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Meng Wu,1,* Shuang Wu,1,* Rui Guo2 1Department of Pharmacy, the First Hospital of China Medical University, Shenyang, 110001, People’s Republic of China; 2Department of Critical Care Medicine, the First Hospital of China Medical University, Shenyang, 110001, People’s Republic of China*These authors contributed equally to this workCorrespondence: Rui Guo, Email 18840047020@163.comBackground: Breast cancer is the leading cause of cancer-related deaths among women worldwide. Identifying robust biomarkers for predicting outcomes is essential for improving patient care and reducing fatalities. ZMAT3, a zinc finger protein with potential carcinogenic properties, has been associated with various cancers. However, its role in breast cancer prognosis remains unclear.Methods: We investigated the expression level of ZMAT3 in breast cancer tissues and its association with clinical outcomes through bioinformatics analysis and experimental validation. We examined the correlation between ZMAT3 expression and immune characteristics. ZMAT3 mRNA expression data from The Cancer Genome Atlas (TCGA) were analysed in relation to overall survival (OS), disease-specific survival (DSS) and progression-free interval (PFI) in patients with breast cancer. Immunohistochemistry (IHC) was performed on breast cancer tissues to assess ZMAT3 protein levels, with findings validated using qPCR and cell experiments.Results: ZMAT3 mRNA levels were significantly upregulated in breast cancer samples compared to normal tissues. High ZMAT3 expression was significantly correlated with the poor OS, DSS and PFI. A significant positive correlation was observed between high ZMAT3 mRNA levels and the abundance of tumour-infiltrating lymphocytes (TILs), especially CD8+T cells and regulatory T cells (Tregs). Multivariate Cox regression analysis identified ZMAT3 as an independent prognostic factor for breast cancer. IHC staining confirmed increased ZMAT3 protein expression in breast cancer tissues, which was further validated by qPCR and cell function tests.Conclusion: Our findings suggest that ZMAT3 is a prognostic biomarker linked to immune invasion in breast cancer. Elevated ZMAT3 expression correlates with adverse clinical outcomes, indicating its potential role in disease progression.Keywords: breast cancer, cancer prognosis, immunotherapy, bioinformatics, ZMAT3

Although TP53 is the most commonly mutated gene in human cancers, the p53-dependent transcriptional programs mediating tumor suppression remain incompletely understood. Here, to uncover critical components downstream of p53 in tumor suppression, we perform unbiased RNAi and CRISPR-Cas9-based genetic screens in vivo. These screens converge upon the p53-inducible gene Zmat3, encoding an RNA-binding protein, and we demonstrate that ZMAT3 is an important tumor suppressor downstream of p53 in mouse KrasG12D-driven lung and liver cancers and human carcinomas. Integrative analysis of the ZMAT3 RNA-binding landscape and transcriptomic profiling reveals that ZMAT3 directly modulates exon inclusion in transcripts encoding proteins of diverse functions, including the p53 inhibitors MDM4 and MDM2, splicing regulators, and components of varied cellular processes. Interestingly, these exons are enriched in NMD signals, and, accordingly, ZMAT3 broadly affects target transcript stability. Collectively, these studies reveal ZMAT3 as a novel RNA-splicing and homeostasis regulator and a key component of p53-mediated tumor suppression.

Disruption of the estrous cycle affects fertility and reproductive health. Follicular dynamics are key to the regularity of the estrous cycle. We identified a novel lincRNA, HEOE, showing significant upregulation in the ovaries during the estrus phase across various pig breeds. Functional analysis revealed that HEOE is responsive to luteinizing hormone (LH) stimulation, modulating transcriptional suppression and alternative splicing in ovarian granulosa cells (GCs). This leads to increased GC apoptosis and inhibition of proliferation. Mechanistically, HEOE inhibits miR-16 maturation in the nucleus, and sequesters miR-16 in the cytoplasm, thereby collectively reducing miR-16's inhibition on ZMAT3, enhancing the expression of ZMAT3, a key factor in the p53 pathway and alternative splicing, thereby regulating follicular development. This effect was validated in both mice and pig follicles. Persistent overexpression or suppression of HEOE throughout the estrous cycle impairs cycle regularity and reduces litter size. These outcomes are associated with HEOE reduced follicular PGF2α levels and modulation of the cAMP signaling pathway. Our data, combined with public databases, indicate that the high expression of HEOE during the estrus phase is crucial for maintaining the estrous cycle. HEOE is a potential therapeutic target for regulating fertility and ensuring estrous cycle regularity in pigs., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

The tumor suppressor p53 and its antagonists MDM2 and MDM4 integrate stress signaling. For instance, dysbalanced assembly of ribosomes in nucleoli induces p53. Here, we show that the ribosomal protein L22 (RPL22; eL22), under conditions of ribosomal and nucleolar stress, promotes the skipping of MDM4 exon 6. Upon L22 depletion, more full-length MDM4 is maintained, leading to diminished p53 activity and enhanced cellular proliferation. L22 binds to specific RNA elements within intron 6 of MDM4 that correspond to a stem-loop consensus, leading to exon 6 skipping. Targeted deletion of these intronic elements largely abolishes L22-mediated exon skipping and re-enables cell proliferation, despite nucleolar stress. L22 also governs alternative splicing of the L22L1 (RPL22L1) and UBAP2L mRNAs. Thus, L22 serves as a signaling intermediate that integrates different layers of gene expression. Defects in ribosome synthesis lead to specific alternative splicing, ultimately triggering p53-mediated transcription and arresting cell proliferation. [Display omitted] • The ribosomal protein L22 binds to the MDM4 pre-mRNA and promotes exon 6 skipping • Consensus sequences in intron 6 are required for L22 binding to the MDM4 pre-mRNA • MDM4 splicing regulation by L22 causes p53 activation upon nucleolar stress • L22 also regulates the splicing of L22L1 and UBAP2L Jansen et al. show that the ribosomal protein L22 binds to the MDM4 pre-mRNA at intron 6 to promote exon 6 skipping. This leads to reduced levels of functional MDM4 when L22 relocalizes to the nucleoplasm upon nucleolar stress, thus representing a mechanism of p53 activation in this context. [ABSTRACT FROM AUTHOR]

Benzene is a common environmental and occupational pollutant, benzene exposure causes damage to hematopoietic system. ZMAT3 is a zinc finger protein which has important biological functions. In this study, benzene-exposed mouse model and ZMAT3 overexpression and low expression hematopoietic stem cells (HSCs) models were constructed to explore the mechanism of ZMAT3 in benzene-induced hematopoietic toxicity. The results showed that benzene increased the expression of ZMAT3 in mouse bone marrow (BM) cells, HSCs and peripheral blood (PB) leukocyte, and the changes in HSCs were more sensitive than BM and PB cells. In addition, overexpression of ZMAT3 decreased the self-renewal ability of HSCs and reduced the HSCs differentiation into myeloid hematopoietic cells, while low expression has the opposite effect. Besides, over and low expression of ZMAT3 both increased the HSCs differentiation into lymphoid progenitor cells. Moreover, bioinformatics analysis suggested that ZMAT3 was associated with TNF-α signaling pathway, and the correlation was confirmed in mouse model. Meanwhile, the results indicated that ZMAT3 promoted TNF-α mRNA processing by binding to the ARE structural domain on TNF-α and interacting with hnRNP A2/B1 and hnRNP A1 proteins, ultimately activating the NF-κB signaling pathway. This study provides a new mechanism for the study of benzene toxicity. [Display omitted] • Benzene induced abnormal increase of ZMAT3 in mouse HSCs, BM and PB leukocyte. • Overexpression of ZMAT3 in HSCs exacerbated the reduction of self-renewal ability. • Overexpression of ZMAT3 in HSCs reduced the differentiation into myeloid cells. • ZMAT3 regulated TNF-α expression, thereby activating the NF-κB signaling pathway. [ABSTRACT FROM AUTHOR]

Senescence of adipose precursor cells (APC) impairs adipogenesis, contributes to the age‐related subcutaneous adipose tissue (SAT) dysfunction, and increases risk of type 2 diabetes (T2D). First‐degree relatives of T2D individuals (FDR) feature restricted adipogenesis, reflecting the detrimental effects of APC senescence earlier in life and rendering FDR more vulnerable to T2D. Epigenetics may contribute to these abnormalities but the underlying mechanisms remain unclear. In previous methylome comparison in APC from FDR and individuals with no diabetes familiarity (CTRL), ZMAT3 emerged as one of the top‐ranked senescence‐related genes featuring hypomethylation in FDR and associated with T2D risk. Here, we investigated whether and how DNA methylation changes at ZMAT3 promote early APC senescence. APC from FDR individuals revealed increases in multiple senescence markers compared to CTRL. Senescence in these cells was accompanied by ZMAT3 hypomethylation, which caused ZMAT3 upregulation. Demethylation at this gene in CTRL APC led to increased ZMAT3 expression and premature senescence, which were reverted by ZMAT3 siRNA. Furthermore, ZMAT3 overexpression in APC determined senescence and activation of the p53/p21 pathway, as observed in FDR APC. Adipogenesis was also inhibited in ZMAT3‐overexpressing APC. In FDR APC, rescue of ZMAT3 methylation through senolytic exposure simultaneously downregulated ZMAT3 expression and improved adipogenesis. Interestingly, in human SAT, aging and T2D were associated with significantly increased expression of both ZMAT3 and the P53 senescence marker. Thus, DNA hypomethylation causes ZMAT3 upregulation in FDR APC accompanied by acquisition of the senescence phenotype and impaired adipogenesis, which may contribute to FDR predisposition for T2D. [ABSTRACT FROM AUTHOR]

The tumor protein p53 (TP53, best known as p53) transcription factor is a critical tumor suppressor, but those p53-inducible genes most important for tumor suppression have remained unclear. Using unbiased RNA interference and CRISPR (Clustered Regularly Interspersed Palindromic Repeats)/Cas9 (CRISPR-associated protein 9) screens, genetically engineered mouse models, human cancer genome analysis, and integrative eCLIP-sequencing and RNA-sequencing analyses, we reveal a new branch of p53-mediated tumor suppression involving the RNA splicing regulator Zinc finger Matrin-type 3, Zmat3.

Though primarily a tumor suppressor, TP53 harboring specific missense mutations located in the region encoding the DNA binding domain exhibits a gain of function by transcriptional activation of oncogenes. We performed microarray‐based messenger RNA profiling of squamous cell carcinoma of the oral tongue (SCCOT) and identified significant elevation of SMARCD1 in samples exhibiting p53 nuclear stabilization. Activation of SMARCD1 by mutant p53 was confirmed by evaluation of additional tongue cancer samples as well as The Cancer Genome Atlas expression datasets. SMARCD1 knockdown in HNSCC cells resulted in a significant reduction in several tumorigenic characteristics including cell viability, ability to form colonies in liquid and solid media and cell migration. We identified significantly increased SMARCD1 transcript levels in tumor versus matched normal samples in SCCOT as well as in other cancer types. Increased SMARCD1 expression predicted poor survival in HNSCC tumors harboring missense p53 mutations. Our results suggest SMARCD1 to be a novel transcriptional target of mutant p53. [ABSTRACT FROM AUTHOR]

Mutations in the TP53 tumour suppressor gene occur in half of all human cancers, indicating its critical importance in inhibiting cancer development. Despite extensive studies, the mechanisms by which mutant p53 enhances tumour progression remain only partially understood. Here, using data from the Cancer Genome Atlas ( TCGA), genomic and transcriptomic analyses were performed on 2256 tumours from 10 human cancer types. We show that tumours with TP53 mutations have altered gene expression profiles compared to tumours retaining two wild-type TP53 alleles. Among 113 known p53-up-regulated target genes identified from cell culture assays, 10 were consistently up-regulated in at least eight of 10 cancer types that retain both copies of wild-type TP53. RPS27L, CDKN1A ( p21CIP1) and ZMAT3 were significantly up-regulated in all 10 cancer types retaining wild-type TP53. Using this p53-based expression analysis as a discovery tool, we used cell-based assays to identify five novel p53 target genes from genes consistently up-regulated in wild-type p53 cancers. Global gene expression analyses revealed that cell cycle regulatory genes and transcription factors E2F1, MYBL2 and FOXM1 were disproportionately up-regulated in many TP53 mutant cancer types. Finally, > 93% of tumours with a TP53 mutation exhibited greatly reduced wild-type p53 messenger expression, due to loss of heterozygosity or copy neutral loss of heterozygosity, supporting the concept of p53 as a recessive tumour suppressor. The data indicate that tumours with wild-type TP53 retain some aspects of p53-mediated growth inhibitory signalling through activation of p53 target genes and suppression of cell cycle regulatory genes. Published by John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

The tumor protein p53 (TP53, best known as p53) transcription factor is a critical tumor suppressor, but those p53-inducible genes most important for tumor suppression have remained unclear. Using unbiased RNA interference and CRISPR (Clustered Regularly Interspersed Palindromic Repeats)/Cas9 (CRISPR-associated protein 9) screens, genetically engineered mouse models, human cancer genome analysis, and integrative eCLIP-sequencing and RNA-sequencing analyses, we reveal a new branch of p53-mediated tumor suppression involving the RNA splicing regulator Zinc finger Matrin-type 3, Zmat3., (© 2021 Taylor & Francis Group, LLC.)

p53 is an intensely studied tumor-suppressive transcription factor. Recent studies suggest that the RNA-binding protein (RBP) ZMAT3 is important in mediating the tumor-suppressive effects of p53. Here, we globally identify ZMAT3-regulated RNAs and their binding sites at nucleotide resolution in intact colorectal cancer (CRC) cells. ZMAT3 binds to thousands of mRNA precursors, mainly at intronic uridine-rich sequences and affects their splicing. The strongest alternatively spliced ZMAT3 target was CD44 , a cell adhesion gene and stem cell marker that controls tumorigenesis. Silencing ZMAT3 increased inclusion of CD44 variant exons, resulting in significant up-regulation of oncogenic CD44 isoforms ( CD44v ) and increased CRC cell growth that was rescued by concurrent knockdown of CD44v Silencing p53 phenocopied the loss of ZMAT3 with respect to CD44 alternative splicing, suggesting that ZMAT3-mediated regulation of CD44 splicing is vital for p53 function. Collectively, our findings uncover a p53-ZMAT3-CD44 axis in growth suppression in CRC cells., (© 2021 Muys et al.; Published by Cold Spring Harbor Laboratory Press.)