Aberrant gene regulation is a hallmark of cancer, and many oncogenic programs are driven by altered transcription factor binding to chromatin. In leukemia, these changes lead to impaired differentiation, dysregulated proliferation, and resistance to therapy, making leukemia a useful model system for studying disease mechanisms dependent on transcription factors.

A standard approach to measuring transcription factor occupancy is chromatin immunoprecipitation coupled with sequencing or targeted quantification. This approach has enabled genome-wide maps of protein-DNA interactions. However, conventional workflows can be limited by crosslinking chemistry, indirect interactions, and background signal. Photochemical, UV-based crosslinking provides a complementary approach because UV irradiation forms so-called zero-length crosslinks between nucleic acids and proteins in immediate contact. This enriches direct binding events in a time-resolved manner.

In this course, we will test the binding of CTCF, a master organizer of chromatin architecture, to selected DNA-binding sites in K562 cells.

Comparative analysis of CTCF ChIP-seq after ultraviolet (UV) laser light or formaldehyde (FA) fixation.
a) Examples of CTCF FA ChIP-seq (gray) and UV laser ChIP-seq using sonication (orange) or MNase digestion (blue) binding profiles at different genomic loci. (b) Venn diagram illustrating the number of detected binding sites in CTCF FA ChIP-seq (gray) and UV laser ChIP-seq after sonication (orange) or MNase digestion (blue).