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Downregulation of the long noncoding RNA DSCR9 (Down syndrome critical region 9) delays breast cancer progression by modulating microRNA-504-5p-dependent G protein-coupled receptor 65

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Abstract

Possible roles of long noncoding RNAs (lncRNAs) in cancer stem cells (CSCs) have often been reported. Here, we focused on the regulatory function of the lncRNA Down syndrome critical region 9 (DSCR9) in breast cancer stem cells (BCSCs). Through bioinformatics analysis, DSCR9, microRNA-504-5p (miR-504-5p), and G protein-coupled receptor 65 (GPR65) were identified as targets implicated in breast cancer development. Then, clinical tissue samples, breast cancer cells, and isolated BCSCs were used to determine the expression of DSCR9, miR-504-5p, and GPR65. The results confirmed the overexpression of DSCR9 and GPR65 but low expression of miR-504-5p in breast cancer tissues and cells as well as in BCSCs. Following mechanistic investigation, it was found that DSCR9 targeted miR-504-5p, and that silencing DSCR9 inhibited the proliferation of BCSCs by elevating the expression of miR-504-5p. Additionally, miR-504-5p targeted GPR65 and inhibited its expression. Moreover, GPR65 activated the MEK/ERK signaling pathway to regulate BCSC proliferation. Finally, animal study verified that depletion of DSCR9 inhibited the proliferation of BCSCs in vivo and that BCSC proliferation was restored by overexpression of GPR65. Altogether, our findings revealed that DSCR9 elevated GPR65 expression by targeting miR-504-5p to exacerbate breast cancer, highlighting a new treatment modality for breast cancer.

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Data availability

The data that supports the findings of this study are available on request from the corresponding author.

Abbreviations

BCSCs:

Breast cancer stem cells

cDNA:

Complementary DNA

DSCR9:

Down syndrome critical region gene 9

GAPDH:

Glyceraldehyde-3-phosphate dehydrogenase

GEO:

Gene expression omnibus

GPR65:

G protein-coupled receptor 65

IHC:

Immunohistochemistry

lncRNAs:

Long non-coding RNAs

MEK/ERK:

Mitogen-activated protein kinase kinase/extracellular regulated protein kinase

miR:

MicroRNA

mRNAs:

Messenger RNAs

shRNA:

Short hairpin RNA

MEK/ERK signaling pathway:

Mitogen-activated protein kinase kinase/extracellular regulated protein kinase signaling pathway

RPMI:

Roswell park memorial institute

ALDH-1:

Aldehyde dehydrogenase-1

NC:

Negative control

RT-qPCR:

Reverse transcription quantitative polymerase chain reaction

FISH:

Fluorescence in situ hybridization

HRP:

Horseradish peroxidase

IgG:

Immunoglobulin G

DMSO:

Dimethylsulfoxide

CCK-8:

Cell counting kit-8

WT:

Wild type

MUT:

Mutant type

RIP:

RNA binding protein immunoprecipitation

AGO2:

Argonaute2

ANOVA:

Analysis of variance

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Funding

This study was supported by Quanzhou Science and Technology Bureau of Fujian Province (No. 2019N038S).

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Authors and Affiliations

  1. Area N4 of Surgical Oncology, Quanzhou First Hospital Affiliated Fujian Medical University, No. 1028, Anji South Road, Fengze District, Quanzhou, 362000, Fujian Province, China

    Mingzhu Li, Conglin Lin & Zhibing Cai

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Contributions

ML: wrote the paper and conceived and designed the experiments; analyzed the data; collected and provided the sample for this study. CL: wrote the paper and conceived and designed the experiments; analyzed the data. ZC: wrote the paper and conceived and designed the experiments; collected and provided the sample for this study.

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Correspondence to Mingzhu Li.

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Conflict of interest

The authors declare that they have no competing interests.

Ethics approval

All research protocols were approved by the Clinical Research Ethics Committee of Quanzhou First Hospital Affiliated Fujian Medical University (FJYKDXFSQZDYYY-CE-2016-0127) and in line with the Declaration of Helsinki. All patients signed informed consent forms prior to the experiments. Additionally, The animal experiments were approved by Quanzhou First Hospital Affiliated Fujian Medical University (FJYKDXFSQZDYYY-AE-2018-0506) and strictly designed and performed conforming to the Guide for the Care and Use of Laboratory Animal by the US National Institutes of Health. All efforts were made for minimizing the number and suffering of the included animals.

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13577_2023_916_MOESM1_ESM.eps

Supplementary file1 Fig. S1 Relationship between DSCR9 and clinicopathologic features of breast cancer patients and overall survival analysis in breast cancer subgroup. A, The relationship between DSCR9 and clinicopathological features of breast cancer patients was analyzed based on the BC-GenExMiner database. B, Overall survival of DSCR9 in breast cancer subgroup was analyzed based on the BC-GenExMiner database. (EPS 2237 KB)

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Li, M., Lin, C. & Cai, Z. Downregulation of the long noncoding RNA DSCR9 (Down syndrome critical region 9) delays breast cancer progression by modulating microRNA-504-5p-dependent G protein-coupled receptor 65. Human Cell 36, 1516–1534 (2023). https://doi.org/10.1007/s13577-023-00916-4

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