The RNA acetyltransferase driven by ATP hydrolysis synthesizes N4-acetylcytidine of tRNA anticodon

The wobble base of Escherichia coli elongator tRNAMet is modified to N4-acetylcytidine (ac4C), which is thought to ensure the precise recognition of the AUG codon by preventing misreading of near-cognate AUA codon. By employing genome-wide screen of uncharacterized genes in Escherichia coli (‘ribonucleome analysis'), we found the ypfI gene, which we named tmcA (tRNAMet cytidine acetyltransferase), to be responsible for ac4C formation. TmcA is an enzyme that contains a Walker-type ATPase domain in its N-terminal region and an N-acetyltransferase domain in its C-terminal region. Recombinant TmcA specifically acetylated the wobble base of E. coli elongator tRNAMet by utilizing acetyl-coenzyme A (CoA) and ATP (or GTP). ATP/GTP hydrolysis by TmcA is stimulated in the presence of acetyl-CoA and tRNAMet. A mutation study revealed that E. coli TmcA strictly discriminates elongator tRNAMet from the structurally similar tRNAIle by mainly recognizing the C27–G43 pair in the anticodon stem. Our findings reveal an elaborate mechanism embedded in tRNAMet and tRNAIle for the accurate decoding of AUA/AUG codons on the basis of the recognition of wobble bases by the respective RNA-modifying enzymes

Genotype–phenotype associations in 1018 individuals with SCN1A‐related epilepsies

Objective: SCN1A variants are associated with epilepsy syndromes ranging from mild genetic epilepsy with febrile seizures plus (GEFS+) to severe Dravet syndrome (DS). Many variants are de novo, making early phenotype prediction difficult, and genotype–phenotype associations remain poorly understood. Methods: We assessed data from a retrospective cohort of 1018 individuals with SCN1A-related epilepsies. We explored relationships between variant characteristics (position, in silico prediction scores: Combined Annotation Dependent Depletion (CADD), Rare Exome Variant Ensemble Learner (REVEL), SCN1A genetic score), seizure characteristics, and epilepsy phenotype. Results: DS had earlier seizure onset than other GEFS+ phenotypes (5.3 vs. 12.0 months, p < .001). In silico variant scores were higher in DS versus GEFS+ (p < .001). Patients with missense variants in functionally important regions (conserved N-terminus, S4–S6) exhibited earlier seizure onset (6.0 vs. 7.0 months, p = .003) and were more likely to have DS (280/340); those with missense variants in nonconserved regions had later onset (10.0 vs. 7.0 months, p = .036) and were more likely to have GEFS+ (15/29, χ2 = 19.16, p < .001). A minority of protein-truncating variants were associated with GEFS+ (10/393) and more likely to be located in the proximal first and last exon coding regions than elsewhere in the gene (9.7% vs. 1.0%, p < .001). Carriers of the same missense variant exhibited less variability in age at seizure onset compared with carriers of different missense variants for both DS (1.9 vs. 2.9 months, p = .001) and GEFS+ (8.0 vs. 11.0 months, p = .043). Status epilepticus as presenting seizure type is a highly specific (95.2%) but nonsensitive (32.7%) feature of DS. Significance: Understanding genotype–phenotype associations in SCN1A-related epilepsies is critical for early diagnosis and management. We demonstrate an earlier disease onset in patients with missense variants in important functional regions, the occurrence of GEFS+ truncating variants, and the value of in silico prediction scores. Status epilepticus as initial seizure type is a highly specific, but not sensitive, early feature of DS

Genome-wide identification and phenotypic characterization of seizure-associated copy number variations in 741,075 individuals

Copy number variants (CNV) are established risk factors for neurodevelopmental disorders with seizures or epilepsy. With the hypothesis that seizure disorders share genetic risk factors, we pooled CNV data from 10,590 individuals with seizure disorders, 16,109 individuals with clinically validated epilepsy, and 492,324 population controls and identified 25 genome-wide significant loci, 22 of which are novel for seizure disorders, such as deletions at 1p36.33, 1q44, 2p21-p16.3, 3q29, 8p23.3-p23.2, 9p24.3, 10q26.3, 15q11.2, 15q12-q13.1, 16p12.2, 17q21.31, duplications at 2q13, 9q34.3, 16p13.3, 17q12, 19p13.3, 20q13.33, and reciprocal CNVs at 16p11.2, and 22q11.21. Using genetic data from additional 248,751 individuals with 23 neuropsychiatric phenotypes, we explored the pleiotropy of these 25 loci. Finally, in a subset of individuals with epilepsy and detailed clinical data available, we performed phenome-wide association analyses between individual CNVs and clinical annotations categorized through the Human Phenotype Ontology (HPO). For six CNVs, we identified 19 significant associations with specific HPO terms and generated, for all CNVs, phenotype signatures across 17 clinical categories relevant for epileptologists. This is the most comprehensive investigation of CNVs in epilepsy and related seizure disorders, with potential implications for clinical practice

Prenylated proteins: demonstration of a thioether linkage to cysteine of proteins.

Prenylated amino acid fragments have been isolated from prenylated proteins of Chinese hamster ovary cells. Gel-exclusion chromatography indicates that these proteins are modified by two different prenyl groups. The prenyl-amino acid fragments are labeled by 35S from cysteine, and this bond is cleaved by Raney-Ni, proving that the prenyl residue is linked to protein via a thioether to cysteine. Hydrazinolysis has been used to demonstrate that the cysteine is carboxy terminal

Nanopatterning of magnetic disks by single-step Ar+ Ion projection

Large-area Ar+ projection has been used to generate planar magnetic nanostructures on a 1¿-format hard disk in a single step (see Figure). The recording pattern was transferred to a Co/Pt multilayer without resist processes or any other contact to the delicate media surface. It is conceivable that magnetic nanostructures can be replicated from a single stencil mask on large quantities of disks for future high-density recording

Ion projection direct structuring for patterning of magnetic media

Ion projection facilitates a direct structuring, which is an attractive potential manufacturing process for patterned storage media. An advantage to this method is that the media roughness remains unchanged. The feasibility of ion projection direct structuring for processing full disk surfaces was investigated using a next generation lithography projector. Co-Pt multilayer films with strong perpendicular anisotropy were deposited on 1-in glass disks as used in the IBM microdrive and on Si substrates. Concentric tracks including data, as well as head positioning servo structures, were patterned in a single exposure step with 45 keV He+ at a 4 × demagnification. In a second experiment, sub-100-nm magnetic islands were produced using projection at 8.7 × demagnification and visualized by magnetic force microscopy