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. 2023 Oct 16;42(47):3514–3528. doi: 10.1038/s41388-023-02858-5

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© The Author(s) 2023

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 7 — A Enrichment plot of cell cycle pathways after DDX3X knockdown in MDA-MB-231 cells by GSEA analysis. B The ssGSEA scores of cell cycle pathways according to KLHL29 mRNA levels based on FUSCC-TNBC dataset. C, D Depletion of DDX3X leads to an accumulation of cell cycle at G0/G1 phase in MDA-MB-231 (C) and SUM159PT (D) cells by flow cytometry analysis. Cells were stained with propidium iodide (PI) for the cell cycle assays. The percentage of cells in the G1/S/G2M phase are shown. E, F Ectopic expression of KLHL29 leads to an accumulation of cell cycle at G0/G1 phase in BT549 (E) and CAL51 (F) cells by flow cytometry analysis. G, H Knockdown of KLHL29 leads to a reduction of cell cycle at G0/G1 phase in MDA-MB-231 (G) and SUM159PT (H) cells by flow cytometry analysis. I, J The accumulation of cell cycle at G0/G1 phase induced by ectopic KLHL29 is restored by re-expression of DDX3X in BT549 (I) and CAL51 (J) cells by flow cytometry analysis. K, L Depletion of DDX3X represses the mRNA expression of CCND1 in MDA-MB-231 (K) and SUM159PT (L) cells by RT-qPCR analysis. M, N Ectopic expression of KLHL29 represses the mRNA expression of CCND1 in BT549 (M) and CAL51 (N) cells by RT-qPCR analysis. O DDX3X binds to the transcripts of CCND1. MDA-MB-231 and HEK293T cells were transfected with plasmids encoding Myc-DDX3X, followed by the RIP assays using anti-Myc antibody. P Depletion of DDX3X promotes the destabilization of CCND1 mRNA. ActD (5 μg/ml) was used to block RNA Pol II-mediated transcription in MDA-MB-231 cells. Q Ectopic expression of KLHL29 promotes the destabilization of CCND1 mRNA. R, S RK33 treatment reduces DDX3X and CCND1 mRNA level in MDA-MB-231 (R) and SUM159PT (S) cells by RT-qPCR analysis. T, U RK33 treatment leads to an accumulation of cell cycle at G0/G1 phase in MDA-MB-231 (T) and SUM159PT (U) cells by flow cytometry analysis. *p < 0.05, **p < 0.01, ***p < 0.001, ns, no significance by two-tailed Student’s t test.