Date of Award

12-2011

Document Type

Campus Access Dissertation

Degree Name

Doctor of Philosophy (PhD)

Department

Biology/Molecular, Cellular, and Organismal Biology

First Advisor

Alexey Veraksa

Second Advisor

Linda Huang

Third Advisor

Katherine Gibson

Abstract

The development of an organism is orchestrated by a small number of signaling pathways. In order to achieve tissue specificity, their activities are controlled by multiple regulatory molecules that fine-tune pathway output and increase the specificity of signaling. To better understand how these pathways function it is imperative that we strive to elucidate all signaling components. Affinity purification followed by Mass Spectrometry (AP-MS) has proved itself as a suitable technique in achieving this aim. Analysis of the Notch pathway via this method has identified Drosophila β-arrestin Kurtz (Krz) as a negative modulator of the pathway. I have further studied Krz activities during embryonic development in Drosophila, and have revealed that Krz functions as a general inhibitor of receptor tyrosine kinase (RTK) signaling pathways. My molecular analysis suggests that the activity of Krz is exerted at the level of the major RTK signaling effector, ERK, by a novel inhibitory mechanism. Genetic analysis further suggests that this mechanism is also employed in regulating RTK signaling in other tissues and developmental stages. In addition, I have shown that Krz also negatively regulates the Toll/Dorsal dorsoventral patterning pathway in the early embryo. My findings reveal a new in vivo function of β-arrestins, and demonstrate that they can employ different mechanisms to regulate diverse developmental signaling pathways. This work began by Krz identification by AP-MS. I have utilized advancements in manipulating large genomic constructs in Drosophila, and continued to implement the AP-MS method to further our understanding of the Notch protein interaction network. In addition to the Notch pathway I have also taken a similar approach in the study of a novel growth regulator myopic (mop). Together with work from collaborators, I have identified Mop as a modulator of the Hippo Kinase pathway through interaction with oncoprotein Yorkie. In sum, this work has uncovered a novel role of Drosophila β-arrestin Krz in development, and demonstrated AP-MS as a powerful tool for dissecting developmental signaling networks in Drosophila.

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