Bclaf1 biomolecular condensates protect nuclear PTK2 from ubiquitin-proteasome system promoting cardiomyocyte survival during oxidative stress

Aparecida Moretto, I.; Rodrigues, B. R. I.; Victor-Carvalho, P.; Carvalho, M. d. G. d. S.; Silva, M. C. d.; Valdivieso-Rivera, F. B.; Araujo, G. L. d.; Samogim, A. P.; Novais, L. B.; de Brito, I. R.; Reis-de-Oliveira, G.; Amaral, A. G.; Baratti, M. O.; Basei, F. L.; Geraldo, M. V.; Carvalho, P. C.; Santos, M. D. M.; Duran, R.; Sponton, C.; Kobarg, J.; Gozzo, F. C.; Carvalho, H. F.; Thomaz, A. A. d.; Santos, A. M. d.

doi:10.1101/2025.02.04.636487

Home/Archive/bioRxiv

bioRxiv · 10.1101/2025.02.04.636487

Bclaf1 biomolecular condensates protect nuclear PTK2 from ubiquitin-proteasome system promoting cardiomyocyte survival during oxidative stress

View original source ↗Export metadata ↓

Abstract

PTK2, a non-receptor tyrosine kinase, plays a critical role in regulating essential cellular functions, including cell survival, by reducing p53 levels and activating the PI3K/AKT pathway. However, the mechanism underlying PTK2 stabilization during stress remains unclear. In this study, we identified Bclaf1, a multifunctional protein known to stabilize partners, as a PTK2 interactor. Using advanced microscopy techniques we identified nuclear Bclaf1 biomolecular condensates containing PTK2 in cardiomyocytes under oxidative stress. While diffuse PTK2 in the nucleus was susceptible to ubiquitination, PTK2 sequestered in the Bclaf1 condensates was protected from the ubiquitin-proteasome system (UPS). The K926 residue was identified as a ubiquitination site on PTK2, and subsequent proteasome inhibition experiments confirmed the role of the UPS in PTK2 homeostasis. Furthermore, disrupting Bclaf1 biomolecular condensates lead to PTK2 degradation, subsequently increasing p53 levels and activating apoptosis. Our findings support the role of Bclaf1 in the formation of pro-survival nuclear condensates that sequester and stabilize PTK2, promoting cardiomyocyte survival during oxidative stress. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=151 SRC="FIGDIR/small/636487v3_ufig1.gif" ALT="Figure 1"> View larger version (33K): org.highwire.dtl.DTLVardef@15377a9org.highwire.dtl.DTLVardef@d5325forg.highwire.dtl.DTLVardef@de0ac9org.highwire.dtl.DTLVardef@196325d_HPS_FORMAT_FIGEXP M_FIG C_FIG In BriefMoretto et al. demonstrated that Bclaf1 undergo liquid-liquid phase separation (LLPS) during doxorubicin (Dox) induced oxidative stress to generate membraneless organelles where PTK2 is sequestered and protected from the ubiquitin-proteasome system (UPS). The maintenance of PTK2 stability may favor the p53 and Mdm2 interaction, which results in p53 ubiquitination and degradation, promoting cell survival. Loss of Bclaf1 disrupts PTK2 stabilization, leading to the ubiquitination and degradation of this kinase and downstream upregulation of p53, increasing cell death. HighlightsO_LIBclaf1 undergoes LLPS to stabilize protein partners during stress. C_LIO_LIPTK2 inside the Bclaf1 biomolecular condensates is protected from the UPS to promote cardiomyocyte survival during dox-induced oxidative stress. C_LIO_LIThe PTK2 ubiquitination site, lysine 926, was identified and the action of the UPS on PTK2 proteostasis was confirmed. C_LIO_LIBclaf1 knockdown results in overall protein ubiquitination and in PTK2 ubiquitination and degradation, resulting in increased levels of p53 and PUMA. C_LI

Explore related subjects

cell biology Preprint

Keep this discovery

Explore connections, maps & timelines

BibTeX RIS

Aparecida Moretto, I., Rodrigues, B. R. I., Victor-Carvalho, P., Carvalho, M. d. G. d. S., Silva, M. C. d., Valdivieso-Rivera, F. B., Araujo, G. L. d., Samogim, A. P., Novais, L. B., de Brito, I. R., Reis-de-Oliveira, G., Amaral, A. G., Baratti, M. O., Basei, F. L., Geraldo, M. V., Carvalho, P. C., Santos, M. D. M., Duran, R., Sponton, C., Kobarg, J., Gozzo, F. C., Carvalho, H. F., Thomaz, A. A. d., Santos, A. M. d.. 2025-02-07. Bclaf1 biomolecular condensates protect nuclear PTK2 from ubiquitin-proteasome system promoting cardiomyocyte survival during oxidative stress. https://doi.org/10.1101/2025.02.04.636487

Cite the original work for its findings. Save a collection to share your selection of sources.

KEEP EXPLORING

Related preprints

bioRxiv2026

Not all TOP RNAs are created equal: 3'UTR length and TSS selection predict the translational regulation of LARP1-bound mRNAs in CD4+ T cells

Naive T cells are poised for activation and contain a pool of translationally repressed ribosomal protein (RP) mRNA prepared to induce ribosome biogenesis to support protein synthesis, cell growth and proliferation. RP mRNA are the prototypical members of a class of transcripts initiating at cytosine followed by a CU rich element called terminal oligo pyrimidine (TOP) RNAs. TOP RNAs are regulated by an RNA binding protein LARP1, which promotes transcript stabilisation and translational repression. We investigated LARP1 function in T cell activation by generating cross-linking immunoprecipitation (CLIP) datasets detailing the LARP1-RNA interactions in naive and activated CD4+ T cells and identifying novel TOP RNAs. TOP RNAs identified by this analysis were functionally diverse. RP mRNAs were typified by high stability, and translational repression in naive T cells followed by MTORC1-dependent translation increases following T cell activation. However, other TOP RNAs varied in these aspects of their regulation. Notably, TOP RNAs with longer 3'UTRs had a relaxed dependency on LARP1 for stability and a reduced dependency on MTORC1 for their translation. Transcription start site heterogeneity also impacted TOP RNA regulation by generating a mixture of transcript isoforms with different TOP motif lengths. Longer terminal oligo pyrimidine stretches were associated with a greater dependency on MTORC1 for translation. Differential regulation of TOP RNAs may allow tuneable translational responses to MTORC1 and indicates potential roles for LARP1 beyond translation regulation and stability.

cell biology↗

bioRxiv2026

Sex-specific metabolic regulation by the Drosophila RNA-binding protein Nab2

Conserved RNA binding proteins (RBPs) regulate key steps of gene expression including mRNA processing, export, localization, stability and translation. Human ZC3H14 is a conserved RBP that regulates pre-mRNA processing in neurons and loss of ZC3H14 leads to neurological defects. Studies of Nab2, the Drosophila orthologue of ZC3H14, have identified potential target RNAs involved in metabolism, suggesting Nab2 may influence neurometabolic circuitry. Here, we show a female-specific increase in dilp2 and dilp5 mRNA levels. The dilps encode insulin-like peptides that signal from the brain insulin producing cells (IPCs) to peripheral tissues. Nab2null females have enlarged lipid droplets in the fat body, a tissue analogous to human adipose tissue and liver. Notably, neuronal depletion of Nab2 increases lipid droplet size while neuronal expression of Nab2 in Nab2null female rescues this phenotype supporting a role for Nab2 in a neuronal circuit that regulates dilp levels. Furthermore, depletion of dilp2 or dilp5 from IPCs rescues the enlarged lipid droplet phenotype in Nab2null females indicating that elevated dilp2/dilp5 contributes to enlarged lipid droplets. Together, these data support a female-specific role for Nab2 in brain neurons to support insulin signaling and fat storage, expanding the known functions of RBPs linking neuronal function and metabolic homeostasis.

cell biology↗

bioRxiv2026

Deep generative embeddings of gene expression and splicing reposition the interpretation of single-cell transcriptomic signatures

Single-cell transcriptomic analysis predominantly derives cell identity from gene expression analysis, while alternative splicing is processed separately despite its fundamental role for cell homeostasis. To overcome the limits of separate investigations, we developed a probabilistic deep learning framework, Crecerelle, enabling resolution of the contributions of gene expression and alternative splicing in each cell. Crecerelle learns cell embeddings from gene expressions and alternative splicing isoforms, to decipher their mutually dependent impact on the functional characterisation of cells in a data-driven manner, exemplified for the Tabula Muris dataset. This is enabled through a zero-and-N-inflated Dirichlet-Multinomial for a variational autoencoder that learns cell embeddings solely from splicing profiles, as well as a bi-modal variational autoencoder with a relevance-weighted mixture-of-experts variational posterior to consolidate the modality-specific contribution at single-cell level. Crecerelle reveals cell-type-specific isoform markers as well as subpopulations with unique isoforms and uncovers regulatory and disease-associated pathways not detected by gene expression analyses alone. This scalable and interpretable framework thus allows a more holistic study of transcriptomic regulation and will open a route to modality-relevance-weighted investigations across single-cell multiomics datasets and their influence on cellular homeostasis, tissue development and disease phenotypes.

cell biology↗