Chemotaxis mediated by chemokine receptors such as CXCR4 plays a key role in lymphocyte homing and hematopoiesis as well as in breast cancer metastasis. We have demonstrated previously that ␤-arrestin2 functions to attenuate CXCR4-mediated G protein activation and to enhance CXCR4 internalization. Here we show further that the expression of ␤-arrestin2 in both HeLa and human embryonic kidney 293 cells significantly enhances the chemotactic efficacy of stromal cell-derived factor 1␣, the specific agonist of CXCR4, whereas the suppression of ␤-arrestin2 endogenous expression by antisense or RNA-mediated interference technology considerably attenuates stromal cell-derived factor 1␣-induced cell migration. Expression of ␤-arrestin2 also augmented chemokine receptor CCR5-mediated but not epidermal growth factor receptor-mediated chemotaxis, indicating the specific effect of ␤-arrestin2. Further analysis reveals that expression of ␤-arrestin2 strengthened CXCR4-mediated activation of both p38 MAPK and ERK, and the suppression of ␤-arrestin2 expression blocked the activation of two kinases. Interestingly, inhibition of p38 MAPK activation (but not ERK activation) by its inhibitors or by expression of a dominantnegative mutant of p38 MAPK effectively blocked the chemotactic effect of ␤-arrestin2. Expression of a dominant-negative mutant of ASK1 also exerted the similar blocking effect. The results of our study suggest that ␤-arrestin2 can function not only as a regulator of CXCR4 signaling but also as a mediator of stromal cellderived factor 1␣-induced chemotaxis and that this activity probably occurs via the ASK1/p38 MAPK pathway.

Norepinephrine released by the sympathetic nerve terminals regulates the immune system primarily via its stimulation of beta(2)-adrenergic receptor (beta(2)AR), but the underlying molecular mechanisms remain to be elicited. Beta(2)AR, a well-studied G protein-coupled receptor (GPCR), is functionally regulated by beta-arrestin2, which not only causes receptor desensitization and internalization but also serves as a signaling molecule in GPCR signal transduction. Here we show that beta-arrestin2 directly interacts with IkappaBalpha (inhibitor of NF-kappaB, the key molecule in innate and adaptive immunity) and thus prevents the phosphorylation and degradation of IkappaBalpha. Consequently, beta-arrestin2 effectively modulates activation of NF-kappaB and expression of NF-kappaB target genes. Moreover, stimulation of beta(2)AR significantly enhances beta-arrestin2-IkappaBalpha interaction and greatly promotes beta-arrestin2 stabilization of IkappaBalpha, indicating that beta-arrestin2 mediates a crosstalk between beta(2)AR and NF-kappaB signaling pathways. Taken together, the current study may present a novel mechanism for regulation of the immune system by the sympathetic nervous system.

Tumor necrosis factor receptor-associated factor 6 (TRAF6) is critical for mediating Toll-like receptor (TLR)-interleukin 1 receptor (IL-1R) signaling and subsequent activation of NF-kappaB and AP-1, transcriptional activators of innate immunity. Here we show that beta-arrestins, a family of multifunctional proteins, directly interacted with TRAF6 after TLR-IL-1R activation. Formation of the beta-arrestin-TRAF6 complex prevented autoubiquitination of TRAF6 and activation of NF-kappaB and AP-1. Endotoxin-treated beta-arrestin 2-deficient mice had higher expression of proinflammatory cytokines and were more susceptible to endotoxic shock. Thus, beta-arrestins are essential negative regulators of innate immune activation via TLR-IL-1R signaling.