PLAMseq enables the proteo-genomic characterization of chromatin-associated proteins and protein interactions in a single workflow.

ABSTRACT: Chromatin Immunoprecipitation (ChIP) and Co-Immunoprecipitation (CoIP) assays are the most common approaches to characterize the genomic localization and protein interactors, respectively, for a protein of interest. However, these approaches require the use of specific antibodies, which are costly reagents that often face sensitivity and specificity issues. Based on TurboID, we developed PLAMseq (Proximity Labelled Affinity-purified Mass spectrometry plus sequencing), which enables, in the same workflow, to identify the genomic loci and the interacting proteome of a protein of interest. Moreover, PLAMseq can also be applied to specifically map protein interactions across the genome or to specifically detect proteins which are ubiquitin(-like) modified. After a short biotin pulse, DNA-protein crosslinks are induced by formaldehyde and the interactors of a protein of interest, together with their associated DNA sequences, are purified simultaneously and identified by mass spectrometry-based proteomics and Next Generation Sequencing, respectively.To validate PLAMseq, we performed the proteo-genomic characterization of two proteins which genomic loci are very well characterized, namely, RNA polymerase II and CTCF, with excellent robustness and reproducibility. Next, we applied PLAMseq to characterize Histone H1 SUMOylation, a histone post-translational modification which study has remained elusive due to the lack of specific reagents. We found that SETDB1 binds to SUMOylated histone H1.2 and H1.4 and, accordingly, SUMOylated histone H1 colocalizes with H3K9me3 at repetitive regions of the genome.

INSTRUMENT(S):

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Cell Culture

SUBMITTER: Román González-Prieto  

LAB HEAD: Román González-Prieto

PROVIDER: PXD062203 | Pride | 2025-10-24

REPOSITORIES: Pride