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Identification and Characterization o f CRIPlb: A Novel CBi Cannabinoid Receptor Interacting ProteinG protein-coupled receptors (GPCRs) transduce extracellular stimuli to intracellular signals through their interaction with heterotrimeric G proteins. Signaling diversity and specificity is imparted primarily through variations o f G protein subunits. Protein-protein interactions between intracellular accessory proteins and GPCRs also modify signaling by altering receptor activity or signaling pathways. The ability of intracellular proteins to interact with the CBi cannabinoid receptor was investigated to determine whether particular signaling properties of CBi resulted from interaction with specific CBi interacting proteins. A novel protein named C RIPlb was discovered to interact with the C-terminal tail o f CB). The interaction between CRIPlb and CBi was characterized using the yeast two-hybrid assay. Functional consequences of the CRIPlb- CB| interaction were investigated by examining protein localization by confocal microscopy and measuring CBi mediated N-type Ca2+ channel activity in the presence of CRIPlb by whole-cell patch clamp recordings. The yeast two-hybrid assay indicated that the last nine amino acids of the CBi C-terminal tail were required for interaction with CRIPlb. Heterologous expression of C RIPlb and CBi in HEK 293 cells did not reveal evidence of colocalization, nor was CBi able to significantly traffic C RIPlb to the plasma membrane. However, CRIPlb and CBi were found to colocalize in superior cervical ganglion (SCG) neurons. Whole-cell voltage-clamp recordings of N-type Ca2+ channels in SCG neurons indicated that CRIPlb had no effect on agonist- or inverse agonist-induced modulation of Ca2+ current by CBi. Furthermore, the level of CBi constitutive activity was not significantly altered by CRIPlb. The high affinity of CBi for G proteins, as demonstrated by the ability of CBi to sequester G proteins from other Gi/0 coupled receptors, was unaffected by expression of CRIP lb. These results provide evidence that CRIPlb is a novel CBi accessory protein that interacts with the C-terminal tail of CBi. While CRIPlb and CBi can apparently interact in a neuronal expression system, the ability of CRIPlb to modify CBi signaling was not detected in any of the pathways investigated. Thus, the distinctive signaling properties of CB|, such as constitutive activity and G protein sequestration do not originate from nor are modified by CRIPlb.