Biology subjects

Koh, C. W. Q.

Publications and source records attributed to Koh, C. W. Q..

2 recordsLinked to original sources

bioRxiv2020

Detection of differential RNA modifications from direct RNA sequencing of human cell lines

Differences in RNA expression can provide insights into the molecular identity of a cell, pathways involved in human diseases, and variation in RNA levels across patients associated with clinical phenotypes. RNA modifications such as m6A have been found to contribute to molecular functions of RNAs. However, quantification of differences in RNA modifications has been challenging. Here we develop a computational method (xPore) to identify differential RNA modifications from direct RNA sequencing data. We evaluate our method on transcriptome-wide m6A profiling data, demonstrating that xPore identifies positions of m6A sites at single base resolution, estimates the fraction of modified RNAs in the cell, and quantifies the differential modification rate across conditions. We apply the method to direct RNA-Sequencing data from 6 cell lines and find that many m6A sites are preserved, while a subset of m6A sites show significant differences in their modification rates across cell types. Together, we show that RNA modifications can be identified from direct RNA-sequencing with high accuracy, enabling the analysis of differential modifications and expression from a single high throughput experiment. AvailabilityxPore is available as open source software (https://github.com/GoekeLab/xpore)

bioinformatics↗

bioRxiv2020

METTL4 catalyzes m6Am methylation in U2 snRNA to regulate pre-mRNA splicing

N6-methylation of 2-O-methyladenosine (Am) in RNA occurs in eukaryotic cells to generate N6,2-O-dimethyladenosine (m6Am). Identification of the methyltransferase responsible for m6Am catalysis has accelerated studies on the function of m6Am in RNA processing. While m6Am is generally found in the first transcribed nucleotide of mRNAs, the modification is also found internally within U2 snRNA. However, the writer required for catalyzing internal m6Am formation had remained elusive. By sequencing transcriptome-wide RNA methylation at single-base-resolution, we identified human METTL4 as the writer that directly methylates Am at U2 snRNA position 30 into m6Am. We found that METTL4 localizes to the nucleus and its conserved methyltransferase catalytic site is required for U2 snRNA methylation. By sequencing human cells with overexpressed Mettl4, we determined METTL4s in vivo target RNA motif specificity. In the absence of Mettl4 in human cells, U2 snRNA lacks m6Am thereby affecting a subset of splicing events that exhibit specific features such as overall 3 splice-site weakness with certain motif positions more affected than others. This study establishes that METTL4 methylation of U2 snRNA regulates splicing of specific pre-mRNA transcripts.

molecular biology↗