Gary Karpen

Professor of Cell Biology, Development and Physiology

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Research Interests

Our current research is focused on understanding how nuclear organization, chromatin composition, and condensate and polymer biophysics impact genome and cellular functions, and organismal health. To generate a deeper view of these essential dynamic processes at multiple spatial scales, we utilize interdisciplinary strategies that integrate cell biological, genomic, biophysical and computational approaches.

Most of our in vivo and in vitro studies have focused on the fruit fly Drosophila melanogaster as a model for chromosome function in metazoans, which allows us to apply interdisciplinary approaches to elucidate molecular mechanisms and consequences in a developing, multicellular animal. However, we also examined the relevance of our findings to human cells and other organisms, and have demonstrated surprising similarities between evolutionarily-distant species.

Current Projects

In recent years we have elucidated the roles of biocondensate formation in the 4D organization of chromatin domains, the impact of chromatin states on DNA repair effectiveness and pathway choice, revealed new aspects of genome and protein evolution, and discovered a novel role for RNAs generated by highly repeated satellite DNAs. Much of our research has focused on the heterochromatin (HC), a major component of eukaryotic genomes that regulates important functions such as nuclear architecture, genome stability, transposon silencing and chromosome inheritance. HC regions are highly enriched for repeated DNAs (satellites, ribosomal DNA, transposons), and nucleosomes containing methylated H3 lysine 9 (H3K9me2/3) and its binding partner Heterochromatin Protein 1a (HP1a). How heterochromatin regulates many dynamic and essential functions, and roles in human diseases such as cancer, remain poorly understood.

Chromatin condensates: Mechanisms and impact. Our major efforts have been focused on the biophysics of chromatin domains and their impact on nuclear architecture and genome functions. Specifically, we showed that the 3D heterochromatin (HC) domain is a biocondensate that displays liquid properties and forms by phase separation or similar mechanisms (Strom et al. Nature 2017). This work generated a fundamentally different conceptualization of how chromatin domains are formed and function, and generated much excitement in the field. Phase separation and emergent biophysical properties provide novel mechanisms for biological regulation at multiple scales, due to exquisite sensitivity to relatively minor changes in molecule concentrations, temperature, ionic environment, and post-translational modifications (PTMs). Exciting impacts include liquid fusions driving contact between distant genomic regions (Lee et al. ELife 2019), spreading of chromatin states through ‘wetting’ rather than progressive histone modification, and regulation of genome functions such as repair, replication and transcription by moving genomic regions into or out of condensates. We continue to study the molecular mechanisms and interactions that drive condensate formation, and perhaps most importantly, if and how emergent biophysical properties impact genome organization and functions in vivo, using advanced imaging, chemical, biochemical, and experimental and theoretical biophysical approaches (Tortora et al. PNAS 2023; Rajshekar et al. Nature Cell Biology 2025; Brennan et al. BioRxiv 2025; Colmenares et al. Nucleic Acids Research 2025) . 

Regulation of DNA repair by chromatin and nuclear dynamics. Repeated DNAs enriched in heterochromatin (HC) pose significant challenges to genome stability. For example, homologous recombination (HR) involving double-strand breaks (DSBs) in repetitive sequences generates chromosome aberrations that cause cell death and contribute to cancer progression. We challenged prevalent views by revealing that a novel, dynamic mechanism regulates HR in HC (Chiolo et al., Cell 2011), and defined the proteins and pathways required for relocalization of DSBs outside the HC domain and completion of HR repair at the nuclear periphery (Ryu et al. NCB 2015). We proposed that spatially separating repeats with DSBs from the rest of the HC reduces the probability of aberrant exchanges that cause genome instability (e.g., translocations) and promotes less harmful HR repair from homologs or sister chromatids. We also developed inducible single DSB systems that, for the first time, allowed direct comparisons of repair pathways and products, foci dynamics and regulatory mechanisms in HC and euchromatin (EC) in animal tissues (Janssen et al., G&D 2016). Overall, we conclude that despite extensive differences in chromatin composition, both the HR and NHEJ pathways are utilized to a similar extent for EC and HC DSB repair, and HC DSBs can switch between NHEJ and HR utilization. Based on recent results (Janssen, Colmenares et al. G&D 2019), we hypothesize that DSB relocalization, repair protein recruitment and pathway choice require removal of HC chromatin marks and replacement with ‘active’ marks (e.g. acetylations). Thus, current and future efforts are focused on using single DSB systems in flies and human cells to understand how chromatin modifications and modifiers regulate repair dynamics and pathway utilization, and how they differ in HC and EC (Kendak et al. 2024 Nucleic Acids Research). 

Evolution of centromeres and heterochromatin

Haplotypes spanning centromeric regions reveal persistence of large blocks of archaic DNA. In collaboration with Dr. Charles Langley (UC Davis), we identified large-scale haplotypes (termed 'cenhaps') that span the pericentromeric heterochromatin, including the centromeres of Drosophila and human chromosomes. Human cenhaps revealed deep diversity and the largest (Mb-size!!) ancient Neanderthal and African introgressions identified to date (Langley et al., ELife 2019). Cenhaps contain variants, including large differences in α-satellite DNA content, which could influence the fidelity and bias of chromosome transmission. Incorporation of Cenhaps into the completed human genome assemblies generated by the Telomere-to-Telomere project helped reveal the evolutionary history of human centromeres (Altemose et al. Science 2022). We are currently investigating cenhap contributions to human variation and disease, and to more incisively model the rich evolution of these challenging genomic regions. 

Unfortunately, we cannot use human cenhaps identified through bioinformatic methods to perform direct experimental interrogations of impact on chromosome and organismal functions. Thus, we also identified highly diverse cenhaps from Drosophila populations isolated from across the world; they display highly significant associations (e.g. with clinal variation), and thus hold promise for studies directly linking haplotypes to functional differences. Ongoing collaborative studies will allow us to propose and test population genomic models and mechanistic explanations of the forces that shape variation in the pericentromeric regions, within and between species.  

RNAs generated from a simple repeat (AAGAG) are required for spermatogenesis. Tandemly-repeated short DNAs, or satellites, are enriched in HC regions of eukaryotic genomes; although some satellites are transcribed, we lacked direct evidence that specific satellite RNAs are required for normal organismal functions. We observed that satellite RNAs derived from AAGAG tandem repeats (4% of the Drosophila genome) are transcribed in many cells throughout Drosophila development, enriched in neurons and testes, localize to HC domains, and required for viability. Strikingly, we find AAGAG transcripts are necessary for male fertility, and that AAGAG RNA depletion results in defective histone-protamine exchange, sperm maturation and chromatin organization (Mills et al. Elife 2019). Since these events happen late in spermatogenesis when the transcripts are not detected, we speculate that AAGAG RNA in primary spermatocytes ‘primes’ post-meiosis steps for sperm maturation. In addition to the surprising discovery of essential functions for AAGAG RNAs, comparisons between closely related Drosophila species suggest that satellites and their transcription evolve quickly to generate new functions. We are currently investigating how fast satellite evolution can be reconciled with encoding essential functions, and potential roles for satellite transcripts in health, disease and evolution.

Selected Publications

Brennan, L., Kim, H., Colmenares, S., Ego, T., Ryu, J., and Karpen, G.H. (2025) HP1a promotes chromatin liquidity and drives spontaneous heterochromatin compartmentalization. BioRxiv doi: 10.1101/2024.10.18.61898 (PMID39868136)

Colmenares, S.U., Tsukamoto, S., Hickmann, C., Brennan, L.D., Khavani, M., Mofrad, M. and Karpen, G.H. (2025) Expanding the HP1a-binding consensus and molecular grammar for heterochromatin assembly. Nucleic Acids Research 53(19): gkaf976. doi:10.1093/nar/gkaf976 (PMID 41099702)

Rajshekar S., Adame-Arana O., Bajpai G., Lin K., Colmenares S., Safran S., Karpen G.H. (2025) Affinity hierarchies and amphiphilic proteins underlie the co-assembly of nucleolar and heterochromatin condensates. Nature Cell Biology (in press).

Kendek A, Sandron A, Lambooij JP, Colmenares SU, Pociunaite SM, Gooijers I, de Groot L, Karpen G.H., Janssen A. (2024) DNA double-strand break movement in heterochromatin depends on the histone acetyltransferase dGcn5. Nucleic Acids Res. 52(19):11753-11767. doi: 10.1093/nar/gkae775. (PMID39258543)

Tortora, M.M., Brennan, L., Karpen, G.H., and Jost, D. (2023) HP1-driven phase separation recapitulates the thermodynamics and kinetics of heterochromatin condensate formation. PNAS 120, e2211855120 (PMID37549295)

Altemose, N. et al. (2022) Complete genomic and epigenetic maps of human centromeres. Science 376, eabl4178 doi: 10.1126/science.abl4178 (PMID35357911)

Lee, Y.C.G., Ogiyama, Y., Martins, N.M.C., Beliveau, B.J., Acevedo, D., Wu, C.T., Cavalli, G., and Karpen, G.H. (2020) Pericentromeric heterochromatin is hierarchically organized and spatially contacts H3K9me2/3 islands located in euchromatic genome. PLoS Genetics 16(3): e1008673 (PMID32203508)

Mills, W.K., Lee, Y.C.G., Kochendoerfer, A.M., Dunleavy, E.M., and Karpen, G.H. (2019) RNA from a simple-tandem repeat is required for sperm maturation and male fertility in Drosophila melanogaster. eLife Nov 5;8.doi: 10.7554/eLife.48940 (PMID31687931)

Langley, S.A., Miga, K., Karpen, G.H. and Langley, C.H. (2019) Haplotypes spanning centromeric regions reveal persistence of large blocks of archaic DNA. eLife 8:e42989 (PMID3123723)

Janssen, A., Colmenares, S.U., Lee, T., and Karpen, G.H. (2019). Timely double-strand break repair and pathway choice in pericentromeric heterochromatin depend on the histone demethylase dKDM4A. Genes and Development 33, 103–115 (PMID30578303)

Strom, A.R. Emelyanov, A.V., Mir, M., Fyodorov, D.V., Darzacq, X. and Karpen, G.H. (2017) Phase separation drives heterochromatin domain formation.  Nature 547, 241 (PMID28636597)
News & Views: Klosen, A. and Hyman, A.A. (2017) Molecular biology: A liquid reservoir for silent chromatin. Nature 547, 168

For full list of publications see https://www.ncbi.nlm.nih.gov/myncbi/gary.karpen.1/bibliography/public/

Photo credit: Mark Hanson at Mark Joseph Studios.

Last Updated 2025-11-10