Nucleolar release of rDNA repeats for repair involves SUMO-mediated untethering by the Cdc48/p97 segregase

Ribosomal RNA genes (rDNA) are highly unstable and susceptible to rearrangement due to their repetitive nature and active transcriptional status. Sequestration of rDNA in the nucleolus suppresses uncontrolled recombination. However, broken repeats must be first released to the nucleoplasm to allow repair by homologous recombination. Nucleolar release of broken rDNA repeats is conserved from yeast to humans, but the underlying molecular mechanisms are currently unknown. Here we show that DNA damage induces phosphorylation of the CLIP-cohibin complex, releasing membrane-tethered rDNA from the nucleolus in Saccharomyces cerevisiae. Downstream of phosphorylation, SUMOylation of CLIP-cohibin is recognized by Ufd1 via its SUMO-interacting motif, which targets the complex for disassembly through the Cdc48/p97 chaperone. Consistent with a conserved mechanism, UFD1L depletion in human cells impairs rDNA release. The dynamic and regulated assembly and disassembly of the rDNA-tethering complex is therefore a key determinant of nucleolar rDNA release and genome integrity

DNA damage-induced histone H1 ubiquitylation is mediated by HUWE1 and stimulates the RNF8-RNF168 pathway

The DNA damage response (DDR), comprising distinct repair and signalling pathways, safeguards genomic integrity. Protein ubiquitylation is an important regulatory mechanism of the DDR. To study its role in the UV-induced DDR, we characterized changes in protein ubiquitylation following DNA damage using quantitative di-Gly proteomics. Interestingly, we identified multiple sites of histone H1 that are ubiquitylated upon UV-damage. We show that UV-dependent histone H1 ubiquitylation at multiple lysines is mediated by the E3-ligase HUWE1. Recently, it was shown that poly-ubiquitylated histone H1 is an important signalling intermediate in the double strand break response. This poly-ubiquitylation is dependent on RNF8 and Ubc13 which extend pre-existi

Somatic insulin signaling regulates a germline starvation response in Drosophila egg chambers

AbstractEgg chambers from starved Drosophila females contain large aggregates of processing (P) bodies and cortically enriched microtubules. As this response to starvation is rapidly reversed upon re-feeding females or culturing egg chambers with exogenous bovine insulin, we examined the role of endogenous insulin signaling in mediating the starvation response. We found that systemic Drosophila insulin-like peptides (dILPs) activate the insulin pathway in follicle cells, which then regulate both microtubule and P body organization in the underlying germline cells. This organization is modulated by the motor proteins Dynein and Kinesin. Dynein activity is required for microtubule and P body organization during starvation, while Kinesin activity is required during nutrient-rich conditions. Blocking the ability of egg chambers to form P body aggregates in response to starvation correlated with reduced progeny survival. These data suggest a potential mechanism to maximize fecundity even during periods of poor nutrient availability, by mounting a protective response in immature egg chambers

A new chromatin flavor to cap chromosomes: Where structure, function, and evolution meet

Soman, A., Wong, S.Y., et al. find that telomeric DNA assembles into a new high-order chromatin structure resembling a columnar stack of nucleosomes with dynamic properties. This raises new questions on telomere biology mechanisms and chromatin evolution

The histone chaperone ANP32B regulates chromatin incorporation of the atypical human histone variant macroH2A

We would like to thank Carla Margulies, all members of the Ladurner and Mattiroli labs, and the Department of Physiological Chemistry for productive discussions. Thanks to Jan Dreyer, Inge Rondeel, and Rosanne van Hooijdonk for assistance with experiments. We thank the Bioimaging, Biophysics, and Flow Cytometry Core Facilities of the LMU Biomedical Center for training and use of their resources. We acknowledge the Protein Analytics Unit at the Biomedical Center, Ludwig-Maximilians University Munich (DFG, RI-00089), for providing services and assistance with data analysis. We thank the Histone Source at Colorado State University for the purification of human histones. pCAGGS-ANP32B was a kind gift from Wendy Barclay and pET29a-YS14 was a kind gift from Jung-Hyun Min (Addgene plasmid # 66890). pQCXIP-GFP1-10 was a gift from Yutaka Hata (Addgene plasmid # 68715) and pRK-flag-GFP11 from Yihong Ye (Addgene plasmid # 78590). This work was supported by funding from the Dutch Research Council (VI.Veni.212.052 to I.K.M.), the European Commission (ERC StG 851564 to F.M.; ERC StG 804182 to L.T.J.), the DFG (German Research Foundation) through Project-ID 213249687 - SFB 1064 and Project-ID 325871075 - SFB 1309, as well as LMU (to A.G.L.) and the national grant PID2021-126907NB-I00 from FEDER/Ministerio de Ciencia e Innovación (MCIN) - Agencia Estatal de Investigación and the Fundació La Marató de TV3 257/C/2019 (to M.B.).We would like to thank Carla Margulies, all members of the Ladurner and Mattiroli labs, and the Department of Physiological Chemistry for productive discussions. Thanks to Jan Dreyer, Inge Rondeel, and Rosanne van Hooijdonk for assistance with experiments. We thank the Bioimaging, Biophysics, and Flow Cytometry Core Facilities of the LMU Biomedical Center for training and use of their resources. We acknowledge the Protein Analytics Unit at the Biomedical Center, Ludwig-Maximilians University Munich (DFG, RI-00089), for providing services and assistance with data analysis. We thank the Histone Source at Colorado State University for the purification of human histones. pCAGGS-ANP32B was a kind gift from Wendy Barclay and pET29a-YS14 was a kind gift from Jung-Hyun Min (Addgene plasmid # 66890). pQCXIP-GFP1-10 was a gift from Yutaka Hata (Addgene plasmid # 68715) and pRK-flag-GFP11 from Yihong Ye (Addgene plasmid # 78590). This work was supported by funding from the Dutch Research Council (VI.Veni.212.052 to I.K.M.), the European Commission (ERC StG 851564 to F.M.; ERC StG 804182 to L.T.J.), the DFG (German Research Foundation) through Project-ID 213249687 - SFB 1064 and Project-ID 325871075 - SFB 1309, as well as LMU (to A.G.L.) and the national grant PID2021-126907NB-I00 from FEDER/Ministerio de Ciencia e Innovación (MCIN) - Agencia Estatal de Investigación and the Fundació La Marató de TV3 257/C/2019 (to M.B.). Conceptualization: I.K.M. and A.G.L.; methodology: I.K.M. C.R. F.M. E.F. and L.T.J.; investigation: I.K.M. E.F. D.C. and C.K.; resources: C.R.; writing - original draft: I.K.M.; writing - review and editing: F.M. and A.G.L.; supervision: L.T.J. F.M. M.B. and A.G.L.; funding acquisition: I.K.M. L.T.J. M.B. F.M. and A.G.L. A.G.L. is a founder, CSO, and managing director of Eisbach Bio GmbH, a biotechnology company in oncology and virology. We support inclusive, diverse, and equitable conduct of research.All vertebrate genomes encode for three large histone H2A variants that have an additional metabolite-binding globular macrodomain module, macroH2A. MacroH2A variants impact heterochromatin organization and transcription regulation and establish a barrier for cellular reprogramming. However, the mechanisms of how macroH2A is incorporated into chromatin and the identity of any chaperones required for histone deposition remain elusive. Here, we develop a split-GFP-based assay for chromatin incorporation and use it to conduct a genome-wide mutagenesis screen in haploid human cells to identify proteins that regulate macroH2A dynamics. We show that the histone chaperone ANP32B is a regulator of macroH2A deposition. ANP32B associates with macroH2A in cells and in vitro binds to histones with low nanomolar affinity. In vitro nucleosome assembly assays show that ANP32B stimulates deposition of macroH2A-H2B and not of H2A-H2B onto tetrasomes. In cells, depletion of ANP32B strongly affects global macroH2A chromatin incorporation, revealing ANP32B as a macroH2A histone chaperone

Measurements of nuclear level densities and gamma-ray strength functions and their interpretations

A method to extract primary $\gamma$-ray spectra from particle-$\gamma$ coincidences at excitation energies up to the neutron binding energy is described. From these spectra, the level density and $\gamma$-ray strength function can be determined. From the level density, several thermodynamical quantities are obtained within the microcanonical and canonical ensemble. Also models for the $\gamma$-ray strength function are discussed

MacroH2A histone variants limit chromatin plasticity through two distinct mechanisms

International audienceMacroH2A histone variants suppress tumor progression and act as epigenetic barriers to induced pluripotency. How they impart their influence on chromatin plasticity is not well understood. Here, we analyze how the different domains of macroH2A proteins contribute to chromatin structure and dynamics. By solving the crystal structure of the macrodomain of human macroH2A2 at 1.7 Å, we find that its putative binding pocket exhibits marked structural differences compared with the macroH2A1.1 isoform, rendering macroH2A2 unable to bind ADP-ribose. Quantitative binding assays show that this specificity is conserved among vertebrate macroH2A isoforms. We further find that macroH2A histones reduce the transient, PARP1-dependent chromatin relaxation that occurs in living cells upon DNA damage through two distinct mechanisms. First, macroH2A1.1 mediates an isoform-specific effect through its ability to suppress PARP1 activity. Second, the unstructured linker region exerts an additional repressive effect that is common to all macroH2A proteins. In the absence of DNA damage, the macroH2A linker is also sufficient for rescuing heterochromatin architecture in cells deficient for macroH2A