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The structure of the METTL3-METTL14 complex, which mediates N.sup.6-adenosine methylation of RNA, suggests that the METTL3 subunit is the catalytic core while METTL14 serves to bind RNA. A window on m.sup.6A epitranscriptomics The various base modifications now known to occur in messenger RNA and long non-coding RNA are reversible, and are utilized to dynamically modify the function of the RNA. The N.sup.6-methyladenosine modification is removed by an enzyme complex comprising METTL3 and METTL14. Ping Yin and colleagues have solved structures of the methyltransferase domains of this heterodimeric complex with and without ligand. Surprisingly, the S-adenosyl methionine ligand was found only the METTL3 pocket, not in METTL14. This suggests a model in which there is a single catalytic subunit, with METTL3 functioning as an RNA binding platform. The reported structures provide unprecedented mechanistic insight into m.sup.6A RNA methylation and suggest new opportunities for the development of therapeutic agents. Chemical modifications of RNA have essential roles in a vast range of cellular processes.sup.1,2,3. N.sup.6-methyladenosine (m.sup.6A) is an abundant internal modification in messenger RNA and long non-coding RNA that can be dynamically added and removed by RNA methyltransferases (MTases) and demethylases, respectively.sup.2,3,4,5. An MTase complex comprising methyltransferase-like 3 (METTL3) and methyltransferase-like 14 (METTL14) efficiently catalyses methyl group transfer.sup.6,7. In contrast to the well-studied DNA MTase.sup.8, the exact roles of these two RNA MTases in the complex remain to be elucidated. Here we report the crystal structures of the METTL3-METTL14 heterodimer with MTase domains in the ligand-free, S-adenosyl methionine (AdoMet)-bound and S-adenosyl homocysteine (AdoHcy)-bound states, with resolutions of 1.9, 1.71 and 1.61 Å, respectively. Both METTL3 and METTL14 adopt a class I MTase fold and they interact with each other via an extensive hydrogen bonding network, generating a positively charged groove. Notably, AdoMet was observed in only the METTL3 pocket and not in METTL14. Combined with biochemical analysis, these results suggest that in the m.sup.6A MTase complex, METTL3 primarily functions as the catalytic core, while METTL14 serves as an RNA-binding platform, reminiscent of the target recognition domain of DNA N.sup.6-adenine MTase.sup.9,10. This structural information provides an important framework for the functional investigation of m.sup.6A., Author(s): Xiang Wang [sup.1] , Jing Feng [sup.1] , Yuan Xue [sup.1] , Zeyuan Guan [sup.1] , Delin Zhang [sup.1] , Zhu Liu [sup.2] , Zhou Gong [sup.3] , Qiang [...]

Author(s): Xiang Wang; Jing Feng; Yuan Xue; Zeyuan Guan; Delin Zhang; Zhu Liu; Zhou Gong; Qiang Wang; Jinbo Huang; Chun Tang; Tingting Zou; Ping Yin Nature 534, 575578 (2016);doi:10.1038/nature18298 In [...]

N 6 -methyldeoxyadenine (6mA) has recently been reported as a prevalent DNA modification in eukaryotes. The Tetrahymena thermophila MTA1 complex consisting of four subunits, namely MTA1, MTA9, p1, and p2, is the first identified eukaryotic 6mA methyltransferase (MTase) complex. Unlike the prokaryotic 6mA MTases which have been biochemically and structurally characterized, the operation mode of the MTA1 complex remains largely elusive. Here, we report the cryogenic electron microscopy structures of the quaternary MTA1 complex in S-adenosyl methionine (SAM)-bound (2.6 Å) and S-adenosyl homocysteine (SAH)-bound (2.8 Å) states. Using an AI-empowered integrative approach based on AlphaFold prediction and chemical cross-linking mass spectrometry, we further modeled a near-complete structure of the quaternary complex. Coupled with biochemical characterization, we revealed that MTA1 serves as the catalytic core, MTA1, MTA9, and p1 likely accommodate the substrate DNA, and p2 may facilitate the stabilization of MTA1. These results together offer insights into the molecular mechanism underpinning methylation by the MTA1 complex and the potential diversification of MTases for N 6 -adenine methylation., (© 2023. The Author(s).)

First discovered in the 1980s, retrons are bacterial genetic elements consisting of a reverse transcriptase and a non-coding RNA (ncRNA). Retrons mediate antiphage defence in bacteria but their structure and defence mechanisms are unknown. Here, we investigate the Escherichia coli Ec86 retron and use cryo-electron microscopy to determine the structures of the Ec86 (3.1 Å) and cognate effector-bound Ec86 (2.5 Å) complexes. The Ec86 reverse transcriptase exhibits a characteristic right-hand-like fold consisting of finger, palm and thumb subdomains. Ec86 reverse transcriptase reverse-transcribes part of the ncRNA into satellite, multicopy single-stranded DNA (msDNA, a DNA-RNA hybrid) that we show wraps around the reverse transcriptase electropositive surface. In msDNA, both inverted repeats are present and the 3' sides of the DNA/RNA chains are close to the reverse transcriptase active site. The Ec86 effector adopts a two-lobe fold and directly binds reverse transcriptase and msDNA. These findings offer insights into the structure-function relationship of the retron-effector unit and provide a structural basis for the optimization of retron-based genome editing systems., (© 2022. The Author(s), under exclusive licence to Springer Nature Limited.)

A newly identified arbitrium communication system regulates the lysis-to-lysogeny decision in a Bacillus bacteriophage. This system contains an arbitrium hexapeptide as a signal, the cellular receptor AimR, and the lysogenic negative regulator AimX. AimR specifically targets the downstream DNA to activate aimX gene expression. The arbitrium peptide binds to AimR, inhibiting its DNA-binding to promote phage lysogeny. Recently, we and other groups have elucidated how arbitrium peptide sensed by AimR. However, the molecular mechanisms of DNA recognition by AimR and the regulation of its DNA-binding activity by the peptide remain largely unknown. Here, we report the crystal structure of the AimR-DNA complex at 2.1 Å resolution. The N-terminal HTH motif recognizes the palindromic DNA sequence, buttressed by interactions between positively charged residues and the DNA phosphate groups. The DNA-bound AimR assembles a more closed dimer than the peptide-bound form. Single-molecule FRET and crosslinking assays revealed that the AimR protein samples both open and closed conformations in solution. Arbitrium peptide binding induces a closed-to-open conformational change of AimR, eliminating DNA targeting. Our structural and functional analysis provides new insights into the DNA recognition mechanism of AimR and its regulation by the arbitrium peptide in the context of phage lysis-lysogeny decisions., Competing Interests: Conflict of interestThe authors declare that they have no conflict of interest.

Perpendicular magnetic materials with low damping constant and high thermal stability have great potential for realizing high-density, non-volatile, and low-power consumption spintronic devices, which can sustain operation reliability for high processing temperatures. In this work, we study the Gilbert damping constant (α) of perpendicularly magnetized W/CoFeB/MgO films with a high perpendicular magnetic anisotropy (PMA) and superb thermal stability. The α of these PMA films annealed at different temperatures (T ann ) is determined via an all-optical Time-Resolved Magneto-Optical Kerr Effect method. We find that α of these W/CoFeB/MgO PMA films decreases with increasing T ann , reaches a minimum of α = 0.015 at T ann  = 350 °C, and then increases to 0.020 after post-annealing at 400 °C. The minimum α observed at 350 °C is rationalized by two competing effects as T ann becomes higher: the enhanced crystallization of CoFeB and dead-layer growth occurring at the two interfaces of the CoFeB layer. We further demonstrate that α of the 400 °C-annealed W/CoFeB/MgO film is comparable to that of a reference Ta/CoFeB/MgO PMA film annealed at 300 °C, justifying the enhanced thermal stability of the W-seeded CoFeB films.

WD40 proteins belong to a big protein family with members identified in every eukaryotic proteome. However, WD40 proteins were only reported in a few prokaryotic proteomes. Using WDSP ( http://wu.scbb.pkusz.edu.cn/wdsp/ ), a prediction tool, we identified thousands of prokaryotic WD40 proteins, among which few proteins have been biochemically characterized. As shown in our previous bioinformatics study, a large proportion of prokaryotic WD40 proteins have higher intramolecular sequence identity among repeats and more hydrogen networks, which may indicate better stability than eukaryotic WD40s. Here we report our biophysical and structural study on the WD40 domain of PkwA from Thermomonospora curvata (referred as tPkwA-C). We demonstrated that the stability of thermophilic tPkwA-C correlated to ionic strength and tPkwA-C exhibited fully reversible unfolding under different denaturing conditions. Therefore, the folding kinetics was also studied through stopped-flow circular dichroism spectra. The crystal structure of tPkwA-C was further resolved and shed light on the key factors that stabilize its beta-propeller structure. Like other WD40 proteins, DHSW tetrad has a significant impact on the stability of tPkwA-C. Considering its unique features, we proposed that tPkwA-C should be a great structural template for protein engineering to study key residues involved in protein-protein interaction of a WD40 protein.