The tumor suppressor HNRNPK induces p53-dependent nucleolar stress to drive ribosomopathies
Pedro Aguilar-Garrido, … , Sean M. Post, Miguel Gallardo
Pedro Aguilar-Garrido, … , Sean M. Post, Miguel Gallardo
Published May 8, 2025
Citation Information: J Clin Invest. 2025;135(12):e183697. https://doi.org/10.1172/JCI183697.
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Research Article Aging Hematology

The tumor suppressor HNRNPK induces p53-dependent nucleolar stress to drive ribosomopathies

Abstract

The nucleolus is a membraneless organelle and an excellent stress sensor. Any changes in its architecture or composition lead to nucleolar stress, resulting in cell cycle arrest and interruption of ribosomal activity, critical factors in aging and cancer. In this study, we identified and described the pivotal role of the RNA-binding protein HNRNPK in ribosome and nucleolar dynamics. We developed an in vitro model of endogenous HNRNPK overexpression and an in vivo mouse model of ubiquitous HNRNPK overexpression. These models showed disruptions in translation as the HNRNPK overexpression caused alterations in the nucleolar structure, resulting in p53-dependent nucleolar stress, cell cycle arrest, senescence, and bone marrow failure phenotype, similar to what is observed in patients with ribosomopathies. Together, our findings identify HNRNPK as a master regulator of ribosome biogenesis and nucleolar homeostasis through p53, providing what we believe to be a new perspective on the orchestration of nucleolar integrity, ribosome function and cellular senescence.

Authors

Pedro Aguilar-Garrido, María Velasco-Estévez, Miguel Ángel Navarro-Aguadero, Álvaro Otero-Sobrino, Marta Ibáñez-Navarro, Miguel Ángel Marugal, María Hernández-Sánchez, Prerna Malaney, Ashley Rodriguez, Oscar Benitez, Xiaroui Zhang, Marisa J.L. Aitken, Alejandra Ortiz-Ruiz, Diego Megías, Manuel Pérez, Gadea Mata, Jesús Gomez, Miguel Lafarga, Orlando Domínguez, Osvaldo Graña-Castro, Eduardo Caleiras, Pilar Ximénez-Embun, Marta Isasa, Paloma Jimena de Andres, Sandra Rodríguez-Perales, Raúl Torres-Ruiz, Enrique Revilla, Rosa María García-Martín, Daniel Azorín, Josune Zubicaray, Julián Sevilla, Oleksandra Sirozh, Vanesa Lafarga, Joaquín Martínez-López, Sean M. Post, Miguel Gallardo

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Figure 4

The HNRNPK overexpression phenotype is p53 dependent.

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The HNRNPK overexpression phenotype is p53 dependent. (A) Western blot m...
(A) Western blot membrane showing HNRNPK and p53 in Tp53lox/WT, HnrnpkTg-cre, and HnrnpkTg-cre/Tp53lox/WT. (B) Representative images of HPG assay and fluorescence intensity values (dot plot, >1.000 cells of representative well; biological replicate analysis, HnrnpkTg-cre vs. HnrnpkTg-cre/Tp53lox/WT, P = 0.0006). Scale bar: 100 μm. (C) Representative confocal microscopy images of NCL (left), FBL (right) HNRNPK, and DAPI staining. For dot plot analysis of Tp53lox/WT, HnrnpkTg-cre, and HnrnpkTg-cre/Tp53lox/WT, Hnrnpk expression was measured with Alexa Fluor 488 intensity (dot plot cell replicates, >1.000; biological replicate analysis: P = 0.0044); NCL relocalization was analyzed with nucleoplasm Alexa Fluor 647 intensity (dot plot cell replicates, >1.000; biological replicate analysis: P = 0.0018); and FBL reload was assessed by Alexa Fluor 647 spot total area (dot plot cell replicates, >1.000; biological replicate analysis: P = 0.0038). Scale bar: 10 μm. (D) Representative cell cycle FCM histogram plots in HnrnpkTg-cre/Tp53lox/WT (n = 4) vs. HnrnpkTg-cre/Tp53lox/WT (n = 3) MEFs and bar graph of the percentage of G1, S and G2/M % (P = 0.02) from FCM analysis. (E) Bright-field microscope images of SA-β-galactosidase staining in HnrnpkTg-cre vs. HnrnpkTg-cre/Tp53lox/WT MEFs and dot plot of cells positive for SA-β-galactosidase staining: HnrnpkTg-cre/Tp53lox/WT (n = 8) vs. HnrnpkTg-cre (n = 6) (P = 0.0007). All graphs are shown as the median or mean. All 2-group statistical analyses were 2-sided Student’s t test; for 3-group statistical analyses, 2-way ANOVA and Dunn’s multiple comparisons test were used: *P < 0.05; **P < 0.01; ***P < 0.001. All experiments comprised at least n = 3 biological replicates and/or n = 3 technical replicates.