Harnessing protein clustering to understand, identify, and manipulate cellular systems

Summary/Abstract The Bugaj Lab develops precision-controlled molecular tools to better understand and control biological systems. This proposal will develop technologies that harness protein clustering for new, enabling methods to study, manipulate, and treat cells. Protein clustering plays important roles throughout cell physiology and disease, yet its potential for biotechnological probes and therapies remains unrealized. Clustering is potent because it achieves rapid and non-linear responses with minimal energetic costs. Over the past decade, optogenetic clustering probes have allowed control over protein clustering with high precision throughout a wide array of cellular processes. However, despite the high utility of optogenetic clustering, there remains predominantly one protein (A. thaliana Cry2) whose light-induced clustering is used for such probes, constraining its application due to inherent limitations, for example response time and ability to control one process at a time. Our first project will address these technological gaps by engineering a distinct photoreceptor that we recently discovered possesses a native ability to form light-induced clusters in mammalian cells, providing a complement to Cry2 with certain favorable properties, including in response kinetics. In our second project, we will develop a suite of single-component optogenetic probes that allow precise control of subcellular localization to a variety of compartments. These probes uniquely harness the ability of clustering to trigger binding through rapid increases in probe avidity. In our third project, we will develop the foundations of synthetic protein therapies that can sense and respond to endogenous protein clusters. Clustering is a hallmark of many pathologies including neurodegeneration, viral infection, and cancer, and thus protein clustering itself could be leveraged as a disease biomarker or therapeutic target. We will engineer autonomous protein systems that detect pathological protein aggregates and execute a strong, appropriate response, for instance killing a cancer cell or secreting cytokines to recruit the immune system. Successful completion of our work will establish protein clustering as a flexible new paradigm for manipulation of cellular function, resulting in broadly applicable probes for study of cell/animal behavior, as well as a new class of “sense-and-respond” smart drugs with application across a diverse array of disease states.