Histone H1 regulates non-coding RNA turnover on chromatin in a m6A-dependent manner

Linker histones are highly abundant chromatin-associated proteins with well-established structural roles in chromatin and as general transcriptional repressors. In addition, it has been long proposed that histone H1 exerts context-specific effects on gene expression. Here, we identify a function of histone H1 in chromatin structure and transcription using a range of genomic approaches. In the absence of histone H1, there is an increase in the transcription of non-coding RNAs, together with reduced levels of m6A modification leading to their accumulation on chromatin and causing replication-transcription conflicts. This strongly suggests that histone H1 prevents non-coding RNA transcription and regulates non-coding transcript turnover on chromatin. Accordingly, altering the m6A RNA methylation pathway rescues the replicative phenotype of H1 loss. This work unveils unexpected regulatory roles of histone H1 on non-coding RNA turnover and m6A deposition, highlighting the intimate relationship between chromatin conformation, RNA metabolism, and DNA replication to maintain genome performance.Work at the M.G. lab was supported by the Spanish Ministry of Sciences and Innovation (BFU2016-78849-P and PID2019-105949GB-I00, co-financed by the European Union FEDER funds), a CSIC grant (2019AEP004), and a Salvador de Madariaga mobility grant (PRX19/00293). J.M.F.-J., C.S.-M., and J.I.-A. were supported by the Spanish Ministry of Sciences and Innovation fellowships (BES-2014-070050, BES-2017-079897, and PRE2020-095071, respectively); S.M.-V. was supported by a predoctoral fellowship from the Spanish Ministry of Education and Universities (FPU18/04794); and M.S.-P. was supported by an AGAUR-FI predoctoral fellowship co-financed by Generalitat de Catalunya and the European Social Fund. A.J. was supported by the Spanish Ministry of Sciences and Innovation (BFU2017-82805-C2-1-P and PID2020-112783GB-C21) and J.F.C. by core funding to the MRC Human Genetics Unit from the Medical Research Council (UK)

Remote sensing techniques applied to seismic vulnerability assessment

Advances in remote sensing and photogrammetry techniques have increased the degree of accuracy and resolution in the record of the earth’s surface. This has expanded the range of possible applications of these data. In this research, we have used these data to document the construction characteristics of the urban environment of Lorca, Spain. An exposure database has been created with the gathered information to be used in seismic vulnerability assessment. To this end, we have used data from photogrammetric flights at different periods, using both orthorectified images in the visible and infrared spectrum. Furthermore, the analysis is completed using LiDAR data. From the combination of these data, it has been possible to delineate the building footprints and characterize the constructions with attributes such as the approximate date of construction, area, type of roof and even building materials. To carry out the calculation, we have developed different algorithms to compare images from different times, segment images, classify LiDAR data, and use the infrared data in order to remove vegetation or to compute roof surfaces with height value, tilt and spectral fingerprint. In addition, the accuracy of our results has been validated with ground truth data. This work is developed in the context of the MERISUR project (ref. CGL2013-40492-R), with funding from the Spanish Ministry of Economy and Competitiveness

Remote sensing techniques applied to seismic vulnerability assessment

Advances in remote sensing and photogrammetry techniques have increased the degree of accuracy and resolution in the record of the earth’s surface. This has expanded the range of possible applications of these data. In this research, we have used these data to document the construction characteristics of the urban environment of Lorca, Spain. An exposure database has been created with the gathered information to be used in seismic vulnerability assessment. To this end, we have used data from photogrammetric flights at different periods, using both orthorectified images in the visible and infrared spectrum. Furthermore, the analysis is completed using LiDAR data. From the combination of these data, it has been possible to delineate the building footprints and characterize the constructions with attributes such as the approximate date of construction, area, type of roof and even building materials. To carry out the calculation, we have developed different algorithms to compare images from different times, segment images, classify LiDAR data, and use the infrared data in order to remove vegetation or to compute roof surfaces with height value, tilt and spectral fingerprint. In addition, the accuracy of our results has been validated with ground truth data. This work is developed in the context of the MERISUR project (ref. CGL2013-40492-R), with funding from the Spanish Ministry of Economy and Competitiveness

Slow RNAPII Transcription Elongation Rate, Low Levels of RNAPII Pausing, and Elevated Histone H1 Content at Promoters Associate with Higher m6A Deposition on Nascent mRNAs

N6-methyladenosine modification (m6A) fine-tunes RNA fate in a variety of ways, thus regulating multiple fundamental biological processes. m6A writers bind to chromatin and interact with RNA polymerase II (RNAPII) during transcription. To evaluate how the dynamics of the transcription process impact m6A deposition, we studied RNAPII elongation rates in mouse embryonic stem cells with altered chromatin configurations, due to reductions in linker histone H1 content. We found that genes transcribed at slow speed are preferentially methylated and display unique signatures at their promoter region, namely high levels of histone H1, together with marks of bivalent chromatin and low RNAPII pausing. They are also highly susceptible to m6A loss upon histone H1 reduction. These results indicate that RNAPII velocity links chromatin structure and the deposition of m6A, highlighting the intricate relationship between different regulatory layers on nascent mRNA molecules

Ferredoxin-linked flavoenzyme defines a family of pyridine nucleotide-independent thioredoxin reductases

Ferredoxin-dependent thioredoxin reductase was identified 35 y ago in the fermentative bacterium Clostridium pasteurianum [Hammel KE, Cornwell KL, Buchanan BB (1983) Proc Natl Acad Sci USA 80:3681-3685]. The enzyme, a flavoprotein, was strictly dependent on ferredoxin as reductant and was inactive with either NADPH or NADH. This early work has not been further pursued. We have recently reinvestigated the problem and confirmed that the enzyme, here designated ferredoxin-dependent flavin thioredoxin reductase (FFTR), is a flavoprotein. The enzyme differs from ferredoxin-thioredoxin reductase (FTR), which has a signature [4Fe-4S] cluster, but shows structural similarities to NADP-dependent thioredoxin reductase (NTR). Comparative amino acid sequence analysis showed that FFTR is present in a number of clostridial species, some of which lack both FTR and an archetypal NTR. We have isolated, crystallized, and determined the structural properties of FFTR from a member of this group, Clostridium acetobutylicum, both alone and in complex with Trx. The structures showed an elongated FFTR homodimer, each monomer comprising two Rossmann domains and a noncovalently bound FAD cofactor that exposes the isoalloxazine ring to the solvent. The FFTR structures revealed an alternative domain organization compared with NTR that enables the enzyme to accommodate Fdx rather than NADPH. The results suggest that FFTR exists in a range of conformations with varying degrees of domain separation in solution and that the stacking between the two redox-active groups for the transfer of reducing equivalents results in a profound structural reorganization. A mechanism in accord with the findings is proposed

A nucleotide-controlled conformational switch modulates the activity of eukaryotic IMP dehydrogenases

Inosine-5′-monophosphate dehydrogenase (IMPDH) is an essential enzyme for nucleotide metabolism and cell proliferation. Despite IMPDH is the target of drugs with antiviral, immunosuppressive and antitumor activities, its physiological mechanisms of regulation remain largely unknown. Using the enzyme from the industrial fungus Ashbya gossypii, we demonstrate that the binding of adenine and guanine nucleotides to the canonical nucleotide binding sites of the regulatory Bateman domain induces different enzyme conformations with significantly distinct catalytic activities. Thereby, the comparison of their high-resolution structures defines the mechanistic and structural details of a nucleotide-controlled conformational switch that allosterically modulates the catalytic activity of eukaryotic IMPDHs. Remarkably, retinopathy-associated mutations lie within the mechanical hinges of the conformational change, highlighting its physiological relevance. Our results expand the mechanistic repertoire of Bateman domains and pave the road to new approaches targeting IMPDHs

Ferredoxin-linked flavoenzyme defines a family of pyridine nucleotide-independent thioredoxin reductases

Ferredoxin-dependent thioredoxin reductase was identified 35 y ago in the fermentative bacterium Clostridium pasteurianum [Hammel KE, Cornwell KL, Buchanan BB (1983) Proc Natl Acad Sci USA 80:3681–3685]. The enzyme, a flavoprotein, was strictly dependent on ferredoxin as reductant and was inactive with either NADPH or NADH. This early work has not been further pursued. We have recently reinvestigated the problem and confirmed that the enzyme, here designated ferredoxin-dependent flavin thioredoxin reductase (FFTR), is a flavoprotein. The enzyme differs from ferredoxin−thioredoxin reductase (FTR), which has a signature [4Fe−4S] cluster, but shows structural similarities to NADP-dependent thioredoxin reductase (NTR). Comparative amino acid sequence analysis showed that FFTR is present in a number of clostridial species, some of which lack both FTR and an archetypal NTR. We have isolated, crystallized, and determined the structural properties of FFTR from a member of this group, Clostridium acetobutylicum, both alone and in complex with Trx. The structures showed an elongated FFTR homodimer, each monomer comprising two Rossmann domains and a noncovalently bound FAD cofactor that exposes the isoalloxazine ring to the solvent. The FFTR structures revealed an alternative domain organization compared with NTR that enables the enzyme to accommodate Fdx rather than NADPH. The results suggest that FFTR exists in a range of conformations with varying degrees of domain separation in solution and that the stacking between the two redox-active groups for the transfer of reducing equivalents results in a profound structural reorganization. A mechanism in accord with the findings is proposed.This work was supported by Spanish Ministerio de Ciencia, Innovación y Universidades Grants BFU2016-80343-P and BIO2016-75634-P. The research leading to these results received funding from the European Community’s Seventh Framework Program FP7/2007–2013 under BioStruct-X Grant Agreement 7687

GuaB3, an overlooked enzyme in cyanobacteria's toolbox that sheds light on IMP dehydrogenase evolution

14 páginas, 1 tabla, 4 figurasIMP dehydrogenase and GMP reductase are enzymes from the same protein family with analogous structures and catalytic mechanisms that have gained attention because of their essential roles in nucleotide metabolism and as potential drug targets. This study focusses on GuaB3, a less-explored enzyme within this family. Phylogenetic analysis uncovers GuaB3's independent evolution from other members of the family and it predominantly occurs in Cyanobacteria. Within this group, GuaB3 functions as a unique IMP dehydrogenase, while its counterpart in Actinobacteria has a yet unknown function. Synechocystis sp. PCC6803 GuaB3 structures demonstrate differences in the active site compared to canonical IMP dehydrogenases, despite shared catalytic mechanisms. These findings highlight the essential role of GuaB3 in Cyanobacteria, provide insights into the diversity and evolution of the IMP dehydrogenase protein family, and reveal a distinctive characteristic in nucleotide metabolism, potentially aiding in combating harmful cyanobacterial blooms-a growing concern for humans and wildlife.This work was funded by the Spanish Ministerio de Ciencia e Innovación-FEDER-Fondo Social Europeo (grants PID2019-109671GB-I00 to Rubén M. Buey, PID2019-110900GB-I00 to Mónica Balsera, and PID2020-118200RBI00 to José Luis Revuelta and Alberto Jiménez). David Fernández-Justel was supported by a pre-doctoral contract from the Junta de Castilla y León. We gratefully acknowledge Paulino Gómez-Puertas for his critical reading of the manuscript and performing supplementary experiments, not covered in the manuscript, that enhanced the comprehensiveness of our findings. We also thank Silvia Domínguez and Marta Santos for excellent technical support. Protein crystallography experiments were performed at the XALOC beamline at ALBA synchrotron with the collaboration and support of the ALBA staff. The atomic coordinates and the structure factors described in this work have been deposited in the Research Collaboratory for Structural Bioinformatics Protein Data Bank under the PDB IDs: 8P4Q and 8P37Peer reviewe