beta-Arrestin 1 and 2 similarly influence mu-opioid receptor mobility and distinctly modulate adenylyl cyclase activity
Abstrakt
beta-Arrestins are known to play a crucial role in GPCR-mediated transmembrane signaling processes. However, there are still many unanswered questions, especially those concerning the presumed similarities and differences of beta-arrestin isoforms.
Here, we examined the roles of beta-arrestin 1 and beta-arrestin 2 at different levels of mu-opioid receptor (MOR)-regulated signaling, including MOR mobility, internalization of MORs, and adenylyl cyclase (AC) activity. For this purpose, naive HEK293 cells or HEK293 cells stably expressing YFP-tagged MOR were transfected with appropriate siRNAs to block in a specific way the expression of beta-arrestin 1 or beta-arrestin 2.
We did not find any significant differences in the ability of beta-arrestin isoforms to influence the lateral mobility of MORs in the plasma membrane. Using FRAP and line-scan FCS, we observed that knockdown of both beta-arrestins similarly increased MOR lateral mobility and diminished the ability of DAMGO and endomorphin-2, respectively, to enhance and slow down receptor diffusion kinetics.
However, beta-arrestin 1 and beta-arrestin 2 diversely affected the process of agonist-induced MOR endocytosis and exhibited distinct modulatory effects on AC function. Knockdown of beta-arrestin 1, in contrast to beta-arrestin 2, more effectively suppressed forskolin-stimulated AC activity and prevented the ability of activated-MORs to inhibit the enzyme activity.
Moreover, we have demonstrated for the first time that beta-arrestin 1, and partially beta-arrestin 2, may somehow interact with AC and that this interaction is strongly supported by the enzyme activation. These data provide new insights into the functioning of beta-arrestin isoforms and their distinct roles in GPCR-mediated signaling.