Cyclin D1 overexpression induces global transcriptional downregulation in lymphoid neoplasms
Robert Albero, … , Elías Campo, Pedro Jares
Robert Albero, … , Elías Campo, Pedro Jares
Published July 10, 2018
Citation Information: J Clin Invest. 2018;128(9):4132-4147. https://doi.org/10.1172/JCI96520.
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Research Article Cell biology

Cyclin D1 overexpression induces global transcriptional downregulation in lymphoid neoplasms

Abstract

Cyclin D1 is an oncogene frequently overexpressed in human cancers that has a dual function as cell cycle and transcriptional regulator, although the latter is widely unexplored. Here, we investigated the transcriptional role of cyclin D1 in lymphoid tumor cells with cyclin D1 oncogenic overexpression. Cyclin D1 showed widespread binding to the promoters of most actively transcribed genes, and the promoter occupancy positively correlated with the transcriptional output of targeted genes. Despite this association, the overexpression of cyclin D1 in lymphoid cells led to a global transcriptional downmodulation that was proportional to cyclin D1 levels. This cyclin D1–dependent global transcriptional downregulation was associated with a reduced nascent transcription and an accumulation of promoter-proximal paused RNA polymerase II (Pol II) that colocalized with cyclin D1. Concordantly, cyclin D1 overexpression promoted an increase in the Poll II pausing index. This transcriptional impairment seems to be mediated by the interaction of cyclin D1 with the transcription machinery. In addition, cyclin D1 overexpression sensitized cells to transcription inhibitors, revealing a synthetic lethality interaction that was also observed in primary mantle cell lymphoma cases. This finding of global transcriptional dysregulation expands the known functions of oncogenic cyclin D1 and suggests the therapeutic potential of targeting the transcriptional machinery in cyclin D1–overexpressing tumors.

Authors

Robert Albero, Anna Enjuanes, Santiago Demajo, Giancarlo Castellano, Magda Pinyol, Noelia García, Cristina Capdevila, Guillem Clot, Helena Suárez-Cisneros, Mariko Shimada, Kennosuke Karube, Mónica López-Guerra, Dolors Colomer, Sílvia Beà, José Ignacio Martin-Subero, Elías Campo, Pedro Jares

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Figure 6

Cyclin D1 colocalizes with RNA Pol II and promotes an increase in the Pol II pausing index.

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Cyclin D1 colocalizes with RNA Pol II and promotes an increase in the Po...
(A) Correlation between normalized cyclin D1 ChIP-Seq tag density in JVM13-cD1T286A and Pol II ChIP-Seq tag density at promoters in JVM13-ctrl and JVM13-D1T286A cells. Promoters were sorted into 50 equal-sized groups based on ChIP-seq tag densities of cyclin D1. The x axis represents mean cyclin D1 normalized tags of the promoters in JVM13-cD1T286A cell lines. The y axis represents Pol II tag density in both cell lines. The linear regression line between cyclin D1 and Pol II presence in promoters is shown. (B) Average signal profiling of Pol II occupancy around the TSS (±3 kb) in JVM13-ctrl and JVM13-D1T286A inducible cell lines. The cyclin D1–binding profile in JVM13-cD1T286A cells is also shown. (C) Western blot showing different phosphorylated forms of Pol II in JVM13-ctrl, JVM13-D1T286A, and JVM13-D1 inducible cell lines. Gels were run in duplicate for the study of the phosphorylation forms. A representative Western blot (n = 3) for each antibody is presented. α-Tubulin of only one of the gels run in duplicate is shown as loading control. (D) Plot representing the pausing index. Lines illustrate rightward shift of pausing ratio at all genes with cyclin D1 in their promoter (–5 kb, TSS) after cyclin D1 induction in JVM13-ctrl and JVM13-D1T286A cells. ***P < 2 × 10–16, Kolmogorov-Smirnov test. (E) Proportion of Pol II (IIo) and Pol (IIa) forms in primary MCL cases. P = 0.01, nonparametric Mann-Whitney U test. (F) Pol II (8WG16) antibody signal in primary MCL cases. P = 0.03, nonparametric Mann-Whitney U test. (G) Coimmunoprecipitation experiment in Z-138 cells using antibodies against cyclin D1 and control IgG. Immunoprecipitated proteins were analyzed by Western blot by blotting with cyclin D1 and Pol II antibody. Input at 1% was loaded as a control. (H) Coimmunoprecipitation experiment in HEK-293T-CDK9-FLAG-D1T286A cells with anti-FLAG resins. Immunoprecipitated proteins were analyzed by Western blot by blotting with CDK9 and cyclin D1 antibodies. HEK-293T–D1T286A immunoprecipitation was used as negative control.