OR2M3: A highly specific and narrowly tuned human odorant receptor for the sensitive detection of onion key food odorant 3-Mercapto-2-methylpentan-1-ol

The detection of key food odorants appears to be an important capability of odorant receptors. Here, thiols occupy an outstanding position among the 230 known key food odorants because of their very low odor thresholds. Members of the homologous series of 3-mercapto-2-methylalkan-1-ols have been described as onion key food odorants or food constituents and are detected at logarithmically different thresholds. 3-Mercapto-2-methylpentan-1-ol being the only key food odorant within this series also has the lowest odor threshold. Most odorants typically activate combinations of odorant receptors, which may be narrowly or broadly tuned. Consequently, a specific receptor activation pattern will define an odor quality. In contrast, here we show that just 1 of the 391 human odorant receptors, OR2M3, responded exclusively to 3-mercapto-2-methylpentan-1-ol of the 190 key food odorants tested, with a half maximal effective concentration at submicromolar concentration. Moreover, neither the Denisovan OR2M3 nor the closest OR2M3 homologs from five species did respond to this compound. This outstanding specificity of extremely narrowly tuned human OR2M3 can explain both odor qualities and odor threshold trend within a homologous series of 3-mercapto-2-methylalkan-1-ols and suggests a modern human-specific, food-related function of OR2M3 in detecting a single onion key food odorant

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

A Poly-ADP-Ribose Trigger Releases the Auto-Inhibition of a Chromatin Remodeling Oncogene

International audienceDNA damage triggers chromatin remodeling by mechanisms that are poorly understood. The oncogene and chromatin remodeler ALC1/CHD1L massively decompacts chromatin in vivo yet is inactive prior to DNA-damage-mediated PARP1 induction. We show that the interaction of the ALC1 macrodomain with the ATPase module mediates auto-inhibition. PARP1 activation suppresses this inhibitory interaction. Crucially, release from auto-inhibition requires a poly-ADP-ribose (PAR) binding macrodomain. We identify tri-ADP-ribose as a potent PAR-mimic and synthetic allosteric effector that abrogates ATPase-macrodomain interactions, promotes an ungated conformation, and activates the remodeler’s ATPase. ALC1 fragments lacking the regulatory macrodomain relax chromatin in vivo without requiring PARP1 activation. Further, the ATPase restricts the macrodomain’s interaction with PARP1 under non-DNA damage conditions. Somatic cancer mutants disrupt ALC1’s auto-inhibition and activate chromatin remodeling. Our data show that the NAD+-metabolite and nucleic acid PAR triggers ALC1 to drive chromatin relaxation. Modular allostery in this oncogene tightly controls its robust, DNA-damage-dependent activation

Structural basis of histone H2A-H2B recognition by the essential chaperone FACT

Facilitates chromatin transcription (FACT) is a conserved histone chaperone that reorganizes nucleosomes and ensures chromatin integrity during DNA transcription, replication and repair(1-6). Key to the broad functions of FACT is its recognition of histones H2A-H2B (ref. 2). However, the structural basis for how histones H2A-H2B are recognized and how this integrates with the other functions of FACT, including the recognition of histones H3-H4 and other nuclear factors, is unknown. Here we reveal the crystal structure of the evolutionarily conserved FACT chaperone domain Spt16M from Chaetomium thermophilum, in complex with the H2A-H2B heterodimer. A novel 'U-turn' motif scaffolded onto a Rtt106-like module(7-10) embraces the alpha 1 helix of H2B. Biochemical and in vivo assays validate the structure and dissect the contribution of histone tails and H3-H4 towards Spt16M binding. Furthermore, we report the structure of the FACT heterodimerization domain that connects FACT to replicative polymerases. Our results show that Spt16M makes several interactions with histones, which we suggest allow the module to invade the nucleosome gradually and block the strongest interaction of H2B with DNA. FACT would thus enhance 'nucleosome breathing' by re-organizing the first 30 base pairs of nucleosomal histone-DNA contacts. Our snapshot of the engagement of the chaperone with H2A-H2B and the structures of all globular FACT domains enable the high-resolution analysis of the vital chaperoning functions of FACT, shedding light on how the complex promotes the activity of enzymes that require nucleosome reorganization