Yeast methionine aminopeptidase type 1 is ribosome-associated and requires its N-terminal zinc finger domain for normal function in vivo

ABSTRACT The erythroid cell-specific transcription factor erythroid Krüppel-like factor (EKLF) is an important activator of β-globin gene expression. It achieves this by binding to the CACCC element at the β-globin promoter via its zinc finger domain. The coactivators CBP and P300 interact with, acetylate, and enhance its activity, helping to explain its role as a transcription activator. Here we show that EKLF can also interact with the corepressors mSin3A and HDAC1 (histone deacetylase 1) through its zinc finger domain. When linked to a GAL4 DNA binding domain, full-length EKLF or its zinc finger domain alone can repress transcription in vivo. This repressive activity can be relieved by the HDAC inhibitor trichostatin A. Although recruitment of EKLF to a promoter is required to show repression, its zinc finger domain cannot bind directly to DNA and repress transcription simultaneously. In addition, the target promoter configuration is important for enabling EKLF to exhibit any repressive activity. These results suggest that EKLF may function in vivo as a transcription repressor and play a previously unsuspected additional role in regulating erythroid gene expression and differentiation.

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Inhibition of multiple phases of human immunodeficiency virus type 1 replication by a dithiane compound that attacks the conserved zinc fingers of retroviral nucleocapsid proteins.

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.41.2.419 ◽

1997 ◽

Vol 41 (2) ◽

pp. 419-426 ◽

Cited By ~ 64

Author(s):

W G Rice ◽

D C Baker ◽

C A Schaeffer ◽

L Graham ◽

M Bu ◽

...

Keyword(s):

Human Immunodeficiency Virus ◽

Zinc Finger ◽

Human Immunodeficiency Virus Type ◽

Virus Type ◽

Zinc Fingers ◽

Immunodeficiency Virus ◽

Zinc Finger Domains ◽

Hiv 1

The human immunodeficiency virus type 1 (HIV-1) nucleocapsid p7 protein contains two retrovirus-type zinc finger domains that are required for multiple phases of viral replication. Chelating residues (three Cys residues and one His residue) of the domains are absolutely conserved among all strains of HIV-1 and other retroviruses, and mutations in these residues in noninfectious virions. These properties establish the zinc finger domains as logical targets for antiviral chemotherapy. Selected dithiobis benzamide (R-SS-R) compounds were previously found to inhibit HIV-1 replication by mediating an electrophilic attack on the zinc fingers. Unfortunately, reaction of these disulfide-based benzamides with reducing agents yields two monomeric structures (two R-SH structures) that can dissociated and no longer react with the zinc fingers, suggesting that in vivo reduction would inactivate the compounds. Through an extensive drug discovery program of the National Cancer Institute, a nondissociable tethered dithiane compound (1,2-dithiane-4,5-diol, 1,1-dioxide, cis; NSC 624151) has been identified. This compound specifically attacks the retroviral zinc fingers, but not other antiviral targets. The lead compound demonstrated broad antiretroviral activity, ranging from field isolates and drug-resistant strains of HIV-1 to HIV-2 and simian immunodeficiency virus. The compound directly inactivated HIV-1 virions and blocked production of infectious virus from cells harboring integrated proviral DNA. NSC 624151 provides a scaffold from which medicinal chemists can develop novel compounds for the therapeutic treatment of HIV infection.

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The Wilms tumour suppressor protein WT1 (+KTS isoform) binds alpha-actinin 1 mRNA via its zinc-finger domain

Biochemistry and Cell Biology ◽

10.1139/o06-065 ◽

2006 ◽

Vol 84 (5) ◽

pp. 789-798 ◽

Cited By ~ 35

Author(s):

A.A. Morrison ◽

J.P. Venables ◽

G. Dellaire ◽

M.R. Ladomery

Keyword(s):

Zinc Finger ◽

Nuclear Extract ◽

Start Codon ◽

Urogenital System ◽

Zinc Finger Domain ◽

Wilms Tumour ◽

Band Shift ◽

Mrna Targets

Mutations in WT1 are associated with developmental syndromes that affect the urogenital system and neoplasms, including Wilms tumour, acute myeloid leukemia, and breast and prostate cancers. The WT1 protein belongs to the early growth response family of zinc-finger transcription factors. Uniquely to WT1, an evolutionarily conserved alternative splice event inserts the tripeptide KTS, between zinc fingers 3 and 4. Whereas –KTS isoforms bind DNA and activate or repress transcription, +KTS isoforms bind DNA less efficiently and interact with splice factors and RNA in vitro and in vivo. Although candidate DNA targets have been found, physiological mRNA targets are yet to be defined. We examined the distribution of WT1 in ribonucleoprotein (RNP) complexes in nuclear extract prepared from M15 cells, a mouse mesonephric fetal kidney cell line. WT1 cofractionated with the splice factor PSF in large RNP particles ≥2 MDa. We also found that PSF co-immunoprecipitated with WT1, suggesting a functional interaction between these 2 multifunctional proteins. Using yeast three-hybrid library constructed from the co-immunoprecipitated RNA we found that WT1 (+KTS) binds close to or at the start codon of alpha-actinin 1 (ACTN1) mRNA. A band shift assay confirmed the ability of the WT1 zinc-finger domain (+KTS) to bind this sequence in vitro. ACTN1 is the first likely physiological mRNA target of WT1.

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Direct Protein Interactions Are Responsible for Ikaros-GATA and Ikaros-Cdk9 Cooperativeness in Hematopoietic Cells

Molecular and Cellular Biology ◽

10.1128/mcb.00296-13 ◽

2013 ◽

Vol 33 (16) ◽

pp. 3064-3076 ◽

Cited By ~ 10

Author(s):

Stefania Bottardi ◽

Lionel Mavoungou ◽

Vincent Bourgoin ◽

Nazar Mashtalir ◽

El Bachir Affar ◽

...

Keyword(s):

Zinc Finger ◽

Protein Interactions ◽

Transcriptional Activation ◽

Target Genes ◽

Transcription Elongation ◽

Hematopoietic Cells ◽

Elongation Factor ◽

Gene Promoters ◽

Zinc Finger Domain

Ikaros (Ik) is a critical regulator of hematopoietic gene expression. Here, we established that the Ik interactions with GATA transcription factors and cyclin-dependent kinase 9 (Cdk9), a component of the positive transcription elongation factor b (P-TEFb), are required for transcriptional activation of Ik target genes. A detailed dissection of Ik-GATA and Ik-Cdk9 protein interactions indicated that the C-terminal zinc finger domain of Ik interacts directly with the C-terminal zinc fingers of GATA1, GATA2, and GATA3, whereas the N-terminal zinc finger domain of Ik is required for interaction with the kinase and T-loop domains of Cdk9. The relevance of these interactions was demonstratedin vivoin COS-7 and primary hematopoietic cells, in which Ik facilitated Cdk9 and GATA protein recruitment to gene promoters and transcriptional activation. Moreover, the oncogenic isoform Ik6 did not efficiently interact with Cdk9 or GATA proteinsin vivoand perturbed Cdk9/P-TEFb recruitment to Ik target genes, thereby affecting transcription elongation. Finally, characterization of a novel nuclear Ik isoform revealed that Ik exon 6 is dispensable for interactions with Mi2 and GATA proteins but is essential for the Cdk9 interaction. Thus, Ik is central to the Ik-GATA-Cdk9 regulatory network, which is broadly utilized for gene regulation in hematopoietic cells.

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Zinc Finger Domain of Murine Leukemia Virus Nucleocapsid Protein Enhances the Rate of Viral DNA Synthesis in Vivo

Journal of Virology ◽

10.1128/jvi.76.15.7473-7484.2002 ◽

2002 ◽

Vol 76 (15) ◽

pp. 7473-7484 ◽

Cited By ~ 29

Author(s):

Wen-hui Zhang ◽

Carey K. Hwang ◽

Wei-Shau Hu ◽

Robert J. Gorelick ◽

Vinay K. Pathak

Keyword(s):

Secondary Structure ◽

Dna Synthesis ◽

Zinc Finger ◽

Rna Secondary Structure ◽

Leukemia Virus ◽

Template Switching ◽

Zinc Finger Domain ◽

Wild Type ◽

Murine Leukemia

ABSTRACT In vitro studies have indicated that retroviral nucleocapsid (NC) protein facilitates both DNA synthesis by reverse transcriptase (RT) and annealing of the nascent DNA with acceptor template. Increasing the rate of DNA synthesis is expected to reduce the frequency of RT template switching, whereas annealing the nascent DNA with acceptor template promotes template switching. We performed a mutational analysis of the murine leukemia virus (MLV) NC zinc finger domain to study its effect on RT template switching in vivo and to explore the role of NC during reverse transcription. The effects of NC mutations on RT template switching were determined by using a previously described in vivo direct-repeat deletion assay. A trans-complementation assay was also developed in which replication-defective NC mutants were rescued by coexpression of replication-defective RT mutants that provided wild-type NC in trans. We found that mutations in the MLV NC zinc finger domain increased the frequency of template switching approximately twofold. When a predicted stem-loop RNA secondary structure was introduced into the template RNA, the template-switching frequency increased 5-fold for wild-type NC and further increased up to an additional 6-fold for NC zinc finger domain mutants, resulting in an overall increase of as much as 30-fold. Thus, wild-type NC increased the efficiency with which RT was able to reverse transcribe through regions of RNA secondary structure that might serve as RT pause sites. These results provide the first in vivo evidence that NC enhances the rate of DNA synthesis by RT in regions of the template possessing stable RNA secondary structure.

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Lambda-interacting protein, a novel protein that specifically interacts with the zinc finger domain of the atypical protein kinase C isotype lambda/iota and stimulates its kinase activity in vitro and in vivo.

Molecular and Cellular Biology ◽

10.1128/mcb.16.1.105 ◽

1996 ◽

Vol 16 (1) ◽

pp. 105-114 ◽

Cited By ~ 85

Author(s):

M T Diaz-Meco ◽

M M Municio ◽

P Sanchez ◽

J Lozano ◽

J Moscat

Keyword(s):

Protein Kinase C ◽

Protein Kinase ◽

Zinc Finger ◽

Zinc Finger Domain ◽

Interacting Protein ◽

Kinase C ◽

Novel Protein

The members of the atypical subfamily of protein kinase C (PKC) show dramatic structural and functional differences from other PKC isotypes. Thus, in contrast to the classical or novel PKCs, they are not activated by diacylglycerol or phorbol esters. However, the atypical PKCs are the target of important lipid second messengers such as ceramide, phosphatidic acid, and 3'-phosphoinositides. The catalytic and pseudosubstrate sequences in the two atypical PKCs (lambda/iota PKC and zeta PKC) are identical but are significantly different from those of conventional or novel PKCs. It has been shown that microinjection of a peptide with the sequence of the pseudosubstrate of the atypical PKC isotypes but not of alpha PKC or epsilon PKC dramatically inhibited maturation and NF-kappa B activation in Xenopus oocytes, as well as reinitiation of DNA synthesis in quiescent mouse fibroblasts. This indicates that either or both atypical isoforms are important in cell signalling. Besides the pseudosubstrate, the major differences in the sequence between lambda/iota PKC and zeta PKC are located in the regulatory domain. Therefore, any functional divergence between the two types of atypical PKCs will presumably reside in that region. We report here the molecular characterization of lambda-interacting protein (LIP), a novel protein that specifically interacts with the zinc finger of lambda/iota PKC but not zeta PKC. We show in this paper that this interaction is detected not only in vitro but also in vivo, that LIP activates lambda/iota PKC but not zeta PKC in vitro and in vivo, and that this interaction is functionally relevant. Thus, expression of LIP leads to the transactivation of a kappa B-dependent promoter in a manner that is dependent on lambda/iota PKC. To our knowledge, this is the first report on the cloning and characterization of a protein activator of a PKC that binds to the zinc finger domain, which has so far been considered a site for binding of lipid modulators. The fact that LIP binds to lambda/iota PKC but not to the highly related zeta PKC isoform suggests that the specificity of the activation of the members of the different PKC subfamilies will most probably be accounted for by proteins like LIP rather than by lipid activators.

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Chemical shift assignments of zinc finger domain of methionine aminopeptidase 1 (MetAP1) from Homo sapiens

Biomolecular NMR Assignments ◽

10.1007/s12104-015-9608-0 ◽

2015 ◽

Vol 9 (2) ◽

pp. 351-353

Author(s):

Kavitha Rachineni ◽

Tarun Arya ◽

Kiran Kumar Singarapu ◽

Anthony Addlagatta ◽

Jagadeesh Bharatam

Keyword(s):

Chemical Shift ◽

Zinc Finger ◽

Homo Sapiens ◽

Zinc Finger Domain ◽

Chemical Shift Assignments ◽

Methionine Aminopeptidase

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Genetic Heterogeneity of KLF1, a Master Regulator of Erythropoiesis, Revealed an Autosomal Recessive Ψβ-Thalassemia and a Very Strong Promoter In Vivo

Blood ◽

10.1182/blood-2020-143018 ◽

2020 ◽

Vol 136 (Supplement 1) ◽

pp. 7-7

Author(s):

Laura Grech ◽

Joseph Borg ◽

Ruth Galdies ◽

Carmen Attard ◽

Christian A. Scerri ◽

...

Keyword(s):

Zinc Finger ◽

Genetic Heterogeneity ◽

Autosomal Recessive ◽

Globin Gene ◽

Constant Region ◽

Zinc Finger Domain ◽

Wild Type ◽

Strong Promoter ◽

The Family

The Erythroid Kruppel-like Factor 1 or KLF1 is a transcription factor that functioned in the early stage genetic programming of erythroid progenitors to promote physiological γ to β globin gene switching. Indeed, we showed that a truncation mutation (p.K288X) in KLF1 that deleted the DNA binding zinc finger domain resulted in marked KLF1 deficiency and a hematological condition that resembled a hereditary persistence of fetal hemoglobin (HbF) or a β thalassemia. Here, we describe five additional families with the same p.K288X mutation but varied hematological and HbF levels together with genetic and phenotypic data on a 600 data-set from the same Maltese population. The data accounted for a strong promoter embedded within a region of genetic heterogeneity of KLF1 that led to a ψβ thalassemia. Whole genome sequencing on 15 subjects of six families (FamF1 - FamF6) segregating (two) p.K288X frameworks of KLF1 had variable degrees of microcytosis (MCV; 76.1fL -77.4 fL) and HbF levels (HbF 2480 mg/dL - 802mg/dL) due to complex heterozygosity between promoter, coding and truncating mutations in KLF1. Case II-6 of FamF1 with the highest HbF (2480 mg/dL) had 2 promoter and 2 coding mutations in cis and in trans to the p.K288X truncation. Nine (9) KLF1 frameworks (A - I) were derived by transmission disequilibrium analyses of the family data, each assembled from 15 mutations and resulting in 7 genotypes among the families. The p.K288X truncation was found on a second rarer framework. Additional, rarer KLF1 frameworks were found with haploview in the population dataset. The population dataset was made up of 198 β thalassemia heterozygotes and 400 others from the clinic and the biobank and that had no β globin gene mutations, variable blood counts or hemoglobin profiles or both. They were older than 2 years of age, not pregnant and had normal iron levels. The number of KLF1 mutations differed from 0 in the wild-type framework A to 6 in one of the rare frameworks X. Six mutations were in the promoter region and 6 were in the coding region that defined a "KLF1 Variable Region" 5' genomic coordinate 12887183 - 12888273, whereas very few were found in the 3' (genomic coordinate 12884589 - 12884752) that defined the KLF1 "Constant Region" The Constant region has also been evolutionary conserved. It included the zinc finger domain and the proteasome binding site. The genotype - phenotype correlations and the family data were consistent with an autosomal recessive condition that resembled a β thalassemia, thus a ψβ thalassemia. It differed from a silent thalassemia because the β globin gene sequence was wild type. It provided a diagnosis for families with iron resistant microcytosis and borderline hemoglobin phenotypes suitable for counselling in the clinical setting. It was consistent with the observation among the families regarding the high strength of the KLF1 promoter. Multiple "hits" were necessary to suppress the biosynthesis of KLF1 for hemoglobin switching to escape perinatal suppression. The effect of the 6 promoter mutations were confirmed in vitro with native and induced K562 and Hek293T cell lines. Presumably, during normal development, the strong promoter served to rapidly drown KLF1 binding sites with KLF1 molecules to direct progenitors to erythropoiesis with the appropriate adult hemoglobin profile in the perinatal period. The diagnosis of the patients with selected genotypes due to compound and double heterozygosities in promoter and coding sequences shall further permit quantification of differential promoter function in vivo. Figure Disclosures No relevant conflicts of interest to declare.

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Structure of the zinc finger domain encompassing residues 13–51 of the nucleocapsid protein from simian immunodeficiency virus

Biochemical Journal ◽

10.1042/bj20051203 ◽

2006 ◽

Vol 393 (3) ◽

pp. 725-732 ◽

Cited By ~ 15

Author(s):

Nelly Morellet ◽

Hervé Meudal ◽

Serge Bouaziz ◽

Bernard Pierre Roques

Keyword(s):

Simian Immunodeficiency Virus ◽

Zinc Finger ◽

Solution Structure ◽

Human Populations ◽

Zinc Finger Domain ◽

Nucleocapsid Proteins ◽

Immunodeficiency Virus ◽

Hiv Type 1 ◽

Hiv 1

The NCps (nucleocapsid proteins) of HIV-1 (HIV type 1), HIV-2 and SIV (simian immunodeficiency virus) are small highly basic proteins, characterized by the presence of two CCHC ZF (zinc finger) domains. NCps, closely associated with the dimeric RNA genome in the core of the virus particle, were shown to promote the specific encapsidation of the viral RNA and are implicated in reverse transcription. Solution structure of the HIV-1 NCp7 and complexes of NCp7 with RNA or DNA showed the critical relationships between the structure and its various functions. HIV-1 and HIV-2 have resulted respectively from transmissions of SIV from chimpanzees and sooty mangabeys. It has been shown that the SIVlhoest (SIV from l'Hoest monkeys) also has the potential to infect human populations. Since monkeys are of great interest for clinical studies of antiviral drugs, the structure of (13-51)NCp8 (zinc finger domain of NCp8, encompassing residues 13–51) from SIVlhoest was determined by NMR to appraise the influence of major differences in the sequence, since Glu21, Gly43 and Met46 in NCp7 are replaced by Pro, Glu and Phe respectively in this particular NCp8. The structure of (13-51)NCp8 is very well defined, and surprisingly the structure of each ZF is similar in NCp7 and NCp8. Moreover, contrary to NCp7, the two ZFs are strongly locked to each other in this NCp8. This first reported structure of a simian NCp8 compared with that of NCp7 shows that the main structural differences occur at the flexible linker between the two ZFs but the essential residues responsible for the interaction with oligonucleotides adopt the same orientation in the two proteins.

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Interplay between HIV-1 Vpr and Sp1 Modulates p21WAF1Gene Expression in Human Astrocytes

Journal of Biological Chemistry ◽

10.1074/jbc.m403792200 ◽

2004 ◽

Vol 279 (44) ◽

pp. 46046-46056 ◽

Cited By ~ 43

Author(s):

Shohreh Amini ◽

Marcus Saunders ◽

Kimberly Kelley ◽

Kamel Khalili ◽

Bassel E. Sawaya

Keyword(s):

Cell Cycle ◽

Human Immunodeficiency Virus ◽

Dna Binding ◽

Zinc Finger ◽

Human Immunodeficiency Virus Type ◽

Binding Activity ◽

Zinc Finger Domain ◽

Dna Binding Activity ◽

Immunodeficiency Virus

The Vpr (viral protein R) of human immunodeficiency virus, type 1, which is expressed during the late stage of the viral infection, has received special attention because of its ability to control transcription of the human immunodeficiency virus, type 1, long terminal repeat and to influence cell cycle progression. Here we demonstrate that Vpr has the ability to regulate transcription of the cyclin-dependent kinase inhibitor, p21WAF1(p21), one of the key regulators of the cell cycle, in human astrocytic cells. The results from transcription assays demonstrated that Vpr augments promoter activity of p21 through the GC-rich region located between nucleotides -84 and -74 with respect to the +1 transcription start site. Activation of p21 by Vpr required cooperativity of Sp1, which binds to the DNA sequence spanning -84 to -74. Results from bandshift assay revealed an increased level of Sp1 DNA binding activity in the presence of Vpr. Furthermore, Vpr was able to associate with Sp1 via the zinc finger domain located in the C-terminal region of Sp1. Functional studies revealed that the cooperativity between Vpr and Sp1 requires the zinc finger domain at the C terminus and the glutamine-rich domain at the N terminus of Sp1. Expression of p53 further enhanced the level of Vpr-Sp1-mediated transcription activation of p21 through the sequence spanning -84 to -74 and increased the DNA binding activity of Sp1 in the presence of Vpr. Results from glutathioneS-transferase pull-down assay showed the association of Vpr with p53 in extracts containing Sp1. Altogether, the outcome of our functional and binding studies suggested that the physical interaction of Vpr with Sp1 and p53 could modulate transcriptional activity of p21.

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