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Updated: May 7, 2026
A Method for Measuring RNA N6-methyladenosine Modifications in Cells and Tissues
Published on: December 5, 2016
Deciphering the "m6A Code" via Antibody-Independent Quantitative Profiling
Miguel Angel Garcia-Campos1, Sarit Edelheit1, Ursula Toth2
1Department of Molecular Genetics, Weizmann Institute of Science, 7610001 Rehovot, Israel.
Abstract:
N6-methyladenosine (m6A) is the most abundant modification on mRNA and is implicated in critical roles in development, physiology, and disease. A major limitation has been the inability to quantify m6A stoichiometry and the lack of antibody-independent methodologies for interrogating m6A. Here, we develop MAZTER-seq for systematic quantitative profiling of m6A at single-nucleotide resolution at 16%-25% of expressed sites, building on differential cleavage by an RNase. MAZTER-seq permits validation and de novo discovery of m6A sites, calibration of the performance of antibody-based approaches, and quantitative tracking of m6A dynamics in yeast gametogenesis and mammalian differentiation. We discover that m6A stoichiometry is "hard coded" in cis via a simple and predictable code, accounting for 33%-46% of the variability in methylation levels and allowing accurate prediction of m6A loss and acquisition events across evolution. MAZTER-seq allows quantitative investigation of m6A regulation in subcellular fractions, diverse cell types, and disease states.
Insights
N6-methyladenosine (m6A) is a key mRNA modification. A new method, MAZTER-seq, quantifies m6A sites, revealing a predictable code that controls methylation levels and dynamics.
Area of Science:
- Molecular Biology
- Epigenetics
- RNA Biology
Background:
- N6-methyladenosine (m6A) is the most prevalent mRNA modification.
- m6A plays crucial roles in biological processes and diseases.
- Quantifying m6A stoichiometry and developing antibody-independent methods are significant challenges.
Purpose of the Study:
- To develop a novel, antibody-independent method for quantitative profiling of m6A.
- To enable single-nucleotide resolution mapping of m6A stoichiometry.
- To investigate the regulatory mechanisms and dynamics of m6A modification.
Main Methods:
- Development of MAZTER-seq, a method utilizing differential RNase cleavage for m6A profiling.
- Systematic quantitative profiling of m6A at single-nucleotide resolution.
- Application of MAZTER-seq in yeast gametogenesis and mammalian differentiation models.
Main Results:
- MAZTER-seq enables quantitative profiling of m6A stoichiometry at 16%-25% of expressed sites.
- The study identified a cis-acting, predictable code governing m6A stoichiometry, explaining 33%-46% of methylation variability.
- MAZTER-seq facilitates validation of existing methods and discovery of novel m6A sites.
Conclusions:
- MAZTER-seq provides a powerful tool for quantitative m6A analysis, overcoming limitations of previous methods.
- m6A stoichiometry is precisely regulated by an intrinsic sequence-based code.
- The findings open new avenues for studying m6A regulation in various biological contexts and diseases.
