Ganez Zapater, A. | bioRxiv Science

bioRxiv2023

Nuclear MTHFD2 secures mitosis progression by preserving centromere integrity

Subcellular compartmentalization of metabolic enzymes may elicit specific cellular functions by establishing a unique metabolic environment. Indeed, the nuclear translocation of certain metabolic enzymes is required for epigenetic regulation and gene expression control. Here, we reveal that, in cancer cells, the mitochondrial enzyme methylenetetrahydrofolate dehydrogenase 2 (MTHFD2) localizes in the nucleus during the G2-M phase of the cell cycle to secure mitosis progression. Nuclear MTHFD2 interacts with proteins involved in mitosis regulation and centromere stability, including the methyltransferases KMT5A and DNMT3B. Loss of MTHFD2 induces centromere overexpression and severe methylation defects and impedes correct mitosis completion. As a consequence, MTHFD2 deficient cells accumulate chromosomal aberrations arising from chromosome congression and segregation defects. Blocking the catalytic nuclear function of MTHFD2 recapitulates the phenotype observed in MTHFD2 deficient cells, attributing to nuclear MTHFD2 an enzymatic active role in controlling mitosis. Our discovery uncovers a nuclear moonlighting role for the cancer target MTHFD2, and emphasizes that cancer metabolism rewiring may encompass the relocation of metabolic enzymes to alternative subcellular compartments.

cell biology↗

bioRxiv2022

The BRD4S-LOXL2-MED1 interaction at the forefront of cell cycle transcriptional control in triple-negative breast cancer

Triple-negative breast cancer often develops resistance to single-agent treatments, which can be circumvented with targeted combinatorial approaches. Here, we demonstrate that the simultaneous inhibition of LOXL2 and BRD4 cooperate to reduce triple-negative breast cancer proliferation in vitro and in vivo. Mechanistically, we reveal that LOXL2 interacts in the nucleus with the short isoform of BRD4 and MED1 to control cell cycle progression at the gene expression level via sustaining the formation of BRD4-MED1 nuclear transcriptional foci. Indeed, the pharmacological or transcriptional repression of LOXL2 provokes downregulation of cell cycle gene expression, G1-S cell cycle arrest, and loss of BRD4-MED1 foci. Our results indicate that the BRD4S-LOXL2-MED1 interaction is fundamental for the proliferation of triple-negative breast cancer. Therefore, targeting such interaction holds potential for the development of novel triple-negative breast cancer therapies.

cancer biology↗

bioRxiv2019

The SWI/SNF subunits BRG1 affects alternative splicing by changing RNA binding factor interactions with RNA

BRG1 and BRM are ATPase core subunits of the human SWI/SNF chromatin remodelling complexes. The function of the SWI/SNF complexes in transcriptional initiation has been well studied, while a function in alternative splicing has only been studied for a few cases for BRM-containing SWI/SNF complexes. Here, we have expressed BRG1 in C33A cells, a BRG1 and BRM-deficient cell line, and we have analysed the effects on the transcriptome by RNA sequencing. We have shown that BRG1 expression affects the splicing of a subset of genes. For some, BRG1 expression favours exon inclusion and for others, exon skipping. Some of the changes in alternative splicing induced by BRG1 expression do not require the ATPase activity of BRG1. Among the exons regulated through an ATPase-independent mechanism, the included exons had signatures of high GC-content and lacked a positioned nucleosome at the exon. By investigating three genes in which the expression of either wild-type BRG1 or a BRG1-ATPase-deficient variant favoured exon inclusion, we showed that expression of the ATPases promotes the local recruitment of RNA binding factors to chromatin and RNA in a differential manner. The hnRNPL, hnRNPU and SAM68 proteins associated to chromatin in C33A cells expressing BRG1 or BRM, but their association with RNA varied. We propose that SWI/SNF can regulate alternative splicing by interacting with splicing-RNA binding factor and altering their binding to the nascent pre-mRNA, which changes RNP structure. Author summarySplicing, in particular alternative splicing, is a combinatorial process which involves splicing factor complexes and many RNA binding splicing regulatory proteins in different constellations. Most splicing events occur during transcription, which also makes the DNA sequence, the chromatin state and the transcription rate at the exons important components that influence the splicing outcome. We show here that the ATP-dependent chromatin remodelling complex SWI/SNF influences the interactions of splicing regulatory factors with RNA during transcription on certain exons that have a high GC-content. The splicing on this type of exon rely on the ATPase BRG1 and favour inclusion of alternative exons in an ATP-independent manner. SWI/SNF complexes are known to alter the chromatin structure at promoters in transcription initiation, and have been previously shown to alter the transcription rate or nucleosome position in splicing. Our results suggests a further mechanism for chromatin remodelling proteins in splicing: to change the interaction patterns of RNA binding splicing regulatory factors at alternative exons to alter the splicing outcome.

molecular biology↗