Exploring a unique class of flavoenzymes: Identification and biochemical characterization of ribosomal RNA dihydrouridine synthase.

Dihydrouridine (D), a prevalent and evolutionarily conserved base in the transcriptome, primarily resides in tRNAs and, to a lesser extent, in mRNAs. Notably, this modification is found at position 2449 in the 23S rRNA, strategically positioned near the ribosome's peptidyl transferase site. Despite the prior identification, in genome, of three dihydrouridine synthases (DUS), a set of NADPH and FMN-dependent enzymes known for introducing D in tRNAs and mRNAs, characterization of the enzyme responsible for D2449 deposition has remained elusive.

Functional redundancy in tRNA dihydrouridylation.

Dihydrouridine (D) is a common modified base found predominantly in transfer RNA (tRNA). Despite its prevalence, the mechanisms underlying dihydrouridine biosynthesis, particularly in prokaryotes, have remained elusive. Here, we conducted a comprehensive investigation into D biosynthesis in Bacillus subtilis through a combination of genetic, biochemical, and epitranscriptomic approaches.

Poly(A) tail degradation in human cells: ATF4 mRNA as a model for biphasic deadenylation.

Eukaryotic mRNA deadenylation is generally considered as a two-step process in which the PAN2-PAN3 complex initiates the poly(A) tail degradation while, in the second step, the CCR4-NOT complex completes deadenylation, leading to decapping and degradation of the mRNA body. However, the mechanism of the biphasic poly(A) tail deadenylation remains enigmatic in several points such as the timing of the switch between the two steps, the role of translation termination and the mRNAs population involved. Here, we have studied the deadenylation of endogenous mRNAs in human cells depleted in either PAN3 or translation termination factor eRF3.

Divergent effects of translation termination factor eRF3A and nonsense-mediated mRNA decay factor UPF1 on the expression of uORF carrying mRNAs and ribosome protein genes.

In addition to its role in translation termination, eRF3A has been implicated in the nonsense-mediated mRNA decay (NMD) pathway through its interaction with UPF1. NMD is a RNA quality control mechanism, which detects and degrades aberrant mRNAs as well as some normal transcripts including those that harbour upstream open reading frames in their 5' leader sequence. In this study, we used RNA-sequencing and ribosome profiling to perform a genome wide analysis of the effect of either eRF3A or UPF1 depletion in human cells.

C3P3-G1: first generation of a eukaryotic artificial cytoplasmic expression system.

Most eukaryotic expression systems make use of host-cell nuclear transcriptional and post-transcriptional machineries. Here, we present the first generation of the chimeric cytoplasmic capping-prone phage polymerase (C3P3-G1) expression system developed by biological engineering, which generates capped and polyadenylated transcripts in host-cell cytoplasm by means of two components. First, an artificial single-unit chimeric enzyme made by fusing an mRNA capping enzyme and a DNA-dependent RNA polymerase.