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. 2024 Jun 6;150(6):294.
doi: 10.1007/s00432-024-05799-y.

LncRNA HOXC-AS3 accelerates malignant proliferation of cervical cancer cells via stabilizing KDM5B

Jie Li #  1 , Fang Hou #  1 , Zhenghua Teng  1 , Weiwei Xia  1 , Jie Peng  2
Affiliations

LncRNA HOXC-AS3 accelerates malignant proliferation of cervical cancer cells via stabilizing KDM5B

Jie Li et al. J Cancer Res Clin Oncol. .

Abstract

Background: Cervical cancer (CC) is a common malignancy amongst women globally. Ubiquitination plays a dual role in the occurrence and development of cancers. This study analyzed the mechanism of long noncoding RNA HOXC cluster antisense RNA 3 (lncRNA HOXC-AS3) in malignant proliferation of CC cells via mediating ubiquitination of lysine demethylase 5B (KDM5B/JARID1B).

Methods: The expression patterns of lncRNA HOXC-AS3 and KDM5B were measured by real-time quantitative polymerase chain reaction or Western blot analysis. After transfection with lncRNA HOXC-AS3 siRNA and pcDNA3.1-KDM5B, proliferation of CC cells was assessed by the cell counting kit-8, colony formation, and 5-Ethynyl-2'-deoxyuridine staining assays. The xenograft tumor model was established to confirm the impact of lncRNA HOXC-AS3 on CC cell proliferation in vivo by measuring tumor size and weight and the immunohistochemistry assay. The subcellular location of lncRNA HOXC-AS3 and the binding of lncRNA HOXC-AS3 to KDM5B were analyzed. After treatment of lncRNA HOXC-AS3 siRNA or MG132, the protein and ubiquitination levels of KDM5B were determined. Thereafter, the interaction and the subcellular co-location of tripartite motif-containing 37 (TRIM37) and KDM5B were analyzed by the co-immunoprecipitation and immunofluorescence assays.

Results: LncRNA HOXC-AS3 and KDM5B were upregulated in CC tissues and cells. Depletion of lncRNA HOXC-AS3 repressed CC cell proliferation and in vivo tumor growth. Mechanically, lncRNA HOXC-AS3 located in the nucleus directly bound to KDM5B, inhibited TRIM37-mediated ubiquitination of KDM5B, and upregulated the protein levels of KDM5B. KDM5B overexpression attenuated the inhibitory role of silencing lncRNA HOXC-AS3 in CC cell proliferation in vivo and in vitro.

Conclusion: Nucleus-located lncRNA HOXC-AS3 facilitated malignant proliferation of CC cells via stabilization of KDM5B protein levels.

Keywords: Cervical cancer; KDM5B; LncRNA HOXC-AS3; Proliferation; TRIM37.

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Conflict of interest statement

All authors have no conflicts of interest.

Figures

Fig. 1
Fig. 1
HOXC-AS3 expression levels were upregulated in CC. A HOXC-AS3 expression levels in cervical squamous cell carcinoma were analyzed via the UALCAN database. B HOXC-AS3 expression levels in 45 pairs of cancer tissues and para-cancerous tissues of CC patients were detected via RT-qPCR, N = 45; C HOXC-AS3 expression levels in normal cervical epithelial cell line (Ect1/E6E7 cells) and in CC cell lines (C33A, Hela, SiHa, and CaSki cells) were examined via RT-qPCR. Cell experiments were independently repeated 3 times. Data in panel C were shown as mean ± standard deviation; data comparisons in panel B were tested via the t test; data comparisons in panel C were tested via one-way ANOVA, followed Tukey’s multiple comparison test. *p < 0.05; **p < 0.01
Fig. 2
Fig. 2
Silencing HOXC-AS3 retards malignant proliferation of CC cells. CaSki cells were transfected with HOXC-AS3 siRNA (si-HOXC-AS3), with NC siRNA (si-NC) as the negative control; HeLa cells were transfected with pcDNA3.1-HOXC-AS3 (HOXC-AS3), with pcDNA3.1-empty vector (NC) as the negative control. A, B HOXC-AS3 expression levels in CaSki and HeLa cells were detected via RT-qPCR. C Cell proliferation in different groups was determined via CCK-8 assay. D HOXC-AS3 expression in cells at 72 h was detected via RT-qPCR. E, F Cell proliferation in different groups was determined via colony formation and EdU assays. Cell experiments were independently repeated 3 times. Data were shown as mean ± standard deviation; data comparisons in panels B, D–F were tested via the t test; data comparisons in panel A were tested via one-way ANOVA and data in panel C were tested via two-way ANOVA, followed Tukey’s multiple comparison test. *p < 0.05; **p < 0.01
Fig. 3
Fig. 3
HOXC-AS3 directly binds to KDM5B. A The RNAInter database predicted the interaction between HOXC-AS3 and KDM5B. B, C The subcellular location of HOXC-AS3 was analyzed by the RNA FISH and nuclear/cytoplasmic fractionation assays. D, E The binding of HOXC-AS3 to KDM5B was analyzed by the RNA pull-down and RIP assays. Cell experiments were independently repeated 3 times. Data in panels were shown as mean ± standard deviation; data comparisons in panel E were tested via one-way ANOVA, followed Tukey’s multiple comparison test. **p < 0.01
Fig. 4
Fig. 4
HOXC-AS3 diminishes KDM5B ubiquitination via binding to KDM5B. A, B KDM5B expression levels in CC tissues (N = 45; representative bands) and CC cells were detected via WB assay. C E3 ubiquitin ligases that can regulate KDM5B were predicted via the Ubibrowser database. CaSki cells were transfected with TRIM37 siRNA (si-TRIM37) with NC siRNA (si-NC) as the negative control. D, E TRIM37 expression levels in CaSki cells were detected via RT-qPCR and WB assays. F The ubiquitination levels of KDM5B cells in each group were detected via the ubiquitination assay. Cell experiments were independently repeated 3 times. Data were shown as mean ± standard deviation; data comparisons in panels A, B (right), and E were tested via the t-test; data comparisons in panels B, D were tested via one-way ANOVA, followed by Tukey’s multiple comparison test. *p < 0.05; **p < 0.01
Fig. 5
Fig. 5
HOXC-AS3 interferes with the interaction between TRIM37 and KDM5B. A The interaction between KDM5B and TRIM37 was analyzed via the Co-IP assay. B The cellular co-location of KDM5B and TRIM37 was determined via the immunofluorescence assay. C The interaction between KDM5B and TRIM37 was analyzed via the Co-IP assay. Cell experiments were independently repeated 3 times
Fig. 6
Fig. 6
KDM5B overexpress neutralizes the function of HOXC-AS3 knockdown in repressing proliferation of CC cells. CaSki cells were transfected with pcDNA3.1-KDM5B (KDM5B) with pcDNA3.1-empty vector (NC) as the negative control. A, B KDM5B expression levels in CaSki cells were detected via RT-qPCR and WB assays. C–E Cell proliferation in different groups was determined via the CCK-8, colony formation, and EdU assays. Cell experiments were independently repeated 3 times. Data in panels were shown as mean ± standard deviation; data comparisons in panel A were tested via the t test; data comparisons in panels B, D, E were tested via one-way ANOVA and data in panel C were tested via two-way ANOVA, followed Tukey’s multiple comparison test. **p < 0.01
Fig. 7
Fig. 7
Silencing HOXC-AS3 represses tumor growth by decreasing KDM5B expression levels. CaSki cells with stable HOXC-AS3 knockdown (sh-HOXC-AS3) were used to establish the xenograft tumor model. A The tumor volume. B The representative picture and weight data of the resected tumors at the 4th week after animal euthanasia. C The positive rate of Ki67 in tumor tissues were measured via the IHC assay. D HOXC-AS3 expression levels in tumor tissues were detected via RT-qPCR. E KDM5B protein levels in tumor tissues were examined via the WB assay. N = 6; Cell experiments were independently repeated 3 times. Data in panels A–C, E were shown as mean ± standard deviation; data comparisons in panels B–E were tested via the t test; data comparisons in panel A were tested via two-way ANOVA, followed Tukey’s multiple comparison test. **p < 0.01
Fig. 8
Fig. 8
HOXC-AS3 promoted malignant proliferation of CC cells via regulating KDM5B. HOXC-AS3 directly bound to KDM5B, which inhibited TRIM37-mediated KDM5B ubiquitination and upregulated KDM5B protein levels, promoting malignant proliferation of CC cells
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References

    1. Aalijahan H, Ghorbian S. Long non-coding RNAs and cervical cancer. Exp Mol Pathol. 2019;106:7–16. doi: 10.1016/j.yexmp.2018.11.010. - DOI - PubMed
    1. Adiga D, Eswaran S, Pandey D, Sharan K, Kabekkodu SP. Molecular landscape of recurrent cervical cancer. Crit Rev Oncol Hematol. 2021;157:103178. doi: 10.1016/j.critrevonc.2020.103178. - DOI - PubMed
    1. Bridges MC, Daulagala AC, Kourtidis A. LNCcation: lncRNA localization and function. J Cell Biol. 2021;220(2):9045. doi: 10.1083/jcb.202009045. - DOI - PMC - PubMed
    1. Bueno MT, Richard S. SUMOylation negatively modulates target gene occupancy of the KDM5B, a histone lysine demethylase. Epigenetics. 2013;8(11):1162–1175. doi: 10.4161/epi.26112. - DOI - PubMed
    1. Chandrashekar DS, Karthikeyan SK, Korla PK, Patel H, Shovon AR, Athar M, et al. UALCAN: an update to the integrated cancer data analysis platform. Neoplasia. 2022;25:18–27. doi: 10.1016/j.neo.2022.01.001. - DOI - PMC - PubMed

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