Modulation of Light-Induced Homo-Oligomerization of Cryptochrome2

Victor Acero1, Liting Duan2, Jen Hope2, Bianxiao Cui2

The Pennsylvania State University1, Stanford University2

Abstract
Protein-protein interactions, the driving force for all cell activity, regulate a multitude of
biological processes ranging from stem cell differentiation to gene expression. Drugs, hormones,
and other stimulants are commonly used to perturb protein interactions. However they offer poor
spatiotemporal resolution. Optogenetic methods seek to develop a platform which allows precise
control of protein-protein interactions via the use of light-sensitive proteins. One of the most
commonly used photosensory proteins, Arabidopsis thaliana Cryptochrome2 (CRY2), can
undergo both light induced homo-oligomerization and hetero-dimerization with its binding
partner CIB1, which can confound results and analysis. Therefor it is crucial to effectively
modulate CRY2 oligomerization, particularly during hetero-dimerization. This work seeks to
determine how fluorescence tags and bulky fusion proteins may modulate homo-oligomerization.
We found that GFP fused CRY2 oligomerizes less than the mCherry fused counterpart.
Furthermore, we found that CRY2 oligomerization can be significantly inhibited by the
tdTomato fused CIB1 during hetero-dimerization; tdTomato is a bulky fluorescence protein
which provides steric hindrance towards oligomerization.