Regulator of G protein signaling 3 modulates Wnt5b calcium dynamics and somite patterning

The process of vertebrate development requires communication among many cells of the embryo in order to define the body axis (front/back, head/tail or left/right). The Wnt signaling network plays a key role in a vast array of cellular processes including body axis patterning and cell polarity. One arm of the Wnt signaling network, the non-canonical Wnt pathway, mediates intracellular Ca²⁺ release via activation of heterotrimeric G proteins. Regulator of G protein Signaling (RGS) proteins can accelerate inactivation of G proteins by acting as G protein GAPs and are uniquely situated to control the amplitude of a Wnt signal. I hypothesize that individual RGS proteins are critical in modulating the frequency and amplitude of Wnt/Ca²⁺ signaling in different tissues and at different developmental stages and this modulation is essential for developmental patterning events. To this end, this thesis is focused on the effects G protein regulation has on intracellular Ca²⁺ release dynamics and developmental patterning events.

I combine cellular, molecular and genetic analyses with high resolution Ca²⁺ imaging to provide new understanding of the role of RGS proteins on Wnt mediated Ca²⁺ release dynamics and developmental patterning events. In chapter 2, I describe endogenous Ca²⁺ dynamics from the very first cell divisions through early somitogenesis in zebrafish embryos. I find that each phase of zebrafish development has a distinct pattern of Ca²⁺ release, highlighting the complexity of Ca²⁺ ion and cellular physiology.

In Chapter 3, I identify rgs3 as potential modulator of Ca²⁺ dynamics and Chapter 4 expands upon these observations by providing data supporting that Rgs3 function is necessary for appropriate frequency and amplitude of Ca²⁺ release during somitogenesis and that Rgs3 functions downstream of Wnt5 activity. My results provide new evidence that a member of the RGS protein family is essential for modulating the non-canonical Wnt network to assure normal tissue patterning during development.

In Chapter 5, I report the identification and characterization of Rgs3b, a paralogue to Rgs3, in zebrafish. I describe results indicating that Rgs3b is poised to interact with Wnt11 indicating that individual RGS genes may have unique roles in modulating Wnt/Ca2+ signaling in different tissues or different stages. In conclusion, this thesis provides data supporting that individual RGS proteins are critical in modulating the frequency and amplitude of Wnt/Ca²⁺ signaling in different tissues and at different developmental stages and this is a substantial breakthrough in understanding how RGS proteins function to fine-tune known signaling pathways