Development of formaldehyde dehydrogenase-coupled assay and antibody-based assays for ALKBH5 activity evaluation

Shen, Dan‐Dan; Suo, Feng-Zhi; Song, Qimeng; Chang, Jiao; Zhang, Ting; Hong, Jing-Jing; Zheng, Yi-Chao; Liu, Hong‐Min

doi:10.1016/j.jpba.2018.09.018

Cited by 12 publications

(5 citation statements)

References 29 publications

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“…The m6A 'writer' is composed of Wilms tumor 1-associated protein (WTAP) (10), methyltransferase-like 3 and 14 (METTL3 and METTL14) (11), and protein virilizer homologue (KIAA1429), while heterogeneous nuclear ribonucleoprotein and YTH domain-containing RNA binding protein function as m6A 'readers' (12). AlkB homolog 5 (ALKBH5) and fat mass and obesity-associated protein (FTO) work as m6A 'erasers' to remove it from the target region (13,14). Cross-talk and dynamic equilibrium among m6A writers, readers, and erasers play important roles in cancer development (15,16).…”

Section: Introductionmentioning

confidence: 99%

METTL14 promotes the migration and invasion of breast cancer cells by modulating N6‑methyladenosine and hsa‑miR‑146a‑5p expression

Wang

Shi

et al. 2020

Oncol Rep

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Breast cancer (BC) is the most frequently diagnosed cancer and the leading cause of cancer-related death among women worldwide. Evidence indicates that posttranscriptional N6-methyladenosine (m6A) modification modulates BC development. In the present study, we assessed BC and normal tissues to investigate this connection. RNA m6A levels were determined by methylation quantification assay. The effects of methyltransferase-like 14 (METTL14) gain-of-expression or co-transfection with an m6A inhibitor on cell migration and invasion abilities were determined by Transwell assays. The levels of differentially expressed (DE) miRNAs were verified by real-time quantitative PCR (RT-qPCR). Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes analyses (KEGG) were performed to analyze potential function of target genes of the DE miRNAs. The effects of candidate miRNAs modulated by METTL14 on cell migration and invasion abilities were confirmed by Transwell assays. We demonstrated that m6A methyltransferase METTL14 was significantly upregulated in BC tissues compared with normal tissues. METTL14 gain-and loss-of-expression regulated m6A levels in MCF-7 and MDA-MB-231 cells. The cell function assays revealed that METTL14 overexpression enhanced the migration and invasion capacities of BC cells. Moreover, treatment with the m6A inhibitor suppressed this enhanced cell migration and invasion. Additionally, aberrant expression of METTL14 reshaped the miRNA profile in BC cell lines. The remodeled DE miRNA/mRNA network was found to be most enriched in cancer pathways, and DE miRNAs were enriched in cell adhesion terms. hsa-miR-146a-5p modulated by METTL14 promoted cell migration and invasion. METTL14 modulates m6A modification and hsa-miR-146a-5p expression, thereby affecting the migration and invasion of breast cancer cells.

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Section: Introductionmentioning

confidence: 99%

METTL14 promotes the migration and invasion of breast cancer cells by modulating N6‑methyladenosine and hsa‑miR‑146a‑5p expression

Wang

Shi

et al. 2020

Oncol Rep

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“…The human ALKBH5 gene, located in the 17 p11.2 region, encodes a protein with 394 amino acid residues and a molecular weight of ~43 kD. ALKBH5, located in the nuclear speckle body, can catalyse m6A demethylation in a ketoglutaric acid‐ and F 2+ ‐dependent manner 27,28 . ALKBH5 can directly catalyse m6A methylation of adenosine to remove methyl groups without intermediate products 29 .…”

Section: Discussionmentioning

confidence: 99%

“…ALKBH5, located in the nuclear speckle body, can catalyse m6A demethylation in a ketoglutaric acid-and F 2+ -dependent manner. 27,28 ALKBH5 can directly catalyse m6A methylation of adenosine to remove methyl groups without intermediate products. 29 Currently, abnormal ALKBH5 levels have been demonstrated to be related to a variety of tumours.…”

Section: Discussionmentioning

confidence: 99%

ALKBH5 inhibits the SIRT3/ACC1 axis to regulate fatty acid metabolism via an m6A‐IGF2BP1‐dependent manner in cervical squamous cell carcinoma

Zhang

Pan

2023

Clin Exp Pharma Physio

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Cervical cancer (CC) is the most common malignancy of the female reproductive system, among which cervical squamous cell carcinoma (CESC) is the most common type. The demethylase ALKBH5 has been previously revealed to be downregulated in CC tissue. N6 methyladenine (m6A) is the most common modification in eukaryotic RNAs and is involved in modulating tumour progression. Therefore, we attempted to clarify the ALKBH5 role and mechanism underlying CESC progression.In CESC, patient tissue and control tissue m6A levels were measured. Reverse transcription quantitative real-time polymerase chain reaction, western blotting and immunochemistry were used to measure ALKBH5 levels. A correlation between CESC patient survival and ALKBH5 levels was evaluated. Wound healing, transwell and colony formation assays were used to detect CESC cellular behaviours. Corresponding kits and BODIPY staining were used to detect CESC lipid metabolism. Bioinformatics, immunoprecipitation, RNA pulldown and RNA immunoprecipitation assays as well as half-life measurements were used to assess the association and mechanism of ALKBH5 with silent mating type information regulation 2 homologue 3 (SIRT3), acetyl-CoA carboxylase 1 (ACC1) and insulin-like growth factor 2 mRNA binding protein 1 (IGF2BP1). The m6A demethylase ALKBH5 was depleted in CESC tissue and cells, and a low level of ALKBH5 predicted an unfavourable prognosis in CESC patients. ALKBH5 overexpression suppressed CESC growth and lipid metabolism in vitro and CESC tumour growth in vivo, and ACC1 overexpression rescued these changes. ALKBH5 downregulated ACC1 levels in CESC cells by facilitating SIRT3 methylation to repress ACC1 deacetylation. ALKBH5 destabilized SIRT3 to downregulate SIRT3 levels in CESCs in an m6A-IGF2BP1-dependent manner.ALKBH5 demethylates and destabilizes SIRT3 in an m6A-IGF2BP1-dependent manner, repressing CESC growth, lipid metabolism and tumorigenesis by downregulating ACC1.

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“…[43][44][45] ALKBH5 also can erase m 6 A methylation in mRNA and affect RNA metabolism. [46][47][48] Analysis revealed that ALKBH5 was similar to FTO in catalytic structure, [35,49] although ALKBH5 only recognizes and reverses methylation of single-stranded RNA/DNA. [42] 2.3.…”

Section: Writersmentioning

confidence: 99%

Dysregulation and implications of N6-methyladenosine modification in renal cell carcinoma

Wu¹,

Zhang²,

Chen³

et al. 2022

Curr Urol

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Increasing evidence indicates that N6-methyladenosine (m6A) methylation modification serves important functions in biological metabolism. Dysregulation of m6A regulators is related to the progression of different malignancies, including renal cell carcinoma (RCC). Recent studies have reported preliminary findings on the influence of m6A regulator dysregulation on RCC tumorigenesis and development. However, no comprehensive review that integrates and analyzes the roles of m6A modification in RCC has been published to date. In this review, we focus on the dysregulation of m6A regulators as it relates to RCC tumorigenesis and development, as well as possible applications of m6A modification in RCC diagnosis and therapeutics.

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Development of formaldehyde dehydrogenase-coupled assay and antibody-based assays for ALKBH5 activity evaluation

Cited by 12 publications

References 29 publications

METTL14 promotes the migration and invasion of breast cancer cells by modulating N6‑methyladenosine and hsa‑miR‑146a‑5p expression

METTL14 promotes the migration and invasion of breast cancer cells by modulating N6‑methyladenosine and hsa‑miR‑146a‑5p expression

ALKBH5 inhibits the SIRT3/ACC1 axis to regulate fatty acid metabolism via an m6A‐IGF2BP1‐dependent manner in cervical squamous cell carcinoma

Dysregulation and implications of N6-methyladenosine modification in renal cell carcinoma

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