p52-ZER6/IGF1R axis maintains cancer stem cell population to promote cancer progression by enhancing pro-survival mitophagy.

Cancer stem cells (CSCs), which are distinct subpopulations of tumor cells, have a substantially higher tumor-initiating capacity and are closely related to poor clinical outcomes. Damage to organelles can trigger CSC pool exhaustion; however, the underlying mechanisms are poorly understood. ZER6 is a zinc-finger protein with two isoforms possessing different amino termini: p52-ZER6 and p71-ZER6. Since their discovery, almost no study reported on their biological and pathological functions. Herein, we found that p52-ZER6 was crucial for CSC population maintenance; p52-ZER6-knocking down almost abolished the tumor initiation capability. Through transcriptomic analyses together with in vitro and in vivo studies, we identified insulin like growth factor 1 receptor (IGF1R) as the transcriptional target of p52-ZER6 that mediated p52-ZER6 regulation of CSC by promoting pro-survival mitophagy. Moreover, this regulation of mitophagy-mediated CSC population maintenance is specific to p52-ZER6, as p71-ZER6 failed to exert the same effect, most possibly due to the presence of the HUB1 domain at its N-terminus. These results provide a new perspective on the regulatory pathway of pro-survival mitophagy in tumor cells and the molecular mechanism underlying p52-ZER6 oncogenic activity, suggesting that targeting p52-ZER6/IGF1R axis to induce CSC pool exhaustion may be a promising anti-tumor therapeutic strategy.

Correction: Histone Demethylase JMJD2B Functions as a Co-Factor of Estrogen Receptor in Breast Cancer Proliferation and Mammary Gland Development.

[This corrects the article DOI: 10.1371/journal.pone.0017830.].

YY2/BUB3 Axis promotes SAC Hyperactivation and Inhibits Colorectal Cancer Progression via Regulating Chromosomal Instability.

Spindle assembly checkpoint (SAC) is a crucial safeguard mechanism of mitosis fidelity that ensures equal division of duplicated chromosomes to the two progeny cells. Impaired SAC can lead to chromosomal instability (CIN), a well-recognized hallmark of cancer that facilitates tumor progression; paradoxically, high CIN levels are associated with better therapeutic response and prognosis. However, the mechanism by which CIN determines tumor cell survival and therapeutic response remains poorly understood. Here, using a cross-omics approach, YY2 is identified as a mitotic regulator that promotes SAC activity by activating the transcription of budding uninhibited by benzimidazole 3 (BUB3), a component of SAC. While both conditions induce CIN, a defect in YY2/SAC activity enhances mitosis and tumor growth. Meanwhile, hyperactivation of SAC mediated by YY2/BUB3 triggers a delay in mitosis and suppresses growth. Furthermore, it is revealed that YY2/BUB3-mediated excessive CIN causes higher cell death rates and drug sensitivity, whereas residual tumor cells that survived DNA damage-based therapy have moderate CIN and increased drug resistance. These results provide insights into the role of SAC activity and CIN levels in influencing tumor cell survival and drug response, as well as suggest a novel anti-tumor therapeutic strategy that combines SAC activity modulators and DNA-damage agents.

YY2-DRP1 Axis Regulates Mitochondrial Fission and Determines Cancer Stem Cell Asymmetric Division.

Cancer stem cells (CSCs) are associated with tumor progression, recurrence, and therapeutic resistance. To maintain their pool while promoting tumorigenesis, CSCs divide asymmetrically, producing a CSC and a highly proliferative, more differentiated transit-amplifying cell. Exhausting the CSC pool has been proposed as an effective antitumor strategy; however, the mechanism underlying CSC division remains poorly understood, thereby largely limiting its clinical application. Here, through cross-omics analysis, yin yang 2 (YY2) is identified as a novel negative regulator of CSC maintenance. It is shown that YY2 is downregulated in stem-like tumor spheres formed by hepatocarcinoma cells and in liver cancer, in which its expression is negatively correlated with disease progression and poor prognosis. Furthermore, it is revealed that YY2 overexpression suppressed liver CSC asymmetric division, leading to depletion of the CSC pool and decreased tumor-initiating capacity. Meanwhile, YY2 knock-out in stem-like tumor spheres caused enrichment in mitochondrial functions. Mechanistically, it is revealed that YY2 impaired mitochondrial fission, and consequently, liver CSC asymmetric division, by suppressing the transcription of dynamin-related protein 1. These results unravel a novel regulatory mechanism of mitochondrial dynamic-mediated CSCs asymmetric division and highlight the role of YY2 as a tumor suppressor and a therapeutic target in antitumor treatment.

YY2/PHGDH axis suppresses tumorigenesis by inhibiting tumor cell de novo serine biosynthesis.

Metabolic reprogramming is one of the key features of tumors facilitating their rapid proliferation and adaptation to harsh microenvironments. Yin Yang 2 (YY2) has recently been reported as a tumor suppressor downregulated in various types of tumors; however, the molecular mechanisms underlying its tumor-suppressive activity remain poorly understood. Furthermore, the involvement of YY2 in tumor cell metabolic reprogramming remains unclear. Herein, we aimed to elucidate the novel regulatory mechanism of YY2 in the suppression of tumorigenesis. Using transcriptomic analysis, we uncovered an unprecedented link between YY2 and tumor cell serine metabolism. YY2 alteration could negatively regulate the expression level of phosphoglycerate dehydrogenase (PHGDH), the first enzyme in the serine biosynthesis pathway, and consequently, tumor cell de novo serine biosynthesis. Mechanistically, we revealed that YY2 binds to the PHGDH promoter and suppresses its transcriptional activity. This, in turn, leads to decreased production of serine, nucleotides, and cellular reductants NADH and NADPH, which subsequently suppresses tumorigenic potential. These findings reveal a novel function of YY2 as a regulator of the serine metabolic pathway in tumor cells and provide new insights into its tumor suppressor activity. Furthermore, our findings suggest the potential of YY2 as a target for metabolic-based antitumor therapeutic strategies.

The p52-ZER6/G6PD axis alters aerobic glycolysis and promotes tumor progression by activating the pentose phosphate pathway.

Abnormal glucose metabolism is a highlight of tumor metabolic reprogramming and is closely related to the development of malignancies. p52-ZER6, a C2H2-type zinc finger protein, promotes cell proliferation and tumorigenesis. However, its role in the regulation of biological and pathological functions remains poorly understood. Here, we examined the role of p52-ZER6 in tumor cell metabolic reprogramming. Specifically, we demonstrated that p52-ZER6 promotes tumor glucose metabolic reprogramming by positively regulating the transcription of glucose-6-phosphate dehydrogenase (G6PD), the rate-limiting enzyme in the pentose phosphate pathway (PPP). By activating the PPP, p52-ZER6 was found to enhance the production of nucleotides and nicotinamide adenine dinucleotide phosphate, thereby providing tumor cells with the building blocks of ribonucleic acids and cellular reductants for reactive oxygen species scavenging, which subsequently promotes tumor cell proliferation and viability. Importantly, p52-ZER6 promoted PPP-mediated tumorigenesis in a p53-independent manner. Taken together, these findings reveal a novel role for p52-ZER6 in regulating G6PD transcription via a p53-independent process, ultimately resulting in tumor cell metabolic reprogramming and tumorigenesis. Our results suggest that p52-ZER6 is a potential target for the diagnosis and treatment of tumors and metabolic disorders.

Surface modulation of extracellular vesicles with cell-penetrating peptide-conjugated lipids for improvement of intracellular delivery to endothelial cells.

Exosomes (diameter 30-200 nm) are a subtype of extracellular vesicles secreted by cells containing DNA, microRNA (miRNA), and proteins. Exosomes are expected to be valuable as a means of delivering drugs or functional miRNAs in treatment of diseases. However, the delivery of exosomes is not sufficiently effective, even though exosomes have intrinsic delivery functions. Cell-penetrating peptides (CPPs) are short peptide families that facilitate cellular intake of molecules and vesicles. We previously reported that the modification of cells, and liposomes with CPP-conjugated-lipids, CPPs conjugated with poly (ethylene glycol)-conjugated phospholipids (PEG-lipid), that induce adhesion by CPPs, can be useful for cell-based assays and harvesting liposomes. In this study, we aimed to modulate the exosome surface using Tat peptide (YGRKKRRQRRR)-PEG-lipids to improve intracellular delivery to endothelial cells. We isolated and characterized exosomes from the medium of HEK 293 T cell cultures. Tat conjugated PEG-lipids with different spacer molecular weights and lipid types were incorporated into exosomes using fluorescein isothiocyanate labeling to optimize the number of Tat-PEG-lipids immobilized on the exosome surface. The exosomes modified with Tat-PEG-lipids were incubated with human umbilical vein endothelial cells (HUVECs) to study the interaction. Tat conjugated with 5 kDa PEG and C16 lipids incorporated on the exosome surface were highly detected inside HUVECs by flow cytometry. Fluorescence was negligible in HUVECs for control groups. Thus, Tat-PEG-lipids can be modified on the exosome surface, improving the intracellular delivery of exosomes.

SGLT2 inhibitor empagliflozin promotes revascularization in diabetic mouse hindlimb ischemia by inhibiting ferroptosis.

Gliflozins are known as SGLT2 inhibitors, which are used to treat diabetic patients by inhibiting glucose reabsorption in kidney proximal tubules. Recent studies show that gliflozins may exert other effects independent of SGLT2 pathways. In this study we investigated their effects on skeletal muscle cell viability and paracrine function, which were crucial for promoting revascularization in diabetic hindlimb ischemia (HLI). We showed that treatment with empagliflozin (0.1-40 µM) dose-dependently increased high glucose (25 mM)-impaired viability of skeletal muscle C2C12 cells. Canagliflozin, dapagliflozin, ertugliflozin, ipragliflozin and tofogliflozin exerted similar protective effects on skeletal muscle cells cultured under the hyperglycemic condition. Transcriptomic analysis revealed an enrichment of pathways related to ferroptosis in empagliflozin-treated C2C12 cells. We further demonstrated that empagliflozin and other gliflozins (10 µM) restored GPX4 expression in high glucose-treated C2C12 cells, thereby suppressing ferroptosis and promoting cell viability. Empagliflozin (10 µM) also markedly enhanced the proliferation and migration of blood vessel-forming cells by promoting paracrine function of skeletal muscle C2C12 cells. In diabetic HLI mice, injection of empagliflozin into the gastrocnemius muscle of the left hindlimb (10 mg/kg, every 3 days for 21 days) significantly enhanced revascularization and blood perfusion recovery. Collectively, these results reveal a novel effect of empagliflozin, a clinical hypoglycemic gliflozin drug, in inhibiting ferroptosis and enhancing skeletal muscle cell survival and paracrine function under hyperglycemic condition via restoring the expression of GPX4. This study highlights the potential of intramuscular injection of empagliflozin for treating diabetic HLI.

p52-ZER6: a determinant of tumor cell sensitivity to MDM2-p53 binding inhibitors.

Targeting MDM2-p53 interaction has emerged as a promising antitumor therapeutic strategy. Several MDM2-p53 inhibitors have advanced into clinical trials, but results are not favorable. The lack of appropriate biomarkers for selecting patients has been assumed as the critical reason for this failure. We previously identified ZER6 isoform p52-ZER6 as an oncogene upregulated in tumor tissues. In this study we investigated whether p52-ZER6 acted as a blocker of MDM2-p53 binding inhibitors, and whether p52-ZER6 could be used as a biomarker of MDM2-p53 binding inhibitors. In p53 wild-type colorectal carcinoma HCT116, hepatocarcinoma HepG2 and breast cancer MCF-7 cells, overexpression of p52-ZER6 enhanced MDM2-p53 binding and promoted p53 ubiquitination/proteasomal degradation. Furthermore, overexpression of p52-ZER6 in the tumor cells dose-dependently reduced their sensitivity to both nutlin and non-nutlin class MDM2-p53 binding inhibitors. We showed that p52-ZER6 restored tumor cell viability, which was suppressed by nutlin-3, through restoring their proliferation potential while suppressing their apoptotic rate, suggesting that MDM2-p53 binding inhibitors might not be effective for patients with high p52-ZER6 levels. We found that nutlin-3 treatment or p52-ZER6 knockdown alone promoted the accumulation of p53 protein in the tumor cells, and their combinatorial treatment significantly increased the accumulation of p53 protein. In HCT116 cell xenograft nude mouse model, administration of shp52-ZER6 combined with an MDM2-p53 binding inhibitor nutlin-3 exerted synergistic antitumor response. In conclusion, this study reveals that p52-ZER6 might be a potential biomarker for determining patients appropriate for MDM2-p53 binding inhibition-based antitumor therapy, and demonstrates the potential of combinatorial therapy using MDM2-p53 binding inhibitors and p52-ZER6 inhibition.

Unspliced XBP1 contributes to cholesterol biosynthesis and tumorigenesis by stabilizing SREBP2 in hepatocellular carcinoma.

Cholesterol biosynthesis plays a critical role in rapidly proliferating tumor cells. X-box binding protein 1 (XBP1), which was first characterized as a basic leucine zipper-type transcription factor, exists in an unspliced (XBP1-u) and spliced (XBP1-s) form. Recent studies showed that unspliced XBP1 (XBP1-u) has unique biological functions independent from XBP1-s and could promote tumorigenesis; however, whether it is involved in tumor metabolic reprogramming remains unknown. Herein, we found that XBP1-u promotes tumor growth by enhancing cholesterol biosynthesis in hepatocellular carcinoma (HCC) cells. Specifically, XBP1-u colocalizes with sterol regulatory element-binding protein 2 (SREBP2) and inhibits its ubiquitination/proteasomal degradation. The ensuing stabilization of SREBP2 activates the transcription of 3-hydroxy-3-methylglutaryl-CoA reductase (HMGCR), a rate-limiting enzyme in cholesterol biosynthesis. We subsequently show that the XBP1-u/SREBP2/HMGCR axis is crucial for enhancing cholesterol biosynthesis and lipid accumulation as well as tumorigenesis in HCC cells. Taken together, these findings reveal a novel function of XBP1-u in promoting tumorigenesis through increased cholesterol biosynthesis in hepatocarcinoma cells. Hence, XBP1-u might be a potential target for anti-tumor therapeutic strategies that focus on cholesterol metabolism in HCC.

Homeostasis Imbalance of YY2 and YY1 Promotes Tumor Growth by Manipulating Ferroptosis.

Ferroptosis is a type of programmed cell death caused by disruption of redox homeostasis and is closely linked to amino acid metabolism. Yin Yang 2 (YY2) and its homolog Yin Yang 1 (YY1) are highly homologous, especially in their zinc-finger domains. Furthermore, they share a consensus DNA binding motif. Increasing evidences have demonstrated the tumor suppressive effect of YY2, in contrast with the oncogenic YY1; however, little is known about the biological and pathological functions of YY2. Here, it is determined that YY2 induces tumor cell ferroptosis and subsequently suppresses tumorigenesis by inhibiting solute carrier family 7 member 11 (SLC7A11) transcription, leading to the decreased glutathione biosynthesis. Furthermore, YY2 and YY1 bind competitively to the same DNA binding site in the SLC7A11 promoter and antagonistically regulate tumor cell ferroptosis, thus suggesting the molecular mechanism underlying their opposite regulation on tumorigenesis. Moreover, mutations of YY2 zinc-finger domains in clinical cancer patients abrogate YY2/SLC7A11 axis and tumor cell ferroptosis. Together, these results provide a new insight regarding the regulatory mechanism of ferroptosis, and a mechanistic explanation regarding the tumor suppressive effect of YY2. Finally, these findings demonstrate that homeostasis between YY1 and YY2 is crucial for maintaining redox homeostasis in tumor cells.

Discovery of Salidroside-Derivated Glycoside Analogues as Novel Angiogenesis Agents to Treat Diabetic Hind Limb Ischemia.

Therapeutic angiogenesis is a potential therapeutic strategy for hind limb ischemia (HLI); however, currently, there are no small-molecule drugs capable of inducing it at the clinical level. Activating the hypoxia-inducible factor-1 (HIF-1) pathway in skeletal muscle induces the secretion of angiogenic factors and thus is an attractive therapeutic angiogenesis strategy. Using salidroside, a natural glycosidic compound as a lead, we performed a structure-activity relationship (SAR) study for developing a more effective and druggable angiogenesis agent. We found a novel glycoside scaffold compound (C-30) with better efficacy than salidroside in enhancing the accumulation of the HIF-1α protein and stimulating the paracrine functions of skeletal muscle cells. This in turn significantly increased the angiogenic potential of vascular endothelial and smooth muscle cells and, subsequently, induced the formation of mature, functional blood vessels in diabetic and nondiabetic HLI mice. Together, this study offers a novel, promising small-molecule-based therapeutic strategy for treating HLI.

NeuroD1 promotes tumor cell proliferation and tumorigenesis by directly activating the pentose phosphate pathway in colorectal carcinoma.

Tumor metabolic reprogramming ensures that cancerous cells obtain sufficient building blocks, energy, and antioxidants to sustain rapid growth and for coping with oxidative stress. Neurogenic differentiation factor 1 (NeuroD1) is upregulated in various types of tumors; however, its involvement in tumor cell metabolic reprogramming remains unclear. In this study, we report that NeuroD1 is positively correlated with glucose-6-phosphate dehydrogenase (G6PD), the rate-limiting enzyme in the pentose phosphate pathway (PPP), in colorectal cancer cells. In addition, the regulation of G6PD by NeuroD1 alters tumor cell metabolism by stimulating the PPP, leading to enhanced production of nucleotides and NADPH. These, in turn, promote DNA and lipid biosynthesis in tumor cells, while decreasing intracellular levels of reactive oxygen species. Mechanistically, we showed that NeuroD1 binds directly to the G6PD promoter to activate G6PD transcription. Consequently, tumor cell proliferation and colony formation are enhanced, leading to increased tumorigenic potential in vitro and in vivo. These findings reveal a novel function of NeuroD1 as a regulator of G6PD, whereby its oncogenic activity is linked to tumor cell metabolic reprogramming and regulation of the PPP. Furthermore, NeuroD1 represents a potential target for metabolism-based anti-tumor therapeutic strategies.

Zn2+-dependent DNAzymes that cleave all combinations of ribonucleotides.

Although several DNAzymes are known, their utility is limited by a narrow range of substrate specificity. Here, we report the isolation of two zinc-dependent DNAzymes, ZincDz1 and ZincDz2, which exhibit compact catalytic core sequences with highly versatile hydrolysis activity. They were selected through in vitro selection followed by deep sequencing analysis. Despite their sequence similarity, each DNAzyme showed different Zn2+-concentration and pH-dependent reaction profiles, and cleaved the target RNA sequences at different sites. Using various substrate RNA sequences, we found that the cleavage sequence specificity of ZincDz2 and its highly active mutant ZincDz2-v2 to be 5'-rN↓rNrPu-3'. Furthermore, we demonstrated that the designed ZincDz2 could cut microRNA miR-155 at three different sites. These DNAzymes could be useful in a broad range of applications in the fields of medicine and biotechnology.

The transcription factor PBX3 promotes tumor cell growth through transcriptional suppression of the tumor suppressor p53.

Pre-B-cell leukemia transcription factor 3 (PBX3) is a member of the PBX family and contains a highly conserved homologous domain. PBX3 is involved in the progression of gastric cancer, colorectal cancer, and prostate cancer; however, the detailed mechanism by which it promotes tumor growth remains to be elucidated. Here, we found that PBX3 silencing induces the expression of the cell cycle regulator p21, leading to an increase in colorectal cancer (CRC) cell apoptosis as well as suppression of proliferation and colony formation. Furthermore, we found that PBX3 is highly expressed in clinical CRC patients, in whom p21 expression is aberrantly low. We found that the regulation of p21 transcription by PBX3 occurs through the upstream regulator of p21, the tumor suppressor p53, as PBX3 binds to the p53 promoter and suppresses its transcriptional activity. Finally, we revealed that PBX3 regulates tumor growth through regulation of the p53/p21 axis. Taken together, our results not only describe a novel mechanism regarding PBX3-mediated regulation of tumor growth but also provide new insights into the regulatory mechanism of the tumor suppressor p53.

Development of aptamer-based inhibitors for CRISPR/Cas system.

The occurrence of accidental mutations or deletions caused by genome editing with CRISPR/Cas9 system remains a critical unsolved problem of the technology. Blocking excess or prolonged Cas9 activity in cells is considered as one means of solving this problem. Here, we report the development of an inhibitory DNA aptamer against Cas9 by means of in vitro selection (systematic evolution of ligands by exponential enrichment) and subsequent screening with an in vitro cleavage assay. The inhibitory aptamer could bind to Cas9 at low nanomolar affinity and partially form a duplex with CRISPR RNA, contributing to its inhibitory activity. We also demonstrated that improving the inhibitory aptamer with locked nucleic acids efficiently suppressed Cas9-directed genome editing in cells and reduced off-target genome editing. The findings presented here might enable the development of safer and controllable genome editing for biomedical research and gene therapy.

Comparative Glycomic Analysis of Exosome Subpopulations Derived from Pancreatic Cancer Cell Lines.

Extracellular vesicles such as exosomes are generally covered with an array of glycans, which are controlled by the host-cell glyco-synthetic machinery, similar to secreted and membrane glycoproteins. Several exosome subpopulations classified by their tetraspanin expression have been investigated in the context of diseases. However, a comparative analysis of their glycomics has never been attempted. Herein, we report a method for the comparative glycomic analysis of exosome subpopulations among pancreatic cancer cell lines. Glycomic profiles were obtained for extracellular vesicles, secreted glycoproteins, and membrane glycoproteins from eight cell lines. Statistical analyses revealed high populations of PHA-L-binding proteins in the vesicles. The surfaces of extracellular vesicles were labeled with Cy3 and captured by magnetic beads with antibodies against tetraspanins (CD9, CD63, and CD81). The coprecipitated vesicles were lysed and subjected to a lectin microarray analysis. A hierarchical clustering analysis using 19 glycomic profiles confirmed that most subpopulations, except CD81-positive exosomes, could be distinguished according to the host-cell species. Principal component analysis and subsequent lectin-affinity capturing of intact exosomes highlighted that CD81-positive exosomes preferentially expressed not PHA-L- but LEL-binding proteins on their surfaces. These data suggested that exosomal glycomics depended on the host-cell type and subpopulation.

Enhanced affinity of racemic phosphorothioate DNA with transcription factor SATB1 arising from diastereomer-specific hydrogen bonds and hydrophobic contacts.

Phosphorothioate modification is commonly introduced into therapeutic oligonucleotides, typically as a racemic mixture in which either of the two non-bridging phosphate oxygens is replaced by sulfur, which frequently increases affinities with proteins. Here, we used isothermal titration calorimetry and X-ray crystallography to investigate the thermodynamic and structural properties of the interaction between the primary DNA-binding domain (CUTr1) of transcription factor SATB1 and dodecamer DNAs with racemic phosphorothioate modifications at the six sites known to contact CUTr1 directly. For both the modified and unmodified DNAs, the binding reactions were enthalpy-driven at a moderate salt concentration (50 mM NaCl), while being entropy-driven at higher salt concentrations with reduced affinities. The phosphorothioate modifications lowered this susceptibility to salt, resulting in a significantly enhanced affinity at a higher salt concentration (200 mM NaCl), although only some DNA molecular species remained interacting with CUTr1. This was explained by unequal populations of the two diastereomers in the crystal structure of the complex of CUTr1 and the phosphorothioate-modified DNA. The preferred diastereomer formed more hydrogen bonds with the oxygen atoms and/or more hydrophobic contacts with the sulfur atoms than the other, revealing the origins of the enhanced affinity.

Neurogenic differentiation factor 1 promotes colorectal cancer cell proliferation and tumorigenesis by suppressing the p53/p21 axis.

Neurogenic differentiation factor 1 (NeuroD1) is a transcription factor critical for promoting neuronal differentiation and maturation. NeuroD1 is involved in neuroblastoma and medulloblastoma; however, its molecular mechanism in promoting tumorigenesis remains unclear. Furthermore, the role of NeuroD1 in non-neural malignancies has not been widely characterized. Here, we found that NeuroD1 is highly expressed in colorectal cancer. NeuroD1-silencing induces the expression of p21, a master regulator of the cell cycle, leading to G2 -M phase arrest and suppression of colorectal cancer cell proliferation as well as colony formation potential. Moreover, NeuroD1-mediated regulation of p21 expression occurs in a p53-dependent manner. Through chromatin immunoprecipitation and point mutation analysis in the predicted NeuroD1 binding site of the p53 promoter, we found that NeuroD1 directly binds to the p53 promoter and suppresses its transcription, resulting in increased p53 expression in NeuroD1-silenced colorectal cancer cells. Finally, xenograft experiments demonstrated that NeuroD1-silencing suppresses colorectal cancer cell tumorigenesis potential by modulating p53 expression. These findings reveal NeuroD1 as a novel regulator of the p53/p21 axis, underscoring its importance in promoting non-neural malignancies. Furthermore, this study provides insight into the transcriptional regulation of p53.

Zinc-finger protein p52-ZER6 accelerates colorectal cancer cell proliferation and tumour progression through promoting p53 ubiquitination.

BACKGROUND: Aberrant expression of p53 and its downstream gene p21 is closely related to alterations in cell cycle and cell proliferation, and is common among cancer patients. However, the underlying molecular mechanism has not been fully unravelled. ZER6 is a zinc-finger protein with two isoforms possessing different amino termini (N-termini) in their proteins, p52-ZER6 and p71-ZER6. The biological function of ZER6 isoforms, as well as their potential involvement in tumourigenesis and the regulation of p53 remain elusive. METHODS: The effect of ZER6 isoforms on p53 and p21 was determined using specific knockdown and overexpression. p52-ZER6 expression in tumours was analysed using clinical specimens, while gene modulation was used to explore p52-ZER6 roles in regulating cell proliferation and tumourigenesis. The mechanism of p52-ZER6 regulation on the p53/p21 axis was studied using molecular biology and biochemical methods. FINDINGS: p52-ZER6 was highly expressed in tumour tissues, and was closely related with tumour progression. Mechanistically, p52-ZER6 bound to p53 through a truncated KRAB (tKRAB) domain in its N-terminus and enhanced MDM2/p53 complex integrity, leading to increased p53 ubiquitination and degradation. p52-ZER6-silencing induced G0-G1 phase arrest, and subsequently reduced cell proliferation and tumourigenesis. Intriguingly, this regulation on p53 was specific to p52-ZER6, whereas p71-ZER6 did not affect p53 stability, most likely due to the presence of a HUB-1 domain. INTERPRETATION: We identified p52-ZER6 as a novel oncogene that enhances MDM2/p53 complex integrity, and might be a potential target for anti-cancer therapy.