Axel Imhof – Chromatin Proteomics

Every cell division challenges the epigenome twice. During DNA replication, parental histones and their modifications are shared between the two daughter strands and diluted by newly made, unmodified histones. During mitosis, chromatin is compacted into chromosomes and must be reset for the next G1 phase. We use quantitative mass spectrometry, isotope labelling and mathematical modelling to follow histone modifications through these transitions.

With the Groth, Rupp and Marr groups we showed that methylation of H3K27, H3K36 and H4K20 is restored slowly after replication, through a domain-based mechanism. In dividing cells, dilution by replication rather than active demethylation shapes H4K20 methylation. Demethylation becomes essential once cells stop dividing, as in Xenopus embryos with a systemic cell-cycle block. Most recently, working with the Earnshaw lab, we built a time-resolved atlas of histone modifications during mitotic entry. It revealed three separate phosphorylation programmes on histone H3 (S10, T3 and S28) and challenged the long-held idea that global changes in other histone marks are needed for chromosome condensation.

Key publications

Kochanova NY et al. (2026) A time-resolved atlas of histone modifications during mitotic entry. Molecular Cell. 10.1016/j.molcel.2026.03.038

Pokrovsky D et al. (2021) A systemic cell cycle block impacts stage-specific histone modification profiles during Xenopus embryogenesis. PLOS Biology. 10.1371/journal.pbio.3001377

Schuh L et al. (2020) H4K20 methylation is differently regulated by dilution and demethylation in proliferating and cell-cycle-arrested Xenopus embryos. Cell Systems. 10.1016/j.cels.2020.11.003

Alabert C et al. (2020) Domain model explains propagation dynamics and stability of histone H3K27 and H3K36 methylation landscapes. Cell Reports. 10.1016/j.celrep.2019.12.060

Heterochromatin is made up largely of repetitive satellite DNA and transposable elements, and it is among the fastest-evolving parts of the genome. We study how this rapid evolution can drive species apart. Our model is the hybrid incompatibility between Drosophila melanogaster and D. simulans, which is caused by the genes Hmr and Lhr. Their expression has diverged between the two species, so hybrids contain higher levels of both proteins, which leads to the death of hybrid males.

We found that HMR and LHR form a six-subunit complex that links centromeric and pericentromeric chromatin. The integrity of this complex is required both for its binding to centromeres and for hybrid lethality. Raising HMR/LHR levels in pure species is enough to cause mitotic defects, derepress transposable elements and reduce fertility. To see how incompatible genomes can still coexist, we have established hybrid Drosophila cell lines. These lines reveal the adaptations that allow survival, including aneuploidy, chromosome loss, dosage compensation and rDNA amplification: in effect, "speciation in reverse".

Key publications

- Lukacs A, Thomae AW et al. (2021) The integrity of the HMR complex is necessary for centromeric binding and reproductive isolation in Drosophila. PLOS Genetics. 10.1371/journal.pgen.1009744

- Kochanova NY et al. (2020) A multi-layered structure of the interphase chromocenter revealed by proximity-based biotinylation. Nucleic Acids Research. 10.1093/nar/gkaa145

- Thomae, A. W. et al. A Pair of Centromeric Proteins Mediates Reproductive Isolation in Drosophila Species. Developmental Cell 10.1016/j.devcel.2013.10.001

Centromeres are defined epigenetically by the histone H3 variant CENP-A rather than by their DNA sequence. In interphase nuclei, centromeres cluster with surrounding heterochromatin into chromocenters. We map the molecular organisation of these domains using proximity labelling combined with quantitative proteomics and super-resolution microscopy.

By labelling the neighbourhoods of CENP-A, HMR and HP1a, we found that the chromocenter has a layered structure: a CENP-A core, a bridging layer containing HMR, LHR and NLP, and an outer shell of HP1a heterochromatin. We also identified the histone chaperone Spt6 as a maintenance factor for CENP-A. Spt6 keeps old CENP-A nucleosomes at the centromere while it is transcribed in mitosis and G1. With AMPL-MS, a new antibody-guided proximity-labelling method for proteins and RNA, we showed that RNA helps hold these chromatin domains together. The same work revealed centromeric R-loops connected to the RNA helicase MLE. We are now extending this work to human centromeres.

Key publications

Parashara, P. et al. PLK1-mediated phosphorylation cascade activates Mis18 complex to ensure centromere inheritance. Science 385, 1098–1104 (2024). 10.1126/science.ado8270

Choudhury R et al. (2024) The role of RNA in the maintenance of chromatin domains as revealed by antibody-mediated proximity labelling coupled to mass spectrometry. eLife 13:e95718. 10.7554/elife.95718

Lukacs A, Thomae AW et al. (2021) PLOS Genetics (see above, "... Origin of Species").

Bobkov GOM et al. (2020) Spt6 is a maintenance factor for centromeric CENP-A. Nature Communications 11. 10.1038/s41467-020-16695-7

Kochanova NY et al. (2020) Nucleic Acids Research (see above, "Timing the histone code ...").