Activation of β- and α2-adrenergic receptors stimulate tubulin polymerization and promote the association of Gβγ with microtubules in cultured NIH3T3 cells

Abstract

Microtubules (MTs) constitute a crucial part of the cytoskeleton and are essential for cell division and differentiation, cell motility, intracellular transport, and cell morphology. Precise regulation of MT assembly and dynamics is essential for the performance of these functions. Although much progress has been made in identifying and characterizing the cellular factors that regulate MT assembly and dynamics, signaling events in this process is not well understood. Gβγ, an important component of the G protein-coupled receptor (GPCR) signaling pathway, has been shown to promote MT assembly in vitro and in cultured NIH3T3 and PC12 cells. Using the MT depolymerizing agent nocodazole, it has been demonstrated that the association of Gβγ with polymerized tubulin is critical for MT assembly. More recently, Gβγ has been shown to play a key role in NGF-induced neuronal differentiation of PC12 cells through its interaction with tubulin/MTs and modulation of MT assembly. Although NGF is known to exert its effect through tyrosine kinase receptor TrkA, the result suggests a possible involvement of GPCRs in this process. The present study was undertaken to determine whether agonist activation of GPCR utilizes Gβγ to promote MT assembly. We used isoproterenol and UK 14,304, agonists for two different GPCRs (β- and α2-adrenergic receptors, respectively) known to activate Gs and Gi respectively, with an opposing effect on production of cAMP. The results demonstrate that the agonist activation of β- and α2-adrenergic receptors promotes the association of Gβγ with MTs and stimulates MT assembly in NIH3T3 cells. Interestingly, the effects of these two agonists were more prominent when the cellular level of MT assembly was low (30% or less). In contrast to MT assembly, actin polymerization was not affected by isoproterenol or UK 14, 304 indicating that the effects of these agonists are limited to MTs. Thus, it appears that, upon cellular demand, GPCRs may utilize Gβγ to promote MT assembly. Stimulation of MT assembly appears to be a novel function of G protein-mediated signaling.

G protein-coupled receptor-G protein interactions: a single-molecule perspective

“…Conversely, it has been suggested that G proteins can serve as regulators of microtubule assembly and dynamics. In particular, Gα has been shown to inhibit microtubule assembly (363) while Gβγ has been suggested to bind to microtubules and promote their assembly (364,387). Furthermore, adenylyl cyclase 8 was suggested to interact directly with actin (13).…”

Microtubules: From understanding their dynamics to using them as potential therapeutic targets

“…Agonist activation of G protein‐coupled receptor utilizes G beta gamma to promote MT assembly. Agonist activation of beta‐ and alpha‐2‐adrenergic receptors promotes G beta gamma association with MT and stimulates MT assembly (Sierra‐Fonseca, Bracamontes, Saldecke, Das, & Roychowdhury, ).…”

The thrombin receptor (PAR1) is associated with microtubules, mitosis and process formation in glioma cells

“…From a mechanistic point of view, it is important to understand whether the thrombin/PAR1 pathway effects on the cellular morphological and proliferative process are in correlation to MTs dynamics. Previous data indicates several membrane bound receptors interact and affect the MT trafficking system, including tyrosine kinase receptors such as endothelial growth factor receptor (EGFR) [ 20 ], and the β2 adrenergic GPCR [ 21 ]. The PARs are known to be mainly localized at the cell membrane and internalized in vesicles following activation [ 22 ].…”