Heterotrimeric G protein a subunits Gα12 and Gα13 are able to stimulate c-Jun N-terminal kinase (JNK) and induce apoptosis via more than one MAP kinase pathways. Gα12 and Gα13 have been reported to stimulate apoptosis signal-regulating kinase (ASK1), one of MAPKK kinases upstream of JNK; the mechanism of this effect has not been studied in detail. Here we report that Gα12 and Gα13 associate with ASK1, as demonstrated by immunoprecipitation assays. Gα12 and Gα13 binding to ASK1 occurs independently of their activation state, as evidenced by ASK1 binding to wild-type Ga subunits both in the presence and in the absence of AlF4-, as well as its similar binding to mutationally activated as compared to wild-type Gα12/Gα13. Using deletion mutants of ASK1, we show that both N- and C-terminal regulatory domains of ASK1 may be involved in the interaction with Gα12/Gα13, while its kinase domain is not required. Coexpression of ASK1 with wild-type Gα13 in COS-7 cells leads to strongly decreased levels of ASK1 protein. In contrast, coexpression of ASK1 with mutationally activated Gα13 increases ASK1 levels as compared to control. Analysis of ASK1 polyubiquitinylation shows that polyubiquitinylated high molecular mass forms of ASK1 are more pronounced in the presence of wild-type Gα13 but are alleviated in the presence of activated Gα13. These results indicate that, in addition to the stimulation of ASK1 kinase activity by Gα12/Gα13 reported previously, ASK1 expression levels can be regulated by these G proteins via control of polyubiquitinylation and following degradation of this kinase.