BiFCo: visualizing cohesin assembly/disassembly cycle in living cells

Abstract

Cohesin is a highly conserved, ring-shaped protein complex found in all eukaryotes. It consists of at least two structural maintenance of chromosomes (SMC) proteins, SMC1 and SMC3 in humans (Psm1 and Psm3 in fission yeast), and the kleisin RAD21 (Rad21 in fission yeast). Mutations in its components or regulators can lead to genetic syndromes, known as cohesinopathies, and various types of cancer. Studies in several organisms have shown that only a small fraction of each subunit assembles into complexes, making it difficult to investigate dynamic chromatin loading and unloading using fluorescent fusions in vivo because of excess soluble components. In this study, we introduce bimolecular fluorescent cohesin (BiFCo), based on bimolecular fluorescent complementation in the fission yeastSchizosaccharomyces pombe. BiFCo selectively excludes signals from individual proteins, enabling the monitoring of complex assembly and disassembly within a physiological context throughout the entire cell cycle in living cells. This versatile system can be expanded and adapted for various genetic backgrounds and other eukaryotic models, including human cells.