Epitranscriptomic mapping of RNA modifications at single-nucleotide resolution using rhodamine sequencing (Rho-seq)

Finet, Olivier; Yague-Sanz, Carlo; Hermand, Damien

doi:10.1016/j.xpro.2022.101369

Cited by 5 publications

(4 citation statements)

References 32 publications

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“…Newly developed high-resolution mapping methods established the presence of D within mRNAs, which further verified its physiological significance. 14 , 23 , 24 D-seq reported that certain (not all) mRNA D sites occur in regions where secondary structure potential is evolutionarily conserved, suggesting biological function possibly analogous to D sites on tRNAs. 23 Another question is raised concerning how D on mRNA involves translation.…”

Section: Discussionmentioning

confidence: 99%

“…Furthermore, wet-lab experimental methods such as D sequencing (D-seq) and rhodamine sequencing (Rho-seq) established the presence of D on mRNAs in yeast and human, which further verified its physiological significance. 14 , 23 , 24 Both methods take advantage of D-specific chemistry and next-generation sequencing to determine the location of D across the transcriptome, resulting in high-confidence single-nucleotide-resolution data.…”

Section: Introductionmentioning

confidence: 99%

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Self-attention enabled deep learning of dihydrouridine (D) modification on mRNAs unveiled a distinct sequence signature from tRNAs

Wang¹,

Wang²,

Cui³

et al. 2023

Molecular Therapy - Nucleic Acids

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Self-attention enabled deep learning of dihydrouridine (D) modification on mRNAs unveiled a distinct sequence signature from tRNAs

Wang¹,

Wang²,

Cui³

et al. 2023

Molecular Therapy - Nucleic Acids

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“…From there, the Rho-seq protocol unfolds as follows; (I) RNA extraction from WT and ∆4 strains (lacking the four dus genes), (II) Rho+ and mock-treatments, (III) ribodepletion, (IV) cDNA synthesis and library preparation, (V) strand-specific deep-sequencing, (VI) data analysis by implementation of a multifactorial analysis. The detailed protocol is available in a separate publication [ 109 ].…”

Section: New High-resolution Methods To Decipher the Transcriptome-wi...mentioning

confidence: 99%

The Dihydrouridine landscape from tRNA to mRNA: a perspective on synthesis, structural impact and function

et al. 2022

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The universal dihydrouridine (D) epitranscriptomic mark results from a reduction of uridine by the Dus family of NADPH-dependent reductases and is typically found within the eponym D-loop of tRNAs. Despite its apparent simplicity, D is structurally unique, with the potential to deeply affect the RNA backbone and many, if not all, RNA-connected processes. The first landscape of its occupancy within the tRNAome was reported 20 years ago. Its potential biological significance was highlighted by observations ranging from a strong bias in its ecological distribution to the predictive nature of Dus enzymes overexpression for worse cancer patient outcomes. The exquisite specificity of the Dus enzymes revealed by a structure-function analyses and accumulating clues that the D distribution may expand beyond tRNAs recently led to the development of new high-resolution mapping methods, including Rho-seq that established the presence of D within mRNAs and led to the demonstration of its critical physiological relevance.

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“…To accomplish this, the septum-synthesizing enzyme Bgs1 must be delivered to the division site. At the same time, new membrane material is delivered to the division site for the development of furrows [23,24].…”

mentioning

confidence: 99%

A novel cell biological tool to explain mechanics and dynamics in fission yeast

Rasheed,

Mohy‐ud‐Din,

Anwar

et al. 2024

J Basic Microbiol

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The Rho guanosine triphosphatase hydrolase enzyme (GTPase) is required for the control of the actin cytoskeleton, but its activation in vivo condition is unknown. The study's goal was to find a new synthetic nanobody VHH (P‐36 tagged with mNeonGreen) that interacts strongly with the Rho GTPase. We present the first novel synthetic nanobody, VHH (P‐36 tagged with mNeonGreen), tested in fission yeast cells and found to have a particular interaction with Rho1GTPase. Plasmids were constructed by using of certain enzymes to digest the pDUAL‐pef1a vector plasmid to produce a protein that was encoded by cloned genes. A varied VHH library was created synthetically, then transformed into yeast cells, and positive clones were chosen using chemical agents. To investigate protein interactions and cellular reactions, several studies were carried out, such as live cell imaging, growth curve analysis, coimmunoprecipitation, structural analysis, and cell therapies. Prism and RStudio were used for the statistical analysis. The presence of VHH (P‐36) has no effect on the growth pattern making it an appropriate model for studying cytokinesis in vivo. According to a computational biological study, its affinity to interact with Rho1GTPase with all the complementarity‐determining region (CDR) regions found on VHH (P‐36) is extremely strong. We were able to track its subcellular target by localization using a fluorescent confocal microscope, ensuring the maintenance of cell polarity and morphology. Spheroplast analysis revealed a circular‐shaped cell with an even distribution of Rho1 tagged VHH (P‐36), indicating that the interaction occurs near the plasma membrane. The introduction of latrunculin‐A (Lat‐A) disrupted Rho GTPase localization, demonstrating the control over actin production, and the cell did not show evidence of mitotic phase commencement while Lat‐A was present. Finally, this important biological tool can aid in our understanding of the mechanics and dynamics of cytokinesis in relation to Rho1GTPase.

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Epitranscriptomic mapping of RNA modifications at single-nucleotide resolution using rhodamine sequencing (Rho-seq)

Cited by 5 publications

References 32 publications

Self-attention enabled deep learning of dihydrouridine (D) modification on mRNAs unveiled a distinct sequence signature from tRNAs

Self-attention enabled deep learning of dihydrouridine (D) modification on mRNAs unveiled a distinct sequence signature from tRNAs

The Dihydrouridine landscape from tRNA to mRNA: a perspective on synthesis, structural impact and function

A novel cell biological tool to explain mechanics and dynamics in fission yeast

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