Characteristics of N6-methyladenosine modification patterns in t(8;21) acute myeloid leukemia.
10.12122/j.issn.1673-4254.2022.05.09
- Author:
Ya Nan WEN
1
;
Shu FANG
2
;
Jing Jing YANG
1
;
Hao WANG
1
;
Yi Fan JIAO
1
;
Nan WANG
1
;
Yan WEI
1
;
Li Li WANG
2
;
Li Ping DOU
2
Author Information
1. Medical School of Chinese PLA, Beijing 100853, China.
2. Department of Hematology, Fifth Medical Center of Chinese PLA General Hospital, Beijing 100853, China.
- Publication Type:Journal Article
- Keywords:
AML1-ETO;
N6-methyladenosine;
acute myeloid leukemia
- MeSH:
Adenosine/analogs & derivatives*;
Humans;
Leukemia, Myeloid, Acute/genetics*;
RNA, Messenger/metabolism*;
Transcriptome
- From:
Journal of Southern Medical University
2022;42(5):690-697
- CountryChina
- Language:Chinese
-
Abstract:
OBJECTIVE:To investigate the relationship between AML1-ETO (AE) fusion gene and intracellular N6-methyladenosine (m6A) modification pattern in t(8;21) acute myeloid leukemia (AML).
METHODS:RNA m6A sequencing was performed in SKNO-1 and AE knockdown SKNO-1 (SKNO-1 siAE) cells using RNA-protein co-immunoprecipitation and high-throughput sequencing (methylated RNA immunoprecipitation sequencing, MeRIP-Seq) to analyze the changes in m6A modification of the entire transcriptome. Transcriptome sequencing (RNA-seq) was performed using high-throughput sequencing. The differentially modified mRNAs were further functionally annotated by Gene Ontology (GO) analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis. The changes in m6A-related enzyme expressions were detected using real-time PCR.
RESULTS:A total of 26 441 genes were identified in AE knockdown AML cells and AE-expressing cells, containing 72 036 m6A peaks. AE knockdown caused a reduction of the number of intracellular m6A peaks from 37 042 to 34 994, among which 1278 m6A peaks were significantly elevated and 1225 were significantly decreased; 1316 genes with newly emerged m6A modification were detected and 1830 genes lost m6A modification after AE knockdown. The differential peaks were mainly enriched in pathways involving cancer and human T-lymphocytic leukemia virus I. RNA-seq results showed that 2483 genes were up-regulated and 3913 genes were down-regulated after AE knockdown. The combined analysis of MeRIP-Seq and RNA-Seq results revealed relatively high expression levels of m6A-modified genes as compared with the genes without m6A modification (SKNO-1: 0.6116±1.263 vs 2.010±1.655, P < 0.0001; SKNO-1 siAE: 0.5528±1.257 vs 2.067±1.686, P < 0.0001). The m6A modified genes located in the 3'UTR or 5 'UTR had significantly higher expression levels than those located in exonic regions (SKNO-1: 2.177± 1.633 vs 1.333 ± 1.470 vs 2.449 ± 1.651, P < 0.0001; SKNO-1 siAE: 2.304 ± 1.671 vs 1.336 ± 1.522 vs 2.394 ± 1.649, P < 0.05). Analysis of RNA-seq data identified 3 m6A-related enzymes that showed significantly elevated mRNA expression after AE knockdown, namely WTAP, METTL14, and ALKBH5 (P < 0.05), but the results of real-time PCR showed that the expressions of WTAP and ALKBH5 were significantly increased while the expression of METTL14 was lowered after AE knockdown (P < 0.05).
CONCLUSION:AE knockdown results in differential expressions of m6A-associated enzymes, suggesting that the AE fusion gene regulates the expression of one or more m6A-associated enzymes to control cellular methylation levels.
