# RuPHOTACs Provide Photocontrol Over Protein Degradation with Optimized Properties for Biological Applications

## TL;DR

This paper introduces RuPHOTACs, a new type of light-controlled molecule that improves the precision and effectiveness of protein degradation in cells.

## Contribution

The novel use of Ru(II) photocages in PROTACs enables precise light-controlled protein degradation with enhanced selectivity and in vivo efficacy.

## Key findings

- RuPHOTACs showed high selectivity for light-triggered activation and improved potency against target proteins.
- The system effectively reduced levels of c-MYC and PIM1 in cells using low energy red light.
- A new reporter system using Dendra2 fusion proteins accurately measures protein degradation rates in live cells.

## Abstract

PROteolysis TArgeting Chimeras (PROTACs) are bifunctional molecules that catalyze degradation of selected proteins by inducing protein:protein interactions (PPIs) between E3 ubiquitin ligases and the protein of interest. A critical limitation is undesired effects in untargeted tissues, necessitating approaches to impose spatiotemporal control over PROTAC function. Here we present Ru(II) photocages that can be released with low energy light, providing triggered PROTAC activity on demand. The systems, termed Ruthenium-based PHOToActivated Chimeras (RuPHOTACs), were validated by targeting bromodomain-containing proteins, which act as crucial epigenetic regulators, and also strongly reduced levels of c-MYC and PIM1. The novel RuPHOTACs demonstrate that the incorporation of metal components within organic PROTACs confers multiple advantages for light-controlled systems for chemical biology applications, including the highest selectivity for activation in the light vs. the dark, improved potency against the target proteins of interest, and increased efficacy in vivo using low energy red light. To efficiently monitor protein degradation via optical means, a new strategy was implemented by creating a fusion of a photoconvertible protein, Dendra2, with the proteins of interest. This bifunctional reporter system for live cell analysis decouples protein degradation efficiency and rates from signals arising from new protein production, and is superior to prior reporter systems described for PROTACs because it directly measures the true degradation rate of the target protein in intact, viable cells while simultaneously tracking newly synthesized protein without perturbing translation, proteostasis, or cell health.