Decoding chromosome organization using CheC-PLS: chromosome conformation by proximity labeling and long-read...

Area of Science:

• Genomics
• Molecular Biology
• Chromatin Biology

Background:

• Genomic techniques offer insights into chromosome structure but often miss connectivity information.
• Large-scale genome organization remains poorly understood due to limitations in current methods.

Purpose of the Study:

• To develop and validate CheC-PLS, a novel proximity-labeling technique for analyzing in vivo genome organization.
• To characterize the 3D organization of chromatin and protein associations at high resolution.

Main Methods:

• CheC-PLS utilizes dam methyltransferase tethered to a protein of interest.
• Nanopore sequencing identifies methylated bases, indicating in vivo proximity along long DNA reads (>100kb).
• The technique was applied to budding yeast and isolated nuclei using nanobodies.

Main Results:

• CheC-PLS successfully recapitulated known cohesin association patterns in budding yeast.
• The method revealed cell-to-cell variability in chromosome organization.
• Single-read data demonstrated cohesin translocation and defined the internal structure of the ribosomal DNA locus.

Conclusions:

• CheC-PLS provides a versatile tool for decoding protein-associated sites and understanding in vivo chromosome conformations.
• The technique illuminates dynamic chromosomal processes and single-chromosome organization.
• This method advances the study of genome architecture and its variability.