Thesis Seminar: Patricia Schneider
Role of Axin-RGS domain during axis formation: maternal vs. zygotic functions: Patricia Schneider 16.April (Friday) at 10:00 in 106 BBE.
Upon sperm entry, the vertebrate egg undergoes a series of cell divisions that create a number of smaller cells without increasing the embryonic mass. This induces an elevation of intracellular calcium transient that is conserved across species. In zebrafish, fertilization occurs through an opening in the chorion, the micropyle and in Xenopus it can occur anywhere in the animal hemisphere. The initial cell divisions cause a rotation in the cortex called cortical rotation. This process ends with the termination of the first cell division and coincides with the translocation of maternal factors to the future dorsal side of the embryo. These maternal factors will drive the initial steps of development and pattern the dorsal axis. wnt signaling is critical during the specification of the dorsal-ventral axis. During gastrulation, a signaling center, the Spemann organizer, is formed and will secrete molecules to pattern the embryo. During this stage, the embryo will form the three germ layers: ectoderm, mesoderm and endoderm. In zebrafish and Xenopus the induction of the neural tissue occurs during gastrulation and is dependent upon the organizer activity.
Wnt signaling activation is required during dorsal-ventral axis specification and it needs to be suppressed during the regionalization of the brain. Axin is a negative regulator of Wnt signaling and contains an RGS (Regulator of G Protein Signaling) domain. RGS domains are typical of RGS proteins, which are involved in a distinct signaling pathway, G-protein signaling. RGS proteins exert a negative effect of G-protein signaling by accelerating the GTPase activity (GAP) of the Ga subunit, thus turning off the signaling. Axin contains an RGS domain, however, it is not clear whether Axin is directly involved in G-protein signaling. In chapter 2, we describe the role of a Wnt antagonist, Axin, during the patterning of the vertebrate brain. We manipulated this protein by creating a point mutation in a critical amino acid within the Axin-RGS domain, known to be detrimental for the GAP function of RGS proteins, Axin1Q162A. We show that this point mutant is not able to restore normal brain development in zebrafish embryos after Axin knockdown. We hypothesize that Axin-RGS like function is necessary during the patterning of the vertebrate brain that occurs after zygotic transcription has been initiated.
Before MBT (mid-blastula transition) the developing embryo is dependent upon maternal contribution of molecules that were deposited from the mother into the egg during oogenesis. In chapter 3, we show that the Axin-RGS like function is dispenable during the formation of the dorsal-ventral axis. Maternal depletion of Axin1 in Xenopus oocytes causes hyperactivation of Wnt signaling and results in dorsalization. Axin1Q162A is able to suppress the dorsalization of maternally depleted embryo and restore normal dorsa-ventral axis formation. We hypothesize that Axin-RGS like function may be dispensable during this stage of development due to a receptor-independent way to activate Wnt signaling during maternal stages.
Finally, we demonstrate that Axin1Q162A localization differs from the wildtype Axin1 and Axin1 but not Axin1Q162A is localized to the plasma membrane upon Ga overexpression in zebrafish embryos.