TINC: a method to dissect transcriptional complexes at single locus resolution
- Department of Anatomy and Developmental Biology, Monash University, VIC, Australia.
- Development and Stem Cells Program, Monash Biomedicine Discovery Institute, VIC, Australia.
- Australian Regenerative Medicine Institute, Monash University, VIC, Australia.
- Australian Research Council Centre of Excellence in Plant Energy Biology, The University of Western Australia, WA, Australia.
- Harry Perkins Institute of Medical Research, WA, Australia.
- Monash Biomedical Proteomics Facility, Monash University, VIC, Australia.
- Australian Centre for Blood Diseases, Monash University, VIC, Australia.
Being able to determine the molecular composition of protein complexes that assemble at specific regulatory elements to activate or repress gene transcription is essential for a better understanding of how gene expression is controlled in normal and diseased states. We have developed a transcription activator-like effectors (TALE) based method termed TINC (TALE-mediated Isolation of Native Chromatin), which enables the isolation of a specific chromatin region from mammalian cells and consequent identification of associated proteins by mass spectrometry. For proof of concept, we targeted the Nanog proximal promoter in mouse embryonic stem cells and were able to identify transcription factors known to bind to this locus and most importantly novel proteins that play an essential role in pluripotency and reprogramming. As TINC does not require any genetic modification of the target sequence, a target-specific antibody nor high copy numbers of the target sequence, we strongly believe that this method is applicable to any scientific field and has immense potential to change the concept of how we study gene regulation.