Heterotrimeric G proteins are key regulators of signaling pathways in mammalian cells. Beyond G protein-coupled receptors, the amount and mutual ratio of specific G protein α, β, and γ subunits determine the G protein signaling. However, little is known about mechanisms that regulate the concentration and composition of G protein subunits at the plasma membrane. Here, we show a novel cross-talk between stimulatory and inhibitory G protein α subunits (Gα) that is mediated by G protein βγ dimers and controls the abundance of specific Gα subunits at the plasma membrane. Firstly, we observed in heart tissue from constitutively Gαi2- and Gαi3-deficient mice that the loss of Gαi2 and Gαi3 was accompanied by a slight increase in the protein content of the nontargeted Gαi isoform. Therefore, we analyzed whether overexpression of selected Gα subunits conversely impairs endogenous G protein α and β subunit levels in cardiomyocytes. Integration of overexpressed Gαi2 subunits into heterotrimeric G proteins was verified by co-immunoprecipitation. Adenoviral expression of increasing amounts of Gαi2 led to a reduction of Gαi3 (up to 90 %) and Gαs (up to 75 %) protein levels. Likewise, increasing amounts of adenovirally expressed Gαs resulted in a linear 75 % decrease in both Gαi2 and Gαi3 protein levels. In contrast, overexpression of either Gαi or Gαs isoform did not influence the amount of Gαo and Gαq, both of which are not involved in the regulation of adenylyl cyclase activity. The mRNA expression of the disappearing endogenous Gα subunits was not affected, indicating a posttranslational mechanism. Interestingly, the amount of endogenous G protein βγ dimers was not altered by any Gα overexpression. However, the increase of Gβγ level by adenoviral expression prevented the loss of endogenous Gαs and Gαi3 in Gαi2 overexpressing cardiomyocytes. Thus, our results provide evidence for a novel mechanism cross-regulating adenylyl cyclase-modulating Gαi isoforms and Gαs proteins. The Gα subunits apparently compete for a limited amount of Gβγ dimers, which are required for G protein heterotrimer formation at the plasma membrane.

Dendritic cells (DCs) as professional antigen-presenting cells play an important role in the initiation and modulation of the adaptive immune response. However, their role in the innate immune response against bacterial infections is not completely defined. Here we have analyzed the role of DCs and their impact on the innate anti-bacterial host defense in an experimental infection model of Yersinia enterocolitica (Ye). We used CD11c-diphtheria toxin (DT) mice to deplete DCs prior to severe infection with Ye. DC depletion significantly increased animal survival after Ye infection. The bacterial load in the spleen of DC-depleted mice was significantly lower than that of control mice throughout the infection. DC depletion was accompanied by an increase in the serum levels of CXCL1, G-CSF, IL-1α, and CCL2 and an increase in the numbers of splenic phagocytes. Functionally, splenocytes from DC-depleted mice exhibited an increased bacterial killing capacity compared to splenocytes from control mice. Cellular studies further showed that this was due to an increased production of reactive oxygen species (ROS) by neutrophils. Adoptive transfer of neutrophils from DC-depleted mice into control mice prior to Ye infection reduced the bacterial load to the level of Ye-infected DC-depleted mice, suggesting that the increased number of phagocytes with additional ROS production account for the decreased bacterial load. Furthermore, after incubation with serum from DC-depleted mice splenocytes from control mice increased their bacterial killing capacity, most likely due to enhanced ROS production by neutrophils, indicating that serum factors from DC-depleted mice account for this effect. In summary, we could show that DC depletion triggers phagocyte accumulation in the spleen and enhances their anti-bacterial killing capacity upon bacterial infection.