GnRH Neuron-Specific Ablation of Gαq/11 Results in Only Partial Inactivation of the Neuroendocrine-Reproductive Axis in Both Male and Female Mice: In Vivo Evidence for Kiss1r-Coupled Gαq/11-Independent GnRH Secretion.

The gonadotropin-releasing hormone (GnRH) is the master regulator of fertility and kisspeptin (KP) is a potent trigger of GnRH secretion from GnRH neurons. KP signals via KISS1R, a Gαq/11-coupled receptor, and mice bearing a global deletion of Kiss1r (Kiss1r(-/-)) or a GnRH neuron-specific deletion of Kiss1r (Kiss1r(d/d)) display hypogonadotropic hypogonadism and infertility. KISS1R also signals via ß-arrestin, and in mice lacking ß-arrestin-1 or -2, KP-triggered GnRH secretion is significantly diminished. Based on these findings, we hypothesized that ablation of Gαq/11 in GnRH neurons would diminish but not completely block KP-triggered GnRH secretion and that Gαq/11-independent GnRH secretion would be sufficient to maintain fertility. To test this, Gnaq (encodes Gαq) was selectively inactivated in the GnRH neurons of global Gna11 (encodes Gα11)-null mice by crossing Gnrh-Cre and Gnaq(fl/fl);Gna11(-/-) mice. Experimental Gnaq(fl/fl);Gna11(-/-);Gnrh-Cre (Gnaq(d/d)) and control Gnaq(fl/fl);Gna11(-/-) (Gnaq(fl/fl)) littermate mice were generated and subjected to reproductive profiling. This process revealed that testicular development and spermatogenesis, preputial separation, and anogenital distance in males and day of vaginal opening and of first estrus in females were significantly less affected in Gnaq(d/d) mice than in previously characterized Kiss1r(-/-) or Kiss1r(d/d) mice. Additionally, Gnaq(d/d) males were subfertile, and although Gnaq(d/d) females did not ovulate spontaneously, they responded efficiently to a single dose of gonadotropins. Finally, KP stimulation triggered a significant increase in gonadotropins and testosterone levels in Gnaq(d/d) mice. We therefore conclude that the milder reproductive phenotypes and maintained responsiveness to KP and gonadotropins reflect Gαq/11-independent GnRH secretion and activation of the neuroendocrine-reproductive axis in Gnaq(d/d) mice. SIGNIFICANCE STATEMENT: The gonadotropin-releasing hormone (GnRH) is the master regulator of fertility. Over the last decade, several studies have established that the KISS1 receptor, KISS1R, is a potent trigger of GnRH secretion and inactivation of KISS1R on the GnRH neuron results in infertility. While KISS1R is best understood as a Gαq/11-coupled receptor, we previously demonstrated that it could couple to and signal via non-Gαq/11-coupled pathways. The present study confirms these findings and, more importantly, while it establishes Gαq/11-coupled signaling as a major conduit of GnRH secretion, it also uncovers a significant role for non-Gαq/11-coupled signaling in potentiating reproductive development and function. This study further suggests that by augmenting signaling via these pathways, GnRH secretion can be enhanced to treat some forms of infertility.

A Galpha subunit controls zoospore motility and virulence in the potato late blight pathogen Phytophthora infestans.

The heterotrimeric G-protein pathway is a ubiquitous eukaryotic signalling module that is known to regulate growth and differentiation in many plant pathogens. We previously identified Pigpa1, a gene encoding a G-protein alpha subunit from the potato late blight pathogen Phytophthora infestans. P. infestans belongs to the class oomycetes, a group of organisms in which signal transduction processes have not yet been studied at the molecular level. To elucidate the function of Pigpa1, PiGPA1-deficient mutants were obtained by homology-dependent gene silencing. The Pigpa1-silenced mutants produced zoospores that turned six to eight times more frequently, causing them to swim only short distances compared with wild type. Attraction to the surface, a phenomenon known as negative geotaxis, was impaired in the mutant zoospores, as well as autoaggregation and chemotaxis towards glutamic and aspartic acid. Zoospore production was reduced by 20-45% in different Pigpa1-silenced mutants. Transformants expressing constitutively active forms of PiGPA1, containing amino acid substitutions (R177H and Q203L), showed no obvious phenotypic differences from the wild-type strain. Infection efficiencies on potato leaves ranged from 3% to 14% in the Pigpa1-silenced mutants, compared with 77% in wild type, showing that virulence is severely impaired. The results prove that PiGPA1 is crucial for zoospore motility and for pathogenicity in an important oomycete plant pathogen.

A Phytophthora infestans G-protein beta subunit is involved in sporangium formation.

The heterotrimeric G-protein pathway regulates cellular responses to a wide range of extracellular signals in virtually all eukaryotes. It also controls various developmental processes in the oomycete plant pathogen Phytophthora infestans, as was concluded from previous studies on the role of the G-protein alpha-subunit PiGPA1 in this organism. The expression of the P. infestans G-protein beta-subunit gene Pigpb1 was induced in nutrient-starved mycelium before the onset of sporangium formation. The gene was hardly expressed in mycelium incubated in rich growth medium. The introduction of additional copies of Pigpb1 into the genome led to silencing of the gene and resulted in transformants deficient in PiGPB1. These Pigpb1-silenced mutants formed very few asexual spores (sporangia) when cultured in rye sucrose medium and produced a denser mat of aerial mycelium than the wild type. Partially Pigpb1-silenced mutants showed intermediate phenotypes with regard to sporulation, and a relatively large number of their sporangia were malformed. The results show that PiGPB1 is important for vegetative growth and sporulation and, therefore, for the pathogenicity of this organism.