Biology subjects

Liu, J. F.

Publications and source records attributed to Liu, J. F..

3 recordsLinked to original sources

bioRxiv2025

FTO depletion does not alter m6A stoichiometry in AML mRNA: a reassessment using direct RNA nanopore sequencing

The RNA demethylase FTO has been proposed to promote acute myeloid leukemia (AML) by demethylating N6-methyladenosine (m6A) from oncogenic transcripts, especially MYC. However, the evidence that supports the idea that FTO demethylates m6A in AML relies on methods that are non-quantitative and unable to reveal m6A stoichiometry changes before or after FTO depletion. To directly test whether FTO regulates m6A in mRNA, we employed Oxford Nanopore direct RNA sequencing to map and quantify m6A at single-nucleotide resolution. We find that the stoichiometry of m6A sites throughout the transcriptome and especially at MYC-specific sites are unaffected despite depletion of FTO activity by knockout, knockdown, or pharmacologic inhibition. This pattern was seen in AML cell lines MONOMAC-6 and MOLM-13, as well as in the non-AML cell line HEK293T. We also find that the anti-leukemia effect of the small-molecule FTO inhibitor FB23-2 is not due to FTO inhibition since it remains cytotoxic to FTO-deficient cells. Instead of regulating m6A, we find that FTO depletion markedly increases N6,2-O-dimethyladenosine (m6Am) in snRNAs, consistent with m6Am in snRNA being a target of FTO. Overall, our findings do not support an m6A eraser role for FTO in AML cell lines under the conditions tested, and they suggest that the reported demethylation functions of FTO on m6A should be reinvestigated using quantitative m6A mapping methods.

molecular biology↗

bioRxiv2024

Decoding m6Am by simultaneous transcription-start mapping and methylation quantification

N6,2-O-dimethyladenosine (m6Am) is a modified nucleotide located at the first transcribed position in mRNA and snRNA that is essential for diverse physiological processes. m6Am mapping methods assume each gene uses a single start nucleotide. However, gene transcription usually involves multiple start sites, generating numerous 5 isoforms. Thus, gene levels annotations cannot capture the diversity of m6Am modification in the transcriptome. Here we describe CROWN-seq, which simultaneously identifies transcription-start nucleotides and quantifies m6Am stoichiometry for each 5 isoform that initiates with adenosine. Using CROWN-seq, we map the m6Am landscape in nine human cell lines. Our findings reveal that m6Am is nearly always a high stoichiometry modification, with only a small subset of cellular mRNAs showing lower m6Am stoichiometry. We find that m6Am is associated with increased transcript expression and provide evidence that m6Am may be linked to transcription initiation associated with specific promoter sequences and initiation mechanisms. These data suggest a potential new function for m6Am in influencing transcription.

molecular biology↗

bioRxiv2024

DirectHRD enables sensitive scar-based classification of homologous recombination deficiency (HRD)

Homologous recombination deficiency (HRD) is a predictive biomarker for efficacy of PARP inhibition and platinum chemotherapy but remains challenging to detect from low tumor fraction samples such as liquid biopsies. Here, we describe DirectHRD, a whole-genome sequencing (WGS) scar-based classifier that is 10x more sensitive than state-of-the-art methods. DirectHRD can detect HRD at >=1% tumor fraction using 50x WGS of cell-free DNA.

bioinformatics↗