XIST loss impairs mammary stem cell differentiation and increases tumorigenicity through Mediator hyperactivation

Richart, Laia; Picod-Chedotel, Mary-Loup; Wassef, Michel; Macario, Manon; Aflaki, Setareh; Salvador, Marion A.; Héry, Tiphaine; Dauphin, Aurélien; Wicinski, Julien; Chevrier, Véronique; Pastor, Sonia; Guittard, Geoffrey; Cam, Samuel Le; Kamhawi, Hanya; Castellano, Rémy; Guasch, Géraldine; Charafe-Jauffret, Emmanuelle; Heard, Edith; Margueron, Raphaël; Ginestier, Christophe

doi:10.1016/j.cell.2022.04.034

Cited by 36 publications

(34 citation statements)

References 134 publications

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“…Importantly, we showed in this study that all these factors are either not subject to XCI or are early-reactivated in female PGC. Med14 codes for a co-unit of Mediator complex, involved in transcription regulation, and is prone to early reactivation in both mouse iPS 31 and human breast cells depleted for XIST 55 . Smc1a gene has recently been shown to be involved in the remodelling and reactivation of Xi in mouse iPSC 56 .…”

Section: Discussionmentioning

confidence: 99%

Spatio-temporal X-linked gene reactivation and site-specific retention of epigenetic silencing in the mouse germline

Roidor

Syx

Beyne

et al. 2023

Preprint

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Random X-chromosome inactivation (XCI) is a hallmark of female mammalian somatic cells. This epigenetic mechanism, mediated by the long non-coding RNA Xist, occurs in the epiblast and is stably maintained to ensure proper dosage compensation of X-linked genes during life. However, this silencing is lost during primordial germ cell (PGC) development. Using a combination of single-cell allele-specific RNA sequencing and low-input chromatin profiling in developing in vivo PGC, we provide unprecedented detailed maps of gene reactivation. We demonstrated that PGC still carry a fully silent X chromosome on embryonic day (E) 9.5, despite the loss of Xist expression. X-linked genes are then gradually reactivated outside the Xist first-bound regions. At E12.5, a significant part of the inactive X chromosome (Xi) still resists reactivation, carrying an epigenetic memory of its silencing. Late-reactivated genes are enriched in repressive chromatin marks, including DNA methylation and H3K27me3 marks. Our results define the timing of reactivation of the silent X chromosome a key event in female PGC reprogramming, with direct implications for reproduction.

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Section: Discussionmentioning

confidence: 99%

Spatio-temporal X-linked gene reactivation and site-specific retention of epigenetic silencing in the mouse germline

Roidor

Syx

Beyne

et al. 2023

Preprint

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“…To calculate MOI use the formula below:

Note: A MOI of 2 was used in our previous study. 1 However other MOIs may be tested to increase transduction efficiency while carefully checking the cell. Centrifuge 24-well plates (1,000 g , 3 h, 37°C, no braking phase).…”

Section: Step-by-step Methods Detailsmentioning

confidence: 99%

“…Note: A MOI of 2 was used in our previous study. 1 However other MOIs may be tested to increase transduction efficiency while carefully checking the cell.…”

Section: Step-by-step Methods Detailsmentioning

confidence: 99%

“…We were able to silence 90% of XIST expression in the CRISPRi-engineered pHMECs with sgXIST compared to pHMECs engineered to express a scrambled sgRNA. 1 Of note, the use of vector expressing two sgRNAs targeting two different genes can be considered but a decrease in the efficiency of gene expression silencing is expected. 2 This CRISPRi-engineered pHMECs were used in different functional assays in vitro (generation of 3D organoids) and in vivo (xenotransplantation in immunocompromised mice).…”

Section: Expected Outcomesmentioning

confidence: 99%

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Production of CRISPRi-engineered primary human mammary epithelial cells with baboon envelope pseudotyped lentiviral vectors

et al. 2023

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“…For example, genetic deletion of XIST in mouse hematopoietic and human mammary epithelial cells promotes the formation of highly aggressive myeloproliferative neoplasm and HRAS G12V -driven mammary carcinoma respectively [22,23], suggesting a role of XIST expression from the inactive X chromosome (Xi) in protecting somatic cells from oncogenesis. However, in late stage breast tumors or established BC cell lines, the Xi, or Barr body, is commonly absent, presumably due to the loss of Xi and replication of the active X chromosome (Xa) [24] and/or epigenetic erosion of the Xi [25], leading to the formation of XIST clouds in the nucleus de cient in XCI [26].…”

Section: Introductionmentioning

confidence: 99%

XIST Regulates Breast Cancer Stem Cells by Activating Proinflammatory IL-6 Signaling

Zhu

et al. 2022

Preprint

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Background Aberrant expression of XIST, a long noncoding RNA initiating X chromosome inactivation (XCI) during early embryogenesis, is a common feature of breast cancer (BC). However, the roles of post-XCI XIST in breast carcinogenesis remain elusive. Methods In this study, we examined the expression of XIST in human BC cell (BCC) lines across the spectrum of BC subtypes. We then investigated the effect of knockdown (KD) of aberrantly expressed XIST in luminal and triple-negative (TN) BCCs on tumor growth, cancer stem cell (CSC) activities, and global gene expression. We identified the most significantly altered genes and pathways in ALDH− bulk tumor cells and ALDH+ CSCs upon XIST KD and validated the roles of these genes in regulating ALDH+ epithelial (E) versus CD24−/loCD44+/hi mesenchymal (M) CSCs. Lastly, we conducted miRNA array and luciferase reporter assays to define the molecular mechanisms of XIST in CSC regulation. Results Doxycycline (DOX) induced XIST KD markedly inhibits spheroid/colony forming capacity, tumor growth and tumor-initiating potential. This phenotype is attributed to impaired E-CSC in luminal and E- and M-CSC activities in TN BCCs. Gene expression profiling demonstrates that XIST KD most significantly affects cytokine-cytokine receptor interactions, resulting in markedly suppressed expression of proinflammatory cytokines IL-6 and IL-8 in the bulk of tumor cells. Exogenous IL-6, but not IL-8, rescues the reduced sphere-forming capacity and proportion of ALDH+ CSCs in luminal and TN BCCs following XIST KD. This suggests a mechanism whereby XIST regulates IL-6 production by bulk tumor cells, which then acts in a paracrine manner on ALDH+ CSCs that display elevated IL-6 receptor (IL6R) expression. XIST functions as a molecular sponge for MicroRNA let-7a-2-3p to derepress IL-6 expression, which in turn promotes self-renewal of ALDH+ CSCs by inducing STAT3 activation and expression of key CSC factors including c-MYC, KLF4 and SOX9. Conclusions This study supports a novel role of XIST by derepressing let-7 controlled paracrine IL-6 proinflammatory signaling to promote CSC self-renewal.

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XIST loss impairs mammary stem cell differentiation and increases tumorigenicity through Mediator hyperactivation

Cited by 36 publications

References 134 publications

Spatio-temporal X-linked gene reactivation and site-specific retention of epigenetic silencing in the mouse germline

Spatio-temporal X-linked gene reactivation and site-specific retention of epigenetic silencing in the mouse germline

Production of CRISPRi-engineered primary human mammary epithelial cells with baboon envelope pseudotyped lentiviral vectors

XIST Regulates Breast Cancer Stem Cells by Activating Proinflammatory IL-6 Signaling

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