The m6A methyltransferase Ime4 epitranscriptionally regulates triacylglycerol metabolism and vacuolar morphology in haploid yeast cells

Abstract

-Methyladenosine (mA) is among the most common modifications in eukaryotic mRNA. The role of yeast mA methyltransferase, Ime4, in meiosis and sporulation in diploid strains is very well studied, but its role in haploid strains has remained unknown. Here, with the help of an immunoblotting strategy and Ime4-GFP protein localization studies, we establish the physiological role of Ime4 in haploid cells. Our data showed that Ime4 epitranscriptionally regulates triacylglycerol metabolism and vacuolar morphology through the long-chain fatty acyl-CoA synthetase Faa1, independently of the RNA methylation complex (MIS complex). The MIS complex consists of the Ime4, Mum2, and Slz1 proteins. Our affinity enrichment strategy (methylated RNA immunoprecipitation assays) using mA polyclonal antibodies coupled with mRNA isolation, quantitative real-time PCR, and standard PCR analyses confirmed the presence of mA-modified transcripts in haploid yeast cells. The term "epitranscriptional regulation" encompasses the RNA modification-mediated regulation of genes. Moreover, we demonstrate that the Aft2 transcription factor up-regulates expression. Because the mA methylation machinery is fundamentally conserved throughout eukaryotes, our findings will help advance the rapidly emerging field of RNA epitranscriptomics. The metabolic link identified here between mA methylation and triacylglycerol metabolism via the Ime4 protein provides new insights into lipid metabolism and the pathophysiology of lipid-related metabolic disorders, such as obesity. Because the yeast vacuole is an analogue of the mammalian lysosome, our findings pave the way to better understand the role of mA methylation in lysosome-related functions and diseases.

The interplay between m6A RNA methylation and noncoding RNA in cancer

“…The genome-wide distribution of n6-methyladenosine (m6A) was unclear until 2012, but m6A is the most common RNA modification of mRNAs. It is enriched in many eukaryotic species of mammals, plants, and yeast [1][2][3][4][5][6][7][8] and is found in prokaryotes, including bacteria and mycoplasma [9,10]. The mRNAs of 7676 mammalian genes were found to be modified by m6A [11].…”

Structural Insights into N 6-Methyladenosine (m6A) Modification in the Transcriptome

“…ALKBH10B, an m 6 A demethylase in Arabidopsis , plays a crucial role in development by delaying flowering and repressing vegetative growth [24] . Using genetic screening, the “MIS” complex have been identified as a core RNA methyltransferase in yeast, which comprises Mum2 (orthologous to mammalian WTAP), Ime4 (orthologous to mammalian METTL3), and sporulation-specific with a leucine zipper motif protein 1 (Slz1), a third ancillary factor [20] , [22] . The MIS complex delays meiosis by abrogating mRNA methylation activity in yeast [20] , [21] , [22] .…”

“…Using genetic screening, the “MIS” complex have been identified as a core RNA methyltransferase in yeast, which comprises Mum2 (orthologous to mammalian WTAP), Ime4 (orthologous to mammalian METTL3), and sporulation-specific with a leucine zipper motif protein 1 (Slz1), a third ancillary factor [20] , [22] . The MIS complex delays meiosis by abrogating mRNA methylation activity in yeast [20] , [21] , [22] . Further studies are warranted to address the discrepancies on the ways these components get involved in m 6 A modification among various species.…”

“…The m 6 A modification, a rising star in the epitranscriptomics field, is the most pervasive modification found in transcripts [13] , [15] . First discovered in 1974 in mRNA from cancer cells [16] , m 6 A was subsequently detected in large quantities in various viruses [17] , [18] , [19] , yeast [20] , [21] , [22] , Arabidopsis [18] , [23] , [24] , Drosophila [25] , [26] , [27] , mice [28] , [29] , [30] , and humans [3] , [4] , [12] , [31] , [32] , [33] . The m 6 A modification is involved in almost all aspects of mRNA metabolism, including splicing [26] , [34] , [35] , stability [35] , [36] , [37] , [38] , and translation efficiency [36] , [39] , [40] , [41] , as well as long ncRNA (lncRNA)-mediated transcriptional repression [42] and miRNA maturation [43] .…”

Chemical RNA Modifications: The Plant Epitranscriptome

“…The MT-A subcomplex carries on a 70 kDa component, the methyltransferase player that was identified and named MT-A70 (Bokar et al 1997). MT-A70 is conserved across eukaryotes and is known as METTL3 in mammals (Liu et al 2014), IME-4 (Inducer of Meiosis-4) in S. cerevisiae (Yadav and Rajasekharan 2017) and Drosophila melanogaster (Lence et al 2016), and MTA70 in A. thaliana (Zhong et al 2008).…”