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METTL14 upregulates TCF1 through m6A mRNA methylation to stimulate osteogenic activity in osteoporosis

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Abstract

Alteration of N6-methyladenosine (m6A) is closely linked to spanning biological processes including osteoporosis (OP) development. This research focuses on the function of methyltransferase like 14 (METTL14) in bone turnover and its interaction with T cell factor 1 (TCF1). A mouse model of OP was established by ovariectomy (OVX). The bone mass parameters were evaluated by micro-CT analysis. Mouse MC3T3-E1 cells and mouse bone marrow macrophages (BMMs) were induced for osteogenic or osteoclastic differentiation, respectively, for in vitro experiments. The osteogenesis or osteoclasis activity was analyzed by measuring the biomarkers such as OPG, ALP, NFATC1, CTSK, RANKL, and TRAP. RT-qPCR and IHC assays identified reduced METTL14 expression in bone tissues of osteoporotic patients and ovariectomized mice. Artificial METTL14 overexpression increased bone mass of mice and promoted osteogenesis whereas suppressed osteoclasis both in vivo and in vitro. METTL14 promoted TCF1 expression through m6A mRNA methylation, and TCF1 increased the osteogenic activity by elevating the protein level of RUNX2, a key molecule linked to bone formation. In rescue experiments, TCF1 restored the RUNX2 level and osteogenic activity of cells suppressed by METTL14 silencing. In summary, this research demonstrates that METTL14 plays a protective role against OP by promoting the TCF1/RUNX2 axis.

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The analyzed data sets generated during the study are available from the corresponding author on reasonable request.

Abbreviations

ANOVA:

Analysis of variance

BMMs:

Bone marrow macrophages

CTSK:

Cathepsin K

CTX-1:

Type I collagen cross-linked C-telopeptide

FBS:

Fetal bovine serum

GAPDH:

Glyceraldehyde-3-phosphate dehydrogenase

HE:

Hematoxylin and eosin

IgG:

Immunoglobulin G

IHC:

Immunohistochemistry

KEGG:

Kyoto Encyclopedia of Genes and Genomes

METTL14:

Methyltransferase like 14

mRNA:

Messenger RNA

M-CSF:

Macrophage colony stimulating factor

m6A:

N6-methyladenosine

NC:

Negative control

NFATC1:

Nuclear factor of activated T cells

LEF:

Lymphoid enhancer factor

Oe:

Overexpression

OP:

Osteoporosis

OPG:

Osteoprotegerin

OVX:

Ovariectomy

PFA:

Paraformaldehyde

RIP:

RNA immunoprecipitation

RT-qPCR:

Reverse transcription-quantitative polymerase chain reaction

RUNX2:

Runt-related transcriptional factor 2

shRNA:

Short hairpin RNA

TCF1:

T cell factor 1

TRAP:

Triiodothyronine receptor auxiliary protein

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Funding

This work was supported by Zhejiang Provincial Public Welfare Research Project (NO. LGF21H060004 and NO. LGF22H060025) and Zhejiang Provincial Medical and Health Technology Project (Grant. 2023KY175).

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Author notes

  1. Xuepeng Wang and Chunchun Zou have contributed equally to this work.

Authors and Affiliations

  1. Department of Orthopedics Surgery, Affiliated Hangzhou First People’s Hospital, Zhejiang University School of Medicine, 261 Huansha Road, Hangzhou, 310006, Zhejiang, People’s Republic of China

    Xuepeng Wang, Maoqiang Li, Changju Hou, Wu Jiang, Zhenyu Bian & Liulong Zhu

  2. Department of Obstetrics and Gynecology, Hangzhou Third People’s Hospital, Hangzhou, 310009, Zhejiang, People’s Republic of China

    Chunchun Zou

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  1. Xuepeng Wang
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Contributions

XPW and CCZ: conception and design of the research, drafting the manuscript and obtaining funding; MQL: acquisition of data and drafting the manuscript; CJH: acquisition of data; WJ: analysis and interpretation of data; ZYB: statistical analysis; LLZ: obtaining funding, revision of manuscript for important intellectual content, and statistical analysis. All authors wrote and reviewed the manuscript.

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Correspondence to Liulong Zhu.

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The sample collection was approved by the Ethics Committee of Hangzhou First People's Hospital (Approval No: 2021170) and adhered to the Declaration of Helsinki. Signed informed consents were obtained from the patients. The usage of animals was ratified by the Animal Ethics Committee of Hangzhou First People’s Hospital (Approval No: 2021175) and the procedures were performed in strict accordance with the Guide for the Care and Use of Laboratory Animals.

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Wang, X., Zou, C., Li, M. et al. METTL14 upregulates TCF1 through m6A mRNA methylation to stimulate osteogenic activity in osteoporosis. Human Cell 36, 178–194 (2023). https://doi.org/10.1007/s13577-022-00825-y

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