Putting a finger on histidine methylation.

Specialized enzymes add methyl groups to the nitrogens of the amino acid histidine, altering the chemical properties of its imidazole ring and, in turn, the function of the modified (poly)peptide. In this issue of Genes & Development, Shimazu and colleagues (pp. 724-742) make the remarkable discovery that CARNMT1 acts as a dual-specificity histidine methyltransferase, modifying both the small-molecule dipeptide carnosine and a set of proteins, predominantly within RNA-binding C3H zinc finger (C3H ZF) motifs. As a result, CARNMT1 modulates the activity of its protein targets to affect RNA processing and metabolism, ultimately contributing an essential function during mammalian development.

Histidine N1-position-specific methyltransferase CARNMT1 targets C3H zinc finger proteins and modulates RNA metabolism.

Histidine (His) residues are methylated in various proteins, but their roles and regulation mechanisms remain unknown. Here, we show that carnosine N-methyltransferase 1 (CARNMT1), a known His methyltransferase of dipeptide carnosine (ßAla-His), is a major His N1-position-specific methyltransferase. We found that 52 His sites in 20 proteins underwent CARNMT1-mediated methylation. The consensus methylation site for CARNMT1 was identified as Cx(F/Y)xH, a C3H zinc finger (C3H ZF) motif. CARNMT1-deficient and catalytically inactive mutant mice showed embryonic lethality. Among the CARNMT1 target C3H ZF proteins, RNA degradation mediated by Roquin and tristetraprolin (TTP) was affected by CARNMT1 and its enzymatic activity. Furthermore, the recognition of the 3' splice site of the CARNMT1 target C3H ZF protein U2AF1 was perturbed, and pre-mRNA alternative splicing (AS) was affected by CARNMT1 deficiency. These findings indicate that CARNMT1-mediated protein His methylation, which is essential for embryogenesis, plays roles in diverse aspects of RNA metabolism by targeting C3H ZF-type RNA-binding proteins and modulating their functions, including pre-mRNA AS and mRNA degradation regulation.

Maintenance of persistent transmission of a plant arbovirus in its insect vector mediated by the Toll-Dorsal immune pathway.

Throughout evolution, arboviruses have developed various strategies to counteract the host's innate immune defenses to maintain persistent transmission. Recent studies have shown that, in addition to bacteria and fungi, the innate Toll-Dorsal immune system also plays an essential role in preventing viral infections in invertebrates. However, whether the classical Toll immune pathway is involved in maintaining the homeostatic process to ensure the persistent and propagative transmission of arboviruses in insect vectors remain unclear. In this study, we revealed that the transcription factor Dorsal is actively involved in the antiviral defense of an insect vector (Laodelphax striatellus) by regulating the target gene, zinc finger protein 708 (LsZN708), which mediates downstream immune-related effectors against infection with the plant virus (Rice stripe virus, RSV). In contrast, an antidefense strategy involving the use of the nonstructural-protein (NS4) to antagonize host antiviral defense through competitive binding to Dorsal from the MSK2 kinase was employed by RSV; this competitive binding inhibited Dorsal phosphorylation and reduced the antiviral response of the host insect. Our study revealed the molecular mechanism through which Toll-Dorsal-ZN708 mediates the maintenance of an arbovirus homeostasis in insect vectors. Specifically, ZN708 is a newly documented zinc finger protein targeted by Dorsal that mediates the downstream antiviral response. This study will contribute to our understanding of the successful transmission and spread of arboviruses in plant or invertebrate hosts.

ZNF91 is an endogenous repressor of the molecular phenotype associated with X-linked dystonia-parkinsonism (XDP).

X-linked dystonia-parkinsonism (XDP) is a severe neurodegenerative disorder resulting from an inherited intronic SINE-Alu-VNTR (SVA) retrotransposon in the TAF1 gene that causes dysregulation of TAF1 transcription. The specific mechanism underlying this dysregulation remains unclear, but it is hypothesized to involve the formation of G-quadruplexes (G4) structures within the XDP-SVA that impede transcription. In this study, we show that ZNF91, a critical repressor of SVA retrotransposons, specifically binds to G4-forming DNA sequences. Further, we found that genetic deletion of ZNF91 exacerbates the molecular phenotype associated with the XDP-SVA insertion in patient cells, while no difference was observed when ZNF91 was deleted from isogenic control cells. Additionally, we observed a significant age-related reduction in ZNF91 expression in whole blood and brain, indicating a progressive loss of repression of the XDP-SVA in XDP. These findings indicate that ZNF91 plays a crucial role in controlling the molecular phenotype associated with XDP. Since ZNF91 binds to G4-forming DNA sequences in SVAs, this suggests that interactions between ZNF91 and G4-forming sequences in the XDP-SVA minimize the severity of the molecular phenotype. Our results showing that ZNF91 expression levels significantly decrease with age provide a potential explanation for the age-related progressive neurodegenerative character of XDP. Collectively, our study provides important insights into the protective role of ZNF91 in XDP pathogenesis and suggests that restoring ZNF91 expression, destabilization of G4s, or targeted repression of the XDP-SVA could be future therapeutic strategies to prevent or treat XDP.

ZBTB20 Regulates SERCA2a Activity and Myocardial Contractility Through Phospholamban.

BACKGROUND: Intracellular Ca2+ cycling determines myocardial contraction and relaxation in response to physiological demands. SERCA2a (sarcoplasmic/endoplasmic reticulum Ca2+-ATPase 2a) is responsible for the sequestration of cytosolic Ca2+ into intracellular stores during cardiac relaxation, and its activity is reversibly inhibited by PLN (phospholamban). However, the regulatory hierarchy of SERCA2a activity remains unclear. METHODS: Cardiomyocyte-specific ZBTB20 knockout mice were generated by crossing ZBTB20flox mice with Myh6-Cre mice. Echocardiography, blood pressure measurements, Langendorff perfusion, histological analysis and immunohistochemistry, quantitative reverse transcription-PCR, Western blot analysis, electrophysiological measurements, and chromatin immunoprecipitation assay were performed to clarify the phenotype and elucidate the molecular mechanisms. RESULTS: Specific ablation of ZBTB20 in cardiomyocyte led to a significant increase in basal myocardial contractile parameters both in vivo and in vitro, accompanied by an impairment in cardiac reserve and exercise capacity. Moreover, the cardiomyocytes lacking ZBTB20 showed an increase in sarcoplasmic reticular Ca2+ content and exhibited a remarkable enhancement in both SERCA2a activity and electrically stimulated contraction. Mechanistically, PLN expression was dramatically reduced in cardiomyocytes at the mRNA and protein levels by ZBTB20 deletion or silencing, and PLN overexpression could largely restore the basal contractility in ZBTB20-deficient cardiomyocytes. CONCLUSIONS: These data point to ZBTB20 as a fine-tuning modulator of PLN expression and SERCA2a activity, thereby offering new perspective on the regulation of basal contractility in the mammalian heart.

The Plasmodium falciparum CCCH zinc finger protein MD3 regulates male gametocytogenesis through its interaction with RNA-binding proteins.

The transmission of malaria parasites to mosquitoes is dependent on the formation of gametocytes. Once fully matured, gametocytes are able to transform into gametes in the mosquito's midgut, a process accompanied with their egress from the enveloping erythrocyte. Gametocyte maturation and gametogenesis require a well-coordinated gene expression program that involves a wide spectrum of regulatory proteins, ranging from histone modifiers to transcription factors to RNA-binding proteins. Here, we investigated the role of the CCCH zinc finger protein MD3 in Plasmodium falciparum gametocytogenesis. MD3 was originally identified as an epigenetically regulated protein of immature gametocytes and recently shown to be involved in male development in a barcode-based screen in P. berghei. We report that MD3 is mainly present in the cytoplasm of immature male P. falciparum gametocytes. Parasites deficient of MD3 are impaired in gametocyte maturation and male gametocytogenesis. BioID analysis in combination with co-immunoprecipitation assays unveiled an interaction network of MD3 with RNA-binding proteins like PABP1 and ALBA3, with translational initiators, regulators and repressors like elF4G, PUF1, NOT1 and CITH, and with further regulators of gametocytogenesis, including ZNF4, MD1 and GD1. We conclude that MD3 is part of a regulator complex crucial for post-transcriptional fine-tuning of male gametocytogenesis.

blf and the drl cluster synergistically regulate cell fate commitment during zebrafish primitive hematopoiesis.

Hematopoiesis is a highly coordinated process that generates all the body's blood cells, and perturbations in embryonic hematopoiesis may result in illnesses ranging from fetal anemia to various leukemias. Correct establishment of hematopoietic progenitor cell fate is essential for the development of adequate blood cell subpopulations, although regulators of cell fate commitment have not been fully defined. Here, we show that primary erythropoiesis and myelopoiesis in zebrafish embryos are synergistically regulated by blf and the drl cluster, as simultaneous depletion led to severe erythrocyte aplasia and excessive macrophage formation at the expense of neutrophil development. Integrative analysis of transcriptome- and genome-wide binding data revealed that blf and drl cluster genes are responsible for constraining the expression of vasculogenesis-promoting genes in the intermediate cell mass and monocytopoiesis-promoting genes in the rostral blood island. This indicates that blf and drl cluster genes act as determinants of the fate commitment of erythroid and myeloid progenitor cells. Furthermore, a rescue screen demonstrated that Zfp932 is a potential mammalian functional equivalent to zebrafish blf and drl cluster genes. Our data provide insight into conserved cell fate commitment mechanisms of primitive hematopoiesis.

Foot-and-mouth disease virus (FMDV) negatively regulates ZFP36 protein expression to alleviate its antiviral activity.

Zinc finger protein 36 (ZFP36) is a key regulator of inflammatory and cytokine production. However, the interplay between swine zinc-finger protein 36 (sZFP36) and foot-and-mouth disease virus (FMDV) has not yet been reported. Here, we demonstrate that overexpression of sZFP36 restricted FMDV replication, while the knockdown of sZFP36 facilitated FMDV replication. To subvert the antagonism of sZFP36, FMDV decreased sZFP36 protein expression through its non-structural protein 3C protease (3Cpro). Our results also suggested that 3Cpro-mediated sZFP36 degradation was dependent on its protease activity. Further investigation revealed that both N-terminal and C-terminal-sZFP36 could be degraded by FMDV and FMDV 3Cpro. In addition, both N-terminal and C-terminal-sZFP36 decreased FMDV replication. Moreover, sZFP36 promotes the degradation of FMDV structural proteins VP3 and VP4 via the CCCH-type zinc finger and NES domains of sZFP36. Together, our results confirm that sZFP36 is a host restriction factor that negatively regulates FMDV replication.IMPORTANCEFoot-and-mouth disease (FMD) is an infectious disease of animals caused by the pathogen foot-and-mouth disease virus (FMDV). FMD is difficult to prevent and control because there is no cross-protection between its serotypes. Thus, we designed this study to investigate virus-host interactions. We first demonstrate that swine zinc-finger protein 36 (sZFP36) impaired FMDV structural proteins VP3 and VP4 to suppress viral replication. To subvert the antagonism of sZFP36, FMDV and FMDV 3Cpro downregulate sZFP36 expression to facilitate FMDV replication. Taken together, the present study reveals a previously unrecognized antiviral mechanism for ZFP36 and elucidates the role of FMDV in counteracting host antiviral activity.

The RNA-binding protein ZC3H11A interacts with the nuclear poly(A)-binding protein PABPN1 and alters polyadenylation of viral transcripts.

Nuclear mRNA metabolism is regulated by multiple proteins, which either directly bind to RNA or form multiprotein complexes. The RNA-binding protein ZC3H11A is involved in nuclear mRNA export, NF-κB signaling, and is essential during mouse embryo development. Furthermore, previous studies have shown that ZC3H11A is important for nuclear-replicating viruses. However, detailed biochemical characterization of the ZC3H11A protein has been lacking. In this study, we established the ZC3H11A protein interactome in human and mouse cells. We demonstrate that the nuclear poly(A)-binding protein PABPN1 interacts specifically with the ZC3H11A protein and controls ZC3H11A localization into nuclear speckles. We report that ZC3H11A specifically interacts with the human adenovirus type 5 (HAdV-5) capsid mRNA in a PABPN1-dependent manner. Notably, ZC3H11A uses the same zinc finger motifs to interact with PABPN1 and viral mRNA. Further, we demonstrate that the lack of ZC3H11A alters the polyadenylation of HAdV-5 capsid mRNA. Taken together, our results suggest that the ZC3H11A protein may act as a novel regulator of polyadenylation of nuclear mRNA.

Modulation of the wheat transcriptome by TaZFP13D under well-watered and drought conditions.

Maintaining global food security in the context of climate changes will be an important challenge in the next century. Improving abiotic stress tolerance of major crops such as wheat can contribute to this goal. This can be achieved by the identification of the genes involved and their use to develop tools for breeding programs aiming to generate better adapted cultivars. Recently, we identified the wheat TaZFP13D gene encoding Zinc Finger Protein 13D as a new gene improving water-stress tolerance. The current work analyzes the TaZFP13D-dependent transcriptome modifications that occur in well-watered and dehydration conditions to better understand its function during normal growth and during drought. Plants that overexpress TaZFP13D have a higher biomass under well-watered conditions, indicating a positive effect of the protein on growth. Survival rate and stress recovery after a severe drought stress are improved compared to wild-type plants. The latter is likely due the higher activity of key antioxidant enzymes and concomitant reduction of drought-induced oxidative damage. Conversely, down-regulation of TaZFP13D decreases drought tolerance and protection against drought-induced oxidative damage. RNA-Seq transcriptome analysis identified many genes regulated by TaZFP13D that are known to improve drought tolerance. The analysis also revealed several genes involved in the photosynthetic electron transfer chain known to improve photosynthetic efficiency and chloroplast protection against drought-induced ROS damage. This study highlights the important role of TaZFP13D in wheat drought tolerance, contributes to unravel the complex regulation governed by TaZFPs, and suggests that it could be a promising marker to select wheat cultivars with higher drought tolerance.

Expression of ZKSCAN3 protein suppresses proliferation, migration, and invasion of pancreatic cancer through autophagy.

Elevated autophagy activity enhances the malignancy of pancreatic cancer (PaCa), and autophagy is recognized as a novel therapeutic target. Zinc finger protein with KRAB and SCAN domains 3 (ZKSCAN3) is a transcription factor that suppresses autophagy, but its association with PaCa is unknown. We analyzed the function of ZKSCAN3 in PaCa and investigated whether autophagy regulation through ZKSCAN3 could become a new therapeutic target for PaCa. Using reverse transcription-quantitative polymerase chain reaction and western blotting, we observed that ZKSCAN3 expression was upregulated in several PaCa cell lines compared with normal pancreatic ductal epithelial cells. Additionally, comparing ZKSCAN3 expression with the prognosis of PaCa patients using web databases, we found that higher ZKSCAN3 expression in PaCa was associated with extended overall survival. Knocking down ZKSCAN3 promoted the proliferation of PaCa cells. Moreover, following ZKSCAN3 knockdown, PaCa cells exhibited significantly enhanced migratory and invasive properties. Conversely, overexpression of ZKSCAN3 significantly suppressed the proliferation, migration and invasion of PaCa cells. Additionally, the knockdown of ZKSCAN3 increased the expression of LC3-II, a marker of autophagy, whereas ZKSCAN3 overexpression decreased LC3-II expression. In a xenograft mouse model, tumors formed by MIA PaCa-2 cells in which ZKSCAN3 was knocked down significantly increased in size compared with the control group. In conclusion, ZKSCAN3 expression was upregulated in several pancreatic cancer cells. Additionally, it was revealed that ZKSCAN3 is negatively correlated with the malignancy of PaCa through autophagy. These results suggest that autophagy regulation via ZKSCAN3 may be a new therapeutic target for PaCa.

ZmC2H2-149 negatively regulates drought tolerance by repressing ZmHSD1 in maize.

Drought is a major abiotic stress that limits maize production worldwide. Therefore, it is of great importance to improve drought tolerance in crop plants for sustainable agriculture. In this study, we examined the roles of Cys2 /His2 zinc-finger-proteins (C2H2-ZFPs) in maize's drought tolerance as C2H2-ZFPs have been implicated for plant stress tolerance. By subjecting 150 Ac/Ds mutant lines to drought stress, we successfully identified a Ds-insertion mutant, zmc2h2-149, which shows increased tolerance to drought stress. Overexpression of ZmC2H2-149 in maize led to a decrease in both drought tolerance and crop yield. DAP-Seq, RNA-Seq, Y1H and LUC assays additionally showed that ZmC2H2-149 directly suppresses the expression of a positive drought tolerance regulator, ZmHSD1 (hydroxysteroid dehydrogenase 1). Consistently, the zmhsd1 mutants exhibited decreased drought tolerance and grain yield under water deficit conditions compared to their respective wild-type plants. Our findings thus demonstrated that ZmC2H2-149 can regulate ZmHSD1 for drought stress tolerance in maize, offering valuable theoretical and genetic resources for maize breeding programmes that aim for improving drought tolerance.

Plasmodiophora brassicae affects host gene expression by secreting the transcription factor-type effector PbZFE1.

The protist pathogen Plasmodiophora brassicae hijacks the metabolism and development of host cruciferous plants and induces clubroot formation, but little is known about its regulatory mechanisms. Previously, the Pnit2int2 sequence, a sequence around the second intron of the nitrilase gene (BrNIT2) involved in auxin biosynthesis in Brassica rapa ssp. pekinensis, was identified as a specific promoter activated during clubroot formation. In this study, we hypothesized that analysis of the transcriptional regulation of Pnit2int2 could reveal how P. brassicae affects the host gene regulatory system during clubroot development. By yeast one-hybrid screening, the pathogen zinc finger protein PbZFE1 was identified to specifically bind to Pnit2int2. Specific binding of PbZFE1 to Pnit2int2 was also confirmed by electrophoretic mobility shift assay. The binding site of PbZFE1 is essential for promoter activity of Pnit2int2 in clubbed roots of transgenic Arabidopsis thaliana (Pnit2int2-2::GUS), indicating that PbZFE1 is secreted from P. brassicae and functions within plant cells. Ectopic expression of PbZEF1 in A. thaliana delayed growth and flowering time, suggesting that PbZFE1 has significant impacts on host development and metabolic systems. Thus, P. brassicae appears to secrete PbZFE1 into host cells as a transcription factor-type effector during pathogenesis.

Lemon zinc finger protein ClSUP induces the accumulation of reactive oxygen species and inhibits citrus yellow vein-clearing virus infection via interactions with ClDOF3.4.

Citrus yellow vein-clearing virus (CYVCV) is an increasing threat to citrus cultivation. Notably, the role of zinc finger proteins (ZFPs) in mediating viral resistance in citrus plants is unclear. In this study, we demonstrate that ZFPs ClSUP and ClDOF3.4 enhance citrus defense responses against CYVCV in Eureka lemon. ClSUP interacted with the coat protein (CP) of CYVCV to reduce CP accumulation and inhibit its silencing suppressor function. Overexpression of CISUP triggered reactive oxygen species (ROS) and salicylic acid (SA) pathways, and enhanced resistance to CYVCV infection. In contrast, ClSUP-silencing resulted in increased CP accumulation and down-regulated ROS and SA-related genes. ClDOF3.4 interacts with ClSUP to facilitate its interactions with CP. Furthermore, ClDOF3.4 synergistically regulated the accumulation of ROS and SA with ClSUP and accelerated the down-regulation of CP accumulation. Transgenic plants co-expressing ClSUP and ClDOF3.4 remarkedly decrease the CYVCV. These findings provide a new reference for understanding the interaction mechanism between the host and CYVCV.

Tenascin C as a novel zinc finger protein 750 target regulating the immunogenicity via DNA damage in lung squamous cell carcinoma.

Modulation of DNA damage repair in lung squamous cell carcinoma (LUSC) can result in the generation of neoantigens and heightened immunogenicity. Therefore, understanding DNA damage repair mechanisms holds significant clinical relevance for identifying targets for immunotherapy and devising therapeutic strategies. Our research has unveiled that the tumor suppressor zinc finger protein 750 (ZNF750) in LUSC binds to the promoter region of tenascin C (TNC), leading to reduced TNC expression. This modulation may impact the malignant behavior of tumor cells and is associated with patient prognosis. Additionally, single-cell RNA sequencing (scRNA-seq) of LUSC tissues has demonstrated an inverse correlation between ZNF750/TNC expression levels and immunogenicity. Manipulation of the ZNF750-TNC axis in vitro within LUSC cells has shown differential sensitivity to CD8+ cells, underscoring its pivotal role in regulating cellular immunogenicity. Further transcriptome sequencing analysis, DNA damage repair assay, and single-strand break analyses have revealed the involvement of the ZNF750-TNC axis in determining the preference for homologous recombination (HR) repair or non-homologous end joining (NHEJ) repair of DNA damage. with involvement of the Hippo/ERK signaling pathway. In summary, this study sheds light on the ZNF750-TNC axis's role in DNA damage repair regulation in LUSC, laying a groundwork for future translational research in immune cell therapy for LUSC.

ZFP207 sustains pluripotency by coordinating OCT4 stability, alternative splicing and RNA export.

The pluripotent state is not solely governed by the action of the core transcription factors OCT4, SOX2, and NANOG, but also by a series of co-transcriptional and post-transcriptional events, including alternative splicing (AS) and the interaction of RNA-binding proteins (RBPs) with defined subpopulations of RNAs. Zinc Finger Protein 207 (ZFP207) is an essential transcription factor for mammalian embryonic development. Here, we employ multiple functional analyses to characterize its role in mouse embryonic stem cells (ESCs). We find that ZFP207 plays a pivotal role in ESC maintenance, and silencing of Zfp207 leads to severe neuroectodermal differentiation defects. In striking contrast to human ESCs, mouse ZFP207 does not transcriptionally regulate neuronal and stem cell-related genes but exerts its effects by controlling AS networks and by acting as an RBP. Our study expands the role of ZFP207 in maintaining ESC identity, and underscores the functional versatility of ZFP207 in regulating neural fate commitment.

ZNF283, a Krüppel-associated box zinc finger protein, inhibits RNA synthesis of porcine reproductive and respiratory syndrome virus by interacting with Nsp9 and Nsp10.

Porcine reproductive and respiratory syndrome virus (PRRSV) is a viral pathogen with substantial economic implications for the global swine industry. The existing vaccination strategies and antiviral drugs offer limited protection. Replication of the viral RNA genome encompasses a complex series of steps, wherein a replication complex is assembled from various components derived from both viral and cellular sources, as well as from the viral genomic RNA template. In this study, we found that ZNF283, a Krüppel-associated box (KRAB) containing zinc finger protein, was upregulated in PRRSV-infected Marc-145 cells and porcine alveolar macrophages and that ZNF283 inhibited PRRSV replication and RNA synthesis. We also found that ZNF283 interacts with the viral proteins Nsp9, an RNA-dependent RNA polymerase, and Nsp10, a helicase. The main regions involved in the interaction between ZNF283 and Nsp9 were determined to be the KRAB domain of ZNF283 and amino acids 178-449 of Nsp9. The KRAB domain of ZNF283 plays a role in facilitating Nsp10 binding. In addition, ZNF283 may have an affinity for the 3' untranslated region of PRRSV. These findings suggest that ZNF283 is an antiviral factor that inhibits PRRSV infection and extend our understanding of the interactions between KRAB-containing zinc finger proteins and viruses.

SETDB1 deletion causes DNA demethylation and upregulation of multiple zinc-finger genes.

BACKGROUND: SETDB1 (SET domain bifurcated-1) is a histone H3-lysine 9 (H3K9)-specific methyltransferase that mediates heterochromatin formation and repression of target genes. Despite the assumed functional link between DNA methylation and SETDB1-mediated H3K9 trimethylations, several studies have shown that SETDB1 operates autonomously of DNA methylation in a region- and cell-specific manner. This study analyzes SETDB1-null HAP1 cells through a linked methylome and transcriptome analysis, intending to explore genes controlled by SETDB1-involved DNA methylation. METHODS AND RESULTS: We investigated SETDB1-mediated regulation of DNA methylation and gene transcription in human HAP1 cells using reduced-representation bisulfite sequencing (RRBS) and RNA sequencing. While two-thirds of differentially methylated CpGs (DMCs) in genic regions were hypomethylated in SETDB1-null cells, we detected a plethora of C2H2-type zinc-finger protein genes (C2H2-ZFP, 223 of 749) among the DMC-associated genes. Most C2H2-ZFPs with DMCs in their promoters were found hypomethylated in SETDB1-KO cells, while other non-ZFP genes with promoter DMCs were not. These C2H2-ZFPs with DMCs in their promoters were significantly upregulated in SETDB1-KO cells. Similarly, C2H2-ZFP genes were upregulated in SETDB1-null 293T cells, suggesting that SETDB1's function in ZFP gene repression is widespread. There are several C2H2-ZFP gene clusters on chromosome 19, which were selectively hypomethylated in SETDB1-KO cells. CONCLUSIONS: SETDB1 collectively and specifically represses a substantial fraction of the C2H2-ZFP gene family. Through the en-bloc silencing of a set of ZFP genes, SETDB1 may help establish a panel of ZFP proteins that are expressed cell-type specifically and thereby can serve as signature proteins for cellular identity.

tRF3-IleAAT reduced extracellular matrix synthesis in diabetic kidney disease mice by targeting ZNF281 and inhibiting ferroptosis.

It is well established that the synthesis of extracellular matrix (ECM) in mesangial cells is a major determinant of diabetic kidney disease (DKD). Elucidating the major players in ECM synthesis may be helpful to provide promising candidates for protecting against DKD progression. tRF3-IleAAT is a tRNA-derived fragment (tRF) produced by nucleases at tRNA-specific sites, which is differentially expressed in the sera of patients with diabetes mellitus and DKD. In this study we investigated the potential roles of tRFs in DKD. Db/db mice at 12 weeks were adapted as a DKD model. The mice displayed marked renal dysfunction accompanied by significantly reduced expression of tRF3-IleAAT and increased ferroptosis and ECM synthesis in the kidney tissues. The reduced expression of tRF3-IleAAT was also observed in high glucose-treated mouse glomerular mesangial cells. We administered ferrostatin-1 (1 mg/kg, once every two days, i.p.) to the mice from the age of 12 weeks for 8 weeks, and found that inhibition of the onset of ferroptosis significantly improved renal function, attenuated renal fibrosis and reduced collagen deposition. Overexpression of tRF3-IleAAT by a single injection of AAV carrying tRF3-IleAAT via caudal vein significantly inhibited ferroptosis and ECM synthesis in DKD model mice. Furthermore, we found that the expression of zinc finger protein 281 (ZNF281), a downstream target gene of tRF3-IleAAT, was significantly elevated in DKD models but negatively regulated by tRF3-IleAAT. In high glucose-treated mesangial cells, knockdown of ZNF281 exerted an inhibitory effect on ferroptosis and ECM synthesis. We demonstrated the targeted binding of tRF3-IleAAT to the 3'UTR of ZNF281. In conclusion, tRF3-IleAAT inhibits ferroptosis by targeting ZNF281, resulting in the mitigation of ECM synthesis in DKD models, suggesting that tRF3-IleAAT may be an attractive therapeutic target for DKD.

circZNF532 promotes endothelial-to-mesenchymal transition in diabetic retinopathy by recruiting TAF15 to stabilize PIK3CD.

Endothelial-to-mesenchymal transition (EndMT) is a pivotal event in diabetic retinopathy (DR). This study explored the role of circRNA zinc finger protein 532 (circZNF532) in regulating EndMT in DR progression. Human retinal microvascular endothelial cells (HRMECs) were exposed to high glucose (HG) to induce the DR cell model. Actinomycin D-treated HRMECs were used to confirm the mRNA stability of phosphoinositide-3 kinase catalytic subunit δ (PIK3CD). The interaction between TATA-box-binding protein-associated factor 15 (TAF15) and circZNF532/PIK3CD was subsequently analyzed using RNA immunoprecipitation (RIP), RNA pull-down. It was found that HG treatment accelerated EndMT process, facilitated cell migration and angiogenesis, and enhanced PIK3CD and p-AKT levels in HRMECs, whereas si-circZNF532 transfection neutralized these effects. Further data showed that circZNF532 recruited TAF15 to stabilize PIK3CD, thus elevating PIK3CD expression. Following rescue experiments suggested that PIK3CD overexpression partially negated the inhibitory effect of circZNF532 silencing on EndMT, migration, and angiogenesis of HG-treated HRMECs. In conclusion, our results suggest that circZNF532 recruits TAF15 to stabilize PIK3CD, thereby facilitating EndMT in DR.