␣ 1 is not a substrate for CKI␣ and has no effect on CKI␣ activity. Centaurin-␣ 1 has been identified as a phosphatidylinositol 3,4,5-trisphosphate-binding protein. Centaurin-␣ 1 contains a cysteine-rich domain that is shared by members of a newly identified family of ADPribosylation factor guanosine trisphosphatase-activating proteins. These proteins are involved in membrane trafficking and actin cytoskeleton rearrangement, thus supporting a role for CKI␣ in these biological events.

Casein kinases I (CKI) are serine/threonine protein kinases widely expressed in a range of eukaryotes including yeast, mammals and plants. They have been shown to play a role in diverse physiological events including membrane trafficking. CKIa is associated with synaptic vesicles and phosphorylates some synaptic vesicle associated proteins including SV2. In this report, we show that syntaxin-1A is phosphorylated in vitro by CKI on Thr21. Casein kinase II (CKII) has been shown previously to phosphorylate syntaxin-1A in vitro and we have identified Ser14 as the CKII phosphorylation site, which is known to be phosphorylated in vivo. As syntaxin-1A plays a key role in the regulation of neurotransmitter release by forming part of the SNARE (soluble N-ethylmaleimide-sensitive factor attachment protein receptor) complex, we propose that CKI may play a role in synaptic vesicle exocytosis.Keywords: CKI; CKII; syntaxin-1A; trafficking.Casein kinase I (CKI) belongs to a family of serine/ threonine protein kinases with seven isoforms identified in mammals (CKI a, b, d, e, c1, c2, and c3; reviewed in [1]). The kinase domain is highly conserved between members of the CKI family but unique N-and C-terminal tails characterize each isoform. In yeast, the functions of CKI have been much more extensively studied compared to their mammalian counterparts. Recently, many reports have linked yeast CKIs to cytokinesis and vesicle trafficking especially in endocytosis [2][3][4][5][6][7][8][9]. Mammalian CKIs appear to have similar functions and also have been involved in DNA repair, circadian rhythms, and wnt signaling. Like their yeast counterparts, CKIcs have been implicated in cytokinesis and in membrane trafficking [10]. CKIa interacts with and phosphorylates the clathrin adapter that is involved in endocytosis. CKIa has been found to colocalize in neurones with synaptic vesicle markers and to phosphorylate some synaptic vesicle associated proteins including SV2 [12]. More importantly, the phosphorylation of SV2 by CKI modulates its ability to interact with synaptotagmin [13]. SV2 plays a role in neurotransmitter release suggesting a role for CKI in this biological process. We have recently identified centaurin-a 1 , a protein shown to associate with presynaptic vesicular structures [14], as a novel CKI partner [15].In this report, we have identified syntaxin-1A as a novel substrate for CKI, which further supports a role for CKI in membrane trafficking. Indeed, the involvement of syntaxin-1A in neurotransmitter release is well documented (reviewed in [16,17]). Regulated neurotransmitter secretion is the key step in synaptic transmission and is the basis of intercellular communication in the nervous system. Synaptic vesicle exocytosis is regulated by Ca 2+ and by a large number of proteins (reviewed in [17,18]). Syntaxin-1A is associated with the presynaptic membrane and associates with the plasma membrane protein SNAP-25 and the synaptic vesicle protein synaptobrevin to form a ÔSNARE complexÕ. Assembly of this complex is necessary ...

No abstract