Sonic Hedgehog-induced Proliferation Requires Specific Gα Inhibitory Proteins

Proliferation of cerebellar granular neuronal precursors (CGNPs) is mediated by Sonic Hedgehog (Shh), which activates the Patched and Smoothened (Smo) receptor complex. Although its protein sequence suggests that Smo is a G protein coupled receptor (GPCR), the evidence that this receptor utilizes heterotrimeric G proteins as downstream effectors is controversial. In Drosophila, G alpha(i) is required for Hedgehog (Hh) activity, but the involvement of heterotrimeric G proteins in vertebrate Shh signaling has not yet been established. Here, we show that Shh-induced proliferation of rat CGNPs is enhanced strongly by the expression of the active forms of G alpha(i/o) proteins (G alpha(i1), G alpha(i2), G alpha(i3), and G alpha(o)) but not by members of another class (G alpha(12)) of heterotrimeric G proteins. Additionally, the mRNAs of these different G alpha(i) members display specific expression patterns in the developing cerebellum; only G alpha(i2) and G alpha(i3) are substantially expressed in the outer external granular layer, where CGNPs proliferate. Consistent with this, Shh-induced proliferation of CGNPs is reduced significantly by knockdowns of G alpha(i2) and G alpha(i3) but not by silencing of other members of the G alpha(i/o) class. Finally, our results demonstrate that G alpha(i2) and G alpha(i3) locate to the primary cilium when expressed in CGNP cultures. In summary, we conclude that the proliferative effects of Shh on CGNPs are mediated by the combined activity of G alpha(i2) and G alpha(i3) proteins.