Date of Award

2017

Degree Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Graduate Group

Chemistry

First Advisor

David M. Chenoweth

Abstract

Fundamental biological processes including cell division, migration, and death,

are driven by protein interactions. Regulation of protein localization is one of the

mechanisms cells utilize to control cellular events with high spatial and temporal

precision. Therefore, several techniques have been developed to provide control of

protein interactions and localization. A number of elegant approaches employ naturally

light-responsive proteins, also known as optogenetics, to reversibly induce protein‒

protein binding interactions with subcellular precision. However, the application of these

light-inducible protein systems to various intracellular locations beyond the plasma

membrane has been limited. Moreover, to achieve sustained interactions in some

applications, most of these optogenetic systems require continuous illumination,

increasing the risk of phototoxicity. Another robust and widely utilized technique to

control protein interactions via small molecules is the chemically-induced dimerization

(CID) of proteins; the most classic example of this technique being rapamycin-induced

dimerization. However, the lack of spatiotemporal control and reversibility in this system

has necessitated the development of new dimerizers in the past two decades. By

combining light-inducible features with the CID technique, we have created a novel

platform to rapidly and reversibly induce protein dimerization using light with high

specificity in living cells. This is accomplished with subcellular spatiotemporal resolution

using a series of novel, cell-permeable, photoactivatable, and photocleavable chemical

dimerizers. The modular design of our system has allowed us to tailor the properties of

our molecules for studying various protein functions and biological pathways inside

living cells. Furthermore, we demonstrate the utility of our system by applying it to

manipulate dynamic biological events including organelle transport and spindle assembly

checkpoint. This work establishes a foundation for optogenetic control over protein

function and highlights the advantages of a hybrid chemical and genetic approach. We

envision our tools to be readily adapted to experimentally probe complex signaling

networks and other cellular processes that depend upon spatiotemporal regulation of

protein localization on biologically-relevant timescales.

Embargoed

Available to all on Monday, January 18, 2021

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