RT Journal Article
SR Electronic
T1 Carboxyl-Terminal Splicing Enhances Physical Interactions between the Cytoplasmic Tails of Purinergic P2X Receptors
JF Molecular Pharmacology
JO Mol Pharmacol
FD American Society for Pharmacology and Experimental Therapeutics
SP 1588
OP 1598
DO 10.1124/mol.105.019802
VO 69
IS 5
A1 Taka-aki Koshimizu
A1 Karla Kretschmannova
A1 Mu-Lan He
A1 Susumu Ueno
A1 Akito Tanoue
A1 Nobuyuki Yanagihara
A1 Stanko S. Stojilkovic
A1 Gozoh Tsujimoto
YR 2006
UL http://molpharm.aspetjournals.org/content/69/5/1588.abstract
AB Purinergic P2X receptors are ion-conducting channels composed of three subunits, each having two transmembrane domains and intracellular amino (N) and carboxyl (C) termini. Although alternative splicing extensively modifies the C-terminal sequences of P2X subunits, the direct influence of such post-transcriptional modifications on receptor architecture and function remains poorly understood. In this study, we focused on mouse pituitary P2X2 receptors. In this tissue, progressive splicing of the P2X2a C terminus generated two functional subunit variants, P2X2b and P2X2e, which exhibited accelerated desensitization rates and attenuated calcium signals when the receptors were in homomeric states. To measure the intersubunit interaction in living cells, the efficient transfer of bioluminescent resonance energy between luciferase and fluorescent proteins attached to the N- or C-subunit termini of these subunits was used. The constitutive interactions between the full-length C termini of P2X2a receptor were detected by a significant increase in fluorescence/luminescence intensity ratio compared with negative controls. Moreover, interactions between C termini and between C- and N termini of adjacent subunits were significantly enhanced in homomeric and heteromeric receptors containing P2X2b or P2X2e subunits. Finally, deletion of two amino acids at the splicing junction, but not at the C-terminal end of the P2X2b receptor, resulted in the enhancement of channel desensitization and luminescence resonance energy transfer. These results indicate that C-terminal structure plays a critical role in the cytoplasmic intersubunit interactions and suggest that the extent of subunit interactions before ATP application could contribute to the subsequent channel activity and conformation changes associated with agonist-dependent desensitization.