Designer Receptors for Nucleotide-Resolution Analysis of Genomic 5-Methylcytosine by Cellular Imaging.
Álvaro Muñoz-LópezBenjamin BuchmullerJan WolffgrammAnne JungMichelle HussongJulian KanneMichal R SchweigerDaniel SummererPublished in: Angewandte Chemie (International ed. in English) (2020)
We report programmable receptors for the imaging-based analysis of 5-methylcytosine (5mC) in user-defined DNA sequences of single cells. Using fluorescent transcription-activator-like effectors (TALEs) that can recognize sequences of canonical and epigenetic nucleobases through selective repeats, we imaged cellular SATIII DNA, the origin of nuclear stress bodies (nSB). We achieve high nucleobase selectivity of natural repeats in imaging and demonstrate universal nucleobase binding by an engineered repeat. We use TALE pairs differing in only one such repeat in co-stains to detect 5mC in SATIII sequences with nucleotide resolution independently of differences in target accessibility. Further, we directly correlate the presence of heat shock factor 1 with 5mC at its recognition sequence, revealing a potential function of 5mC in its recruitment as initial step of nSB formation. This opens a new avenue for studying 5mC functions in chromatin regulation in situ with nucleotide, locus, and cell resolution.
Keyphrases
- single molecule
- high resolution
- heat shock
- gene expression
- induced apoptosis
- circulating tumor
- transcription factor
- cell free
- stem cells
- copy number
- bone marrow
- cell cycle arrest
- heat stress
- mesenchymal stem cells
- cell therapy
- genome wide
- nuclear factor
- quantum dots
- oxidative stress
- risk assessment
- heat shock protein
- mass spectrometry
- dna binding
- amino acid
- genetic diversity