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1.
Sci Rep ; 14(1): 10662, 2024 05 09.
Artigo em Inglês | MEDLINE | ID: mdl-38724606

RESUMO

DBF4 zinc finger (DBF4) is a critical component involved in DNA replication and cell proliferation. It acts as a positive regulator of the cell division cycle 7 kinase. In this study, our investigation encompassed the impact of DBF4 on hepatocellular carcinoma (HCC) progression and delved into the potential mechanisms. We utilized open-access databases like TCGA and GEO to analyze the association between DBF4 and 33 different tumor types. We also conducted immunohistochemistry experiments to validate the expression of DBF4 in HCC, STAD, COAD, READ, PAAD, and LGG. Furthermore, we employed lentiviral transduction to knockdown DBF4 in HLF and SMMC cells, as well as to overexpress DBF4 in Huh7 cells. Subsequently, we evaluated the impact of DBF4 on proliferation, migration, and invasion of hepatocellular carcinoma cells. RNA sequencing and KEGG pathway enrichment analysis were also conducted to identify potential pathways, which were further validated through WB experiments. Finally, pathway inhibitor was utilized in rescue experiments to confirm whether DBF4 exerts its effects on tumor cells via the implicated pathway. Our findings revealed that DBF4 exhibited significant expression levels in nearly all examined tumors, which were further substantiated by the results of immunohistochemistry analysis. High DBF4 expression was correlated with poor overall survival (OS), disease-specific survival (DSS), progression-free interval (PFI), disease-free interval (DFI), relapse-free interval (RFI) in majority of tumor types, particularly in patients with HCC. In vitro experiments demonstrated that inhibition of DBF4 impaired the proliferative, migratory, and invasive abilities of HCC cells, whereas overexpression of DBF4 promoted these phenotypes. Sequencing results indicated that DBF4 may induce these changes through the ERBB signaling pathway. Further experimental validation revealed that DBF4 activates the ERBB signaling pathway, leading to alterations in the JNK/STAT, MAPK, and PI3K/AKT signaling pathways, thereby impacting the proliferative, migratory, and invasive abilities of tumor cells. Lastly, treatment of Huh7 cells overexpressing DBF4 with the ERBB2 inhibitor dacomitinib demonstrated the ability of ERBB2 inhibition to reverse the promoting effect of DBF4 overexpression on the proliferative, migratory, and invasive abilities of HCC cells. DBF4 plays a pivotal oncogenic role in HCC by promoting the ERBB signaling pathway and activating its downstream PI3K/AKT, JNK/STAT3, and MAPK signaling pathways. DBF4 may serve as a prognostic biomarker for patients with HCC.


Assuntos
Carcinoma Hepatocelular , Proteínas de Ciclo Celular , Neoplasias Hepáticas , Feminino , Humanos , Masculino , Biomarcadores Tumorais/metabolismo , Biomarcadores Tumorais/genética , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/patologia , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Regulação Neoplásica da Expressão Gênica , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/patologia , Prognóstico , Transdução de Sinais , Dedos de Zinco , Proteínas de Ciclo Celular/genética
2.
Int J Mol Sci ; 25(8)2024 Apr 17.
Artigo em Inglês | MEDLINE | ID: mdl-38673993

RESUMO

Cucumber (Cucumis sativus L.) is a globally prevalent and extensively cultivated vegetable whose yield is significantly influenced by various abiotic stresses, including drought, heat, and salinity. Transcription factors, such as zinc finger-homeodomain proteins (ZHDs), a plant-specific subgroup of Homeobox, play a crucial regulatory role in stress resistance. In this study, we identified 13 CsZHDs distributed across all six cucumber chromosomes except chromosome 7. Phylogenetic analysis classified these genes into five clades (ZHDI-IV and MIF) with different gene structures but similar conserved motifs. Collinearity analysis revealed that members of clades ZHD III, IV, and MIF experienced amplification through segmental duplication events. Additionally, a closer evolutionary relationship was observed between the ZHDs in Cucumis sativus (C. sativus) and Arabidopsis thaliana (A. thaliana) compared to Oryza sativa (O. sativa). Quantitative real-time PCR (qRT-PCR) analysis demonstrated the general expression of CsZHD genes across all tissues, with notable expression in leaf and flower buds. Moreover, most of the CsZHDs, particularly CsZHD9-11, exhibited varying responses to drought, heat, and salt stresses. Virus-induced gene silencing (VIGS) experiments highlighted the potential functions of CsZHD9 and CsZHD10, suggesting their positive regulation of stomatal movement and responsiveness to drought stress. In summary, these findings provide a valuable resource for future analysis of potential mechanisms underlying CsZHD genes in response to stresses.


Assuntos
Cucumis sativus , Evolução Molecular , Regulação da Expressão Gênica de Plantas , Filogenia , Proteínas de Plantas , Estresse Fisiológico , Cucumis sativus/genética , Cucumis sativus/metabolismo , Estresse Fisiológico/genética , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Dedos de Zinco/genética , Secas , Cromossomos de Plantas/genética , Perfilação da Expressão Gênica
3.
BMC Genomics ; 25(1): 385, 2024 Apr 19.
Artigo em Inglês | MEDLINE | ID: mdl-38641598

RESUMO

BACKGROUND: The C2H2 zinc finger protein family plays important roles in plants. However, precisely how C2H2s function in Opisthopappus (Opisthopappus taihangensis and Opisthopappus longilobus) remains unclear. RESULTS: In this study, a total of 69 OpC2H2 zinc finger protein genes were identified and clustered into five Groups. Seven tandem and ten fragment repeats were found in OpC2H2s, which underwent robust purifying selection. Of the identified motifs, motif 1 was present in all OpC2H2s and conserved at important binding sites. Most OpC2H2s possessed few introns and exons that could rapidly activate and react when faced with stress. The OpC2H2 promoter sequences mainly contained diverse regulatory elements, such as ARE, ABRE, and LTR. Under salt stress, two up-regulated OpC2H2s (OpC2H2-1 and OpC2H2-14) genes and one down-regulated OpC2H2 gene (OpC2H2-7) might serve as key transcription factors through the ABA and JA signaling pathways to regulate the growth and development of Opisthopappus species. CONCLUSION: The above results not only help to understand the function of C2H2 gene family but also drive progress in genetic improvement for the salt tolerance of Opisthopappus species.


Assuntos
Dedos de Zinco CYS2-HIS2 , Dedos de Zinco CYS2-HIS2/genética , Estresse Salino/genética , Genoma de Planta , Fatores de Transcrição/metabolismo , Dedos de Zinco/genética , Proteínas de Plantas/metabolismo , Regulação da Expressão Gênica de Plantas , Filogenia
4.
Comput Biol Med ; 174: 108397, 2024 May.
Artigo em Inglês | MEDLINE | ID: mdl-38603896

RESUMO

The equilibrium of cellular protein levels is pivotal for maintaining normal physiological functions. USP5 belongs to the deubiquitination enzyme (DUBs) family, controlling protein degradation and preserving cellular protein homeostasis. Aberrant expression of USP5 is implicated in a variety of diseases, including cancer, neurodegenerative diseases, and inflammatory diseases. In this paper, a multi-level virtual screening (VS) approach was employed to target the zinc finger ubiquitin-binding domain (ZnF-UBD) of USP5, leading to the identification of a highly promising candidate compound 0456-0049. Molecular dynamics (MD) simulations were then employed to assess the stability of complex binding and predict hotspot residues in interactions. The results indicated that the candidate stably binds to the ZnF-UBD of USP5 through crucial interactions with residues ARG221, TRP209, GLY220, ASN207, TYR261, TYR259, and MET266. Binding free energy calculations, along with umbrella sampling (US) simulations, underscored a superior binding affinity of the candidate relative to known inhibitors. Moreover, US simulations revealed conformational changes of USP5 during ligand dissociation. These insights provide a valuable foundation for the development of novel inhibitors targeting USP5.


Assuntos
Simulação de Dinâmica Molecular , Dedos de Zinco , Humanos , Endopeptidases/química , Endopeptidases/metabolismo , Ligação Proteica , Domínios Proteicos
5.
Proc Natl Acad Sci U S A ; 121(17): e2320938121, 2024 Apr 23.
Artigo em Inglês | MEDLINE | ID: mdl-38635637

RESUMO

The MYC-Associated Zinc Finger Protein (MAZ) plays important roles in chromatin organization and gene transcription regulation. Dysregulated expression of MAZ causes diseases, such as glioblastoma, breast cancer, prostate cancer, and liposarcoma. Previously, it has been reported that MAZ controls the proinflammatory response in colitis and colon cancer via STAT3 signaling, suggesting that MAZ is involved in regulating immunity-related pathways. However, the molecular mechanism underlying this regulation remains elusive. Here, we investigate the regulatory effect of MAZ on interferon-gamma (IFN-γ)-stimulated genes via STAT1, a protein that plays an essential role in immune responses to viral, fungal, and mycobacterial pathogens. We demonstrate that about 80% of occupied STAT1-binding sites colocalize with occupied MAZ-binding sites in HAP1/K562 cells after IFN-γ stimulation. MAZ depletion significantly reduces STAT1 binding in the genome. By analyzing genome-wide gene expression profiles in the RNA-Seq data, we show that MAZ depletion significantly suppresses a subset of the immune response genes, which include the IFN-stimulated genes IRF8 and Absent in Melanoma 2. Furthermore, we find that MAZ controls expression of the immunity-related genes by changing the epigenetic landscape in chromatin. Our study reveals an important role for MAZ in regulating immune-related gene expression.


Assuntos
Cromatina , Interferon gama , Masculino , Humanos , Interferon gama/genética , Interferon gama/farmacologia , Cromatina/genética , Regulação da Expressão Gênica , Ligação Proteica , Dedos de Zinco/genética , Fator de Transcrição STAT1/genética
6.
Biochem Biophys Res Commun ; 710: 149862, 2024 May 28.
Artigo em Inglês | MEDLINE | ID: mdl-38593618

RESUMO

Zinc is an important trace element in the human body, and its homeostasis is closely related to amyotrophic lateral sclerosis (ALS). Cytoplasmic FUS proteins from patients with ALS aggregate their important pathologic markers. Liquid-liquid phase separation (LLPS) of FUS can lead to its aggregation. However, whether and how zinc homeostasis affects the aggregation of disease-associated FUS proteins in the cytoplasm remains unclear. Here, we found that zinc ion enhances LLPS and promotes the aggregation in the cytoplasm for FUS protein. In the FUS, the cysteine of the zinc finger (ZnF), recognizes and binds to zinc ions, reducing droplet mobility and enhancing protein aggregation in the cytoplasm. The mutation of FUS cysteine disrupts the dynamic regulatory switch of zinc ions and ZnF, resulting in insensitivity to zinc ions. These results suggest that the dynamic regulation of LLPS by binding with zinc ions may be a widespread mechanism and provide a new understanding of neurological diseases such as ALS and other ZnF protein-related diseases.


Assuntos
Esclerose Lateral Amiotrófica , Proteína FUS de Ligação a RNA , Humanos , Esclerose Lateral Amiotrófica/genética , Esclerose Lateral Amiotrófica/metabolismo , Cisteína/genética , Mutação , Separação de Fases , Proteína FUS de Ligação a RNA/química , Proteína FUS de Ligação a RNA/genética , Proteína FUS de Ligação a RNA/metabolismo , Zinco/metabolismo , Dedos de Zinco , Agregados Proteicos
7.
Biochim Biophys Acta Mol Cell Res ; 1871(4): 119707, 2024 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-38493895

RESUMO

Ribosomes across species contain subsets of zinc finger proteins that play structural roles by binding to rRNA. While the majority of these zinc fingers belong to the C2-C2 type, the large subunit protein L36 in bacteria and mitochondria exhibits an atypical C2-CH motif. To comprehend the contribution of each coordinating residue in S. cerevisiae bL36m to mitoribosome assembly and function, we engineered and characterized strains carrying single and double mutations in the zinc coordinating residues. Our findings reveal that although all four residues markedly influence protein stability, C to A mutations in C66 and/or C69 have a more pronounced effect compared to those at C82 and H88. Importantly, protein stability directly correlates with the assembly and function of the mitoribosome and the growth rate of yeast in respiratory conditions. Mass spectrometry analysis of large subunit particles indicates that strains deleted for bL36m or expressing mutant variants have defective assembly of the L7/L12 stalk base, limiting their functional competence. Furthermore, we employed a synthetic bL36m protein collection, including both wild-type and mutant proteins, to elucidate their ability to bind zinc. Our data indicate that mutations in C82 and, particularly, H88 allow for some zinc binding albeit inefficient or unstable, explaining the residual accumulation and activity in mitochondria of bL36m variants carrying mutations in these residues. In conclusion, stable zinc binding by bL36m is essential for optimal mitoribosome assembly and function. MS data are available via ProteomeXchange with identifierPXD046465.


Assuntos
Ribossomos Mitocondriais , Saccharomyces cerevisiae , Ribossomos Mitocondriais/química , Ribossomos Mitocondriais/metabolismo , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Dedos de Zinco/genética , Subunidades Ribossômicas Maiores/genética , Zinco/metabolismo
8.
Science ; 383(6689): 1284-1289, 2024 Mar 22.
Artigo em Inglês | MEDLINE | ID: mdl-38513035

RESUMO

Can the course of fatal prion diseases be changed by removing the protein before it goes bad?


Assuntos
Doenças Priônicas , Príons , Humanos , Doenças Priônicas/genética , Doenças Priônicas/prevenção & controle , Príons/antagonistas & inibidores , Príons/genética , Mutação , Dedos de Zinco
9.
Science ; 383(6688): eadk4422, 2024 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-38484051

RESUMO

Conditional protein degradation tags (degrons) are usually >100 amino acids long or are triggered by small molecules with substantial off-target effects, thwarting their use as specific modulators of endogenous protein levels. We developed a phage-assisted continuous evolution platform for molecular glue complexes (MG-PACE) and evolved a 36-amino acid zinc finger (ZF) degron (SD40) that binds the ubiquitin ligase substrate receptor cereblon in complex with PT-179, an orthogonal thalidomide derivative. Endogenous proteins tagged in-frame with SD40 using prime editing are degraded by otherwise inert PT-179. Cryo-electron microscopy structures of SD40 in complex with ligand-bound cereblon revealed mechanistic insights into the molecular basis of SD40's activity and specificity. Our efforts establish a system for continuous evolution of molecular glue complexes and provide ZF tags that overcome shortcomings associated with existing degrons.


Assuntos
Degrons , Evolução Molecular Direcionada , Proteólise , Ubiquitina-Proteína Ligases , Dedos de Zinco , Microscopia Crioeletrônica , Talidomida/química , Ubiquitina-Proteína Ligases/química , Ubiquitinação , Degrons/genética , Dedos de Zinco/genética , Quimera de Direcionamento de Proteólise , Evolução Molecular Direcionada/métodos , Humanos
10.
Microbiol Res ; 283: 127695, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38554651

RESUMO

Cap expansion in agaricoid mushroom species is an important event for sexual reproduction because meiosis occurs in basidia under the cap, and basidiospores can be released by opening the cap. However, molecular mechanisms underlying cap expansion in basidiomycetes remain poorly understood. We aimed to elucidate the molecular mechanisms of cap expansion in basidiomycetes by analyzing the unique cap-expansionless UV mutant #13 (exp2-1) in Coprinopsis cinerea. Linkage analysis and consequent genome sequence analysis revealed that the gene responsible for the mutant phenotypes encodes a putative transcription factor with two C2H2 zinc finger motifs. The mutant that was genome-edited to lack exp2 exhibited an expansionless phenotype. Some of the genes encoding cell wall degradation-related enzymes showed decreased expression during cap expansion and autolysis in the exp2 UV and genome-edited mutant. The exp2 gene is widely conserved in Agaricomycetes, suggesting that Exp2 homologs regulate fruiting body maturation in Agaricomycetes, especially cap expansion in Agaricoid-type mushroom-forming fungi. Therefore, exp2 homologs could be a target for mushroom breeding to maintain shape after harvest for some cultivating mushrooms, presenting a promising avenue for further research in breeding techniques.


Assuntos
Agaricales , Basidiomycota , Carpóforos/genética , Agaricales/genética , Dedos de Zinco/genética , Proteínas Fúngicas/genética , Proteínas Fúngicas/metabolismo
11.
J Integr Plant Biol ; 66(4): 787-809, 2024 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-38477645

RESUMO

The recretohalophyte Limonium bicolor thrives in high-salinity environments because salt glands on the above-ground parts of the plant help to expel excess salt. Here, we characterize a nucleus-localized C3HC4 (RING-HC)-type zinc finger protein of L. bicolor named  RING  ZINC  FINGER PROTEIN  1 (LbRZF1). LbRZF1 was expressed in salt glands and in response to NaCl treatment. LbRZF1 showed no E3 ubiquitin ligase activity. The phenotypes of overexpression and knockout lines for LbRZF1 indicated that LbRZF1 positively regulated salt gland development and salt tolerance in L. bicolor. lbrzf1 mutants had fewer salt glands and secreted less salt than did the wild-type, whereas LbRZF1-overexpressing lines had opposite phenotypes, in keeping with the overall salt tolerance of these plants. A yeast two-hybrid screen revealed that LbRZF1 interacted with LbCATALASE2 (LbCAT2) and the transcription factor LbMYB113, leading to their stabilization. Silencing of LbCAT2 or LbMYB113 decreased salt gland density and salt tolerance. The heterologous expression of LbRZF1 in Arabidopsis thaliana conferred salt tolerance to this non-halophyte. We also identified the transcription factor LbMYB48 as an upstream regulator of LbRZF1 transcription. The study of LbRZF1 in the regulation network of salt gland development also provides a good foundation for transforming crops and improving their salt resistance.


Assuntos
Arabidopsis , Plumbaginaceae , Animais , Tolerância ao Sal/genética , Plumbaginaceae/genética , Plumbaginaceae/metabolismo , Glândula de Sal/metabolismo , Zinco/metabolismo , Arabidopsis/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Dedos de Zinco , Regulação da Expressão Gênica de Plantas , Plantas Geneticamente Modificadas/metabolismo , Estresse Fisiológico/genética , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo
12.
Open Biol ; 14(3): 230270, 2024 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-38471568

RESUMO

The Drosophila male-specific lethal (MSL) complex binds to the male X chromosome to activate transcription. It comprises five proteins (MSL1, MSL2, MSL3, male absent on the first (MOF), and maleless (MLE)) and two long noncoding RNAs (lncRNAs; roX1 and roX2). The MLE helicase remodels the roX lncRNAs, enabling the lncRNA-mediated assembly of the Drosophila dosage compensation complex. MSL2 is expressed only in males and interacts with the N-terminal zinc finger of the transcription factor chromatin-linked adapter for MSL proteins (CLAMP), which is important for the specific recruitment of the MSL complex to the male X chromosome. Here, we found that MLE's unstructured C-terminal region interacts with the sixth zinc-finger domain of CLAMP. In vitro, 4-5 zinc fingers are critical for the specific DNA-binding of CLAMP with GA repeats, which constitute the core motif at the high affinity binding sites for MSL proteins. Deleting the CLAMP binding region in MLE decreases the association of MSL proteins with the male X chromosome and increases male lethality. These results suggest that interactions of unstructured regions in MSL2 and MLE with CLAMP zinc finger domains are important for the specific recruitment of the MSL complex to the male X chromosome.


Assuntos
Proteínas de Drosophila , RNA Longo não Codificante , Animais , Masculino , Drosophila/metabolismo , Proteínas de Ligação a DNA/metabolismo , Drosophila melanogaster/genética , Proteínas de Drosophila/metabolismo , RNA Longo não Codificante/metabolismo , Proteínas Nucleares/metabolismo , Mecanismo Genético de Compensação de Dose , Dedos de Zinco , Cromossomo X/metabolismo
13.
Elife ; 122024 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-38488661

RESUMO

R-loops are non-canonical DNA structures that form during transcription and play diverse roles in various physiological processes. Disruption of R-loop homeostasis can lead to genomic instability and replication impairment, contributing to several human diseases, including cancer. Although the molecular mechanisms that protect cells against such events are not fully understood, recent research has identified fork protection factors and DNA damage response proteins as regulators of R-loop dynamics. In this study, we identify the Werner helicase-interacting protein 1 (WRNIP1) as a novel factor that counteracts transcription-associated DNA damage upon replication perturbation. Loss of WRNIP1 leads to R-loop accumulation, resulting in collisions between the replisome and transcription machinery. We observe co-localization of WRNIP1 with transcription/replication complexes and R-loops after replication perturbation, suggesting its involvement in resolving transcription-replication conflicts. Moreover, WRNIP1-deficient cells show impaired replication restart from transcription-induced fork stalling. Notably, transcription inhibition and RNase H1 overexpression rescue all the defects caused by loss of WRNIP1. Importantly, our findings highlight the critical role of WRNIP1 ubiquitin-binding zinc finger (UBZ) domain in preventing pathological persistence of R-loops and limiting DNA damage, thereby safeguarding genome integrity.


Assuntos
ATPases Associadas a Diversas Atividades Celulares , Replicação do DNA , Proteínas de Ligação a DNA , Humanos , ATPases Associadas a Diversas Atividades Celulares/metabolismo , DNA , Dano ao DNA , Proteínas de Ligação a DNA/metabolismo , Instabilidade Genômica , Hidrolases/genética , Dedos de Zinco
14.
Int J Mol Sci ; 25(6)2024 Mar 19.
Artigo em Inglês | MEDLINE | ID: mdl-38542422

RESUMO

Using an established human primary cell culture model, we previously demonstrated that the promyelocytic leukemia zinc finger (PLZF) transcription factor is a direct target of the progesterone receptor (PGR) and is essential for progestin-dependent decidualization of human endometrial stromal cells (HESCs). These in vitro findings were supported by immunohistochemical analysis of human endometrial tissue biopsies, which showed that the strongest immunoreactivity for endometrial PLZF is detected during the progesterone (P4)-dominant secretory phase of the menstrual cycle. While these human studies provided critical clinical support for the important role of PLZF in P4-dependent HESC decidualization, functional validation in vivo was not possible due to the absence of suitable animal models. To address this deficiency, we recently generated a conditional knockout mouse model in which PLZF is ablated in PGR-positive cells of the mouse (Plzf d/d). The Plzf d/d female was phenotypically analyzed using immunoblotting, real-time PCR, and immunohistochemistry. Reproductive function was tested using the timed natural pregnancy model as well as the artificial decidual response assay. Even though ovarian activity is not affected, female Plzf d/d mice exhibit an infertility phenotype due to an inability of the embryo to implant into the Plzf d/d endometrium. Initial cellular and molecular phenotyping investigations reveal that the Plzf d/d endometrium is unable to develop a transient receptive state, which is reflected at the molecular level by a blunted response to P4 exposure with a concomitant unopposed response to 17-ß estradiol. In addition to a defect in P4-dependent receptivity, the Plzf d/d endometrium fails to undergo decidualization in response to an artificial decidual stimulus, providing the in vivo validation for our earlier HESC culture findings. Collectively, our new Plzf d/d mouse model underscores the physiological importance of the PLZF transcription factor not only in endometrial stromal cell decidualization but also uterine receptivity, two uterine cellular processes that are indispensable for the establishment of pregnancy.


Assuntos
Leucemia , Fatores de Transcrição , Gravidez , Feminino , Camundongos , Animais , Humanos , Fatores de Transcrição/metabolismo , Decídua/metabolismo , Endométrio/metabolismo , Camundongos Knockout , Dedos de Zinco , Leucemia/metabolismo , Células Estromais/metabolismo
15.
Nature ; 627(8003): 416-423, 2024 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-38418872

RESUMO

Permanent epigenetic silencing using programmable editors equipped with transcriptional repressors holds great promise for the treatment of human diseases1-3. However, to unlock its full therapeutic potential, an experimental confirmation of durable epigenetic silencing after the delivery of transient delivery of editors in vivo is needed. To this end, here we targeted Pcsk9, a gene expressed in hepatocytes that is involved in cholesterol homeostasis. In vitro screening of different editor designs indicated that zinc-finger proteins were the best-performing DNA-binding platform for efficient silencing of mouse Pcsk9. A single administration of lipid nanoparticles loaded with the editors' mRNAs almost halved the circulating levels of PCSK9 for nearly one year in mice. Notably, Pcsk9 silencing and accompanying epigenetic repressive marks also persisted after forced liver regeneration, further corroborating the heritability of the newly installed epigenetic state. Improvements in construct design resulted in the development of an all-in-one configuration that we term evolved engineered transcriptional repressor (EvoETR). This design, which is characterized by a high specificity profile, further reduced the circulating levels of PCSK9 in mice with an efficiency comparable with that obtained through conventional gene editing, but without causing DNA breaks. Our study lays the foundation for the development of in vivo therapeutics that are based on epigenetic silencing.


Assuntos
Epigênese Genética , Epigenoma , Edição de Genes , Inativação Gênica , Animais , Camundongos , Colesterol/metabolismo , Epigênese Genética/genética , Epigenoma/genética , Edição de Genes/métodos , Hepatócitos/metabolismo , Fígado/metabolismo , Regeneração Hepática , Nanopartículas , Pró-Proteína Convertase 9/sangue , Pró-Proteína Convertase 9/deficiência , Pró-Proteína Convertase 9/genética , Proteínas Repressoras/administração & dosagem , Proteínas Repressoras/metabolismo , Dedos de Zinco
16.
Int Immunopharmacol ; 130: 111670, 2024 Mar 30.
Artigo em Inglês | MEDLINE | ID: mdl-38373386

RESUMO

Type 2 immune responses are critical for host defense, mediate allergy and Th2-high asthma. The transcription factor, promyelocytic leukemia zinc finger (PLZF), has emerged as a significant regulator of type 2 inflammation in the lung; however, its exact mechanism remains unclear. In this review, we summarized recent findings regarding the ability of PLZF to control the development and function of innate lymphoid cells (ILCs), iNKT cells, memory T cells, basophils, and other immune cells that drive type 2 responses. We discussed the important role of PLZF in the pathogenesis of Th2-high asthma. Collectively, prior studies have revealed the critical role of PLZF in the regulation of innate and adaptive immune cells involved in type 2 inflammation in the lung. Therefore, targeting PLZF signaling represents a promising therapeutic approach to suppress Th2-high asthma.


Assuntos
Asma , Leucemia , Humanos , Proteína com Dedos de Zinco da Leucemia Promielocítica , Imunidade Inata , Linfócitos/metabolismo , Pulmão/metabolismo , Inflamação , Dedos de Zinco , Fatores de Transcrição Kruppel-Like/genética , Fatores de Transcrição Kruppel-Like/metabolismo
17.
Gene ; 908: 148290, 2024 May 25.
Artigo em Inglês | MEDLINE | ID: mdl-38367853

RESUMO

Viral diseases are crucial determinants affecting tobacco cultivation, leading to a substantial annual decrease in production. Previous studies have demonstrated the regulatory function of the C3HC4 family of plant zinc finger proteins in combating bacterial diseases. However, it remains to be clarified whether this protein family also plays a role in regulating resistance against plant viruses. In this study, the successful cloning of the zinc finger protein coding gene NbZFP1 from Nicotiana benthamiana has been achieved. The full-length coding sequence of NbZFP1 is 576 bp. Further examination and analysis of this gene revealed its functional properties. The induction of NbZFP1 transcription in N. benthamiana has been observed in response to TMV, CMV, and PVY. Transgenic N. benthamiana plants over-expressing NbZFP1 demonstrated a notable augmentation in the production of chlorophyll a (P < 0.05). Moreover, NbZFP1-overexpressing tobacco exhibited significant resistance to TMV, CMV, and PVY, as evidenced by a decrease in virus copies (P < 0.05). In addition, the defense enzymes activities of PAL, POD, and CAT experienced a significant increase (P < 0.05). The up-regulated expression of genes of NbPAL, NbNPR1 and NbPR-1a, which play a crucial role in SA mediated defense, indicated that the NbZFP1 holds promise in enhancing the virus resistance of tobacco plant. Importantly, the results demonstrate that NbZFP1 can be considered as a viable candidate gene for the cultivation of crops with enhanced virus resistance.


Assuntos
Infecções por Citomegalovirus , Nicotiana , Nicotiana/genética , Clorofila A , Dedos de Zinco/genética , Antivirais , Doenças das Plantas/genética , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo
18.
Biomolecules ; 14(2)2024 Feb 04.
Artigo em Inglês | MEDLINE | ID: mdl-38397429

RESUMO

Zinc finger and BTB domain-containing 20 (ZBTB20), which was initially identified in human dendritic cells, belongs to a family of transcription factors (TFs) with an N-terminal BTB domain and one or more C-terminal DNA-binding zinc finger domains. Under physiological conditions, ZBTB20 acts as a transcriptional repressor in cellular development and differentiation, metabolism, and innate immunity. Interestingly, multiple lines of evidence from mice and human systems have revealed the importance of ZBTB20 in the pathogenesis and development of cancers. ZBTB20 is not only a hotspot of genetic variation or fusion in many types of human cancers, but also a key TF or intermediator involving in the dysregulation of cancer cells. Given the diverse functions of ZBTB20 in both health and disease, we herein summarize the structure and physiological roles of ZBTB20, with an emphasis on the latest findings on tumorigenesis and cancer progression.


Assuntos
Domínio BTB-POZ , Neoplasias , Animais , Humanos , Diferenciação Celular , Neoplasias/genética , Dedos de Zinco
19.
Cancer Res ; 84(6): 808-826, 2024 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-38345497

RESUMO

Heterochromatin loss and genetic instability enhance cancer progression by favoring clonal diversity, yet uncontrolled replicative stress leads to mitotic catastrophe and inflammatory responses that promote immune rejection. KRAB domain-containing zinc finger proteins (KZFP) contribute to heterochromatin maintenance at transposable elements (TE). Here, we identified an association of upregulation of a cluster of primate-specific KZFPs with poor prognosis, increased copy-number alterations, and changes in the tumor microenvironment in diffuse large B-cell lymphoma (DLBCL). Depleting two of these KZFPs targeting evolutionarily recent TEs, ZNF587 and ZNF417, impaired the proliferation of cells derived from DLBCL and several other tumor types. ZNF587 and ZNF417 depletion led to heterochromatin redistribution, replicative stress, and cGAS-STING-mediated induction of an interferon/inflammatory response, which enhanced susceptibility to macrophage-mediated phagocytosis and increased surface expression of HLA-I, together with presentation of a neoimmunopeptidome. Thus, cancer cells can exploit KZFPs to dampen TE-originating surveillance mechanisms, which likely facilitates clonal expansion, diversification, and immune evasion. SIGNIFICANCE: Upregulation of a cluster of primate-specific KRAB zinc finger proteins in cancer cells prevents replicative stress and inflammation by regulating heterochromatin maintenance, which could facilitate the development of improved biomarkers and treatments.


Assuntos
Heterocromatina , Neoplasias , Animais , Heterocromatina/genética , Dedos de Zinco/genética , Elementos de DNA Transponíveis , Primatas/genética , Inflamação/genética , Neoplasias/genética
20.
Int J Biol Sci ; 20(4): 1142-1159, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38385086

RESUMO

Human embryonic stem cells (hESCs) can proliferate infinitely (self-renewal) and give rise to almost all types of somatic cells (pluripotency). Hence, understanding the molecular mechanism of pluripotency regulation is important for applications of hESCs in regenerative medicine. Here we report that PATZ1 is a key factor that regulates pluripotency and metabolism in hESCs. We found that depletion of PATZ1 is associated with rapid downregulation of master pluripotency genes and prominent deceleration of cell growth. We also revealed that PATZ1 regulates hESC pluripotency though binding the regulatory regions of OCT4 and NANOG. In addition, we demonstrated PATZ1 is a key node in the OCT4/NANOG transcriptional network. We further revealed that PATZ1 is essential for cell growth in hESCs. Importantly, we discovered that depletion of PATZ1 drives hESCs to exploit glycolysis which energetically compensates for the mitochondrial dysfunction. Overall, our study establishes the fundamental role of PATZ1 in regulating pluripotency in hESCs. Moreover, PATZ1 is essential for maintaining a steady metabolic homeostasis to refine the stemness of hESCs.


Assuntos
Células-Tronco Embrionárias Humanas , Células-Tronco Pluripotentes , Humanos , Células-Tronco Embrionárias Humanas/metabolismo , Células-Tronco Pluripotentes/metabolismo , Zinco , Motivos AT-Hook , Diferenciação Celular/genética , Fatores de Transcrição/metabolismo , Dedos de Zinco , Proteínas Repressoras/metabolismo , Fatores de Transcrição Kruppel-Like/metabolismo
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