Zinc finger transcription factors designed for bispecific coregulation of erbb2 and erbb3 receptors : insights into ErbB receptor biology. (2005 Oct)
Zinc finger transcription factors designed for bispecific coregulation of erbb2 and erbb3 receptors : insights into ErbB receptor biology . signaling through the ErbB family of tyrosine kinase receptors in normal and cancer derived cell lines contributes to cell growth and differentiation . In this work , we altered the levels of erbb2 and erbb3 receptors , individually and in combination , by using 6 finger and 12 finger synthetic zinc finger protein artificial transcription factors ( ATFs ) in an epidermoid squamous cell carcinoma line , A431 . We successfully designed 12 finger ATFs capable of coregulating erbb3 and ICAM 1 or erbb2 and erbb3 . With ATFs , the effects of changes in erbb2 and erbb3 receptor levels were evaluated by using cell proliferation , cell migration , and cell signaling assays . Cell proliferation was increased when erbb2 and erbb3 were both overexpressed . Cell migration on collagen was decreased when erbb2 was down regulated , yet migration on laminin was significantly increased with erbb3 overexpression . erbb2 and erbb3 overexpression also stimulated the phosphatidylinositol 3 kinase and mitogen activated protein kinase pathways . Our ATF approach has elucidated differences in ErbB receptor mediated proliferation , migration , and intracellular signaling that cannot be explained merely by the presence or absence of particular ErbB receptors and emphasizes the dynamic nature of the ErbB signaling system . The transcription factor approach developed here provides a gene economical route to the regulation of multiple genes and may be important …

Genomic organisation and characterisation of the neural sex determination gene fruitless ( fru ) in the hawaiian species drosophila heteroneura. (2000 May)
genomic organisation and characterisation of the neural sex determination gene fruitless ( fru ) in the hawaiian species drosophila heteroneura . there are several mechanisms for the determination of sex . sexual behaviour is part of the sex determination cascade , and in drosophila melanogaster male courtship is controlled in part by the fruitless gene . As part of a study of sexual behaviour in hawaiian drosophila , we have cloned the neural sex determination gene fru from the hawaiian picture wing species drosophila heteroneura . The fru gene has at least seven exons covering a region of 18kb and encodes three transcripts , fruA , fruB and fruC . Each transcript encodes a single ORF of 841 , 678 and 691aa , respectively . The FRUA and FRUB proteins have a BTB protein protein binding domain and two zinc finger like domains and are well conserved with the D . melanogaster proteins . The FRUC protein has a BTB domain but no zinc finger like domains . The fru gene is expressed in 1 7 day old adult males as a 5 . 1kb transcript . This transcript is not seen in adult females , so the fru gene has a different pattern of sex differential expression in the hawaiian drosophila compared with D . melanogaster .

Synthetic zinc finger peptides : old and novel applications. (2004 Jul)
synthetic zinc finger peptides : old and novel applications . In the last decade , the efforts in clarifying the interaction between zinc finger proteins and DNA targets strongly stimulated the creativity of scientists in the field of protein engineering . In particular , the versatility and the modularity of zinc finger ( ZF ) motives make these domains optimal building blocks for generating artificial zinc finger peptides ( ZFPs ) . ZFPs can act as transcription modulators potentially able to control the expression of any desired gene , when fused to an appropriate effector domain . artificial ZFPs open the possibility to re program the expression of specific genes at will and can represent a powerful tool in basic science , biotechnology and gene therapy . In this review we will focus on old , novel and possible future applications of artificial ZFPs .

Combining structure based design with phage display to create new Cys ( 2 ) His ( 2 ) zinc finger … (2000 Oct)
combining structure based design with phage display to create new Cys ( 2 ) His ( 2 ) zinc finger dimers . background : several strategies have been reported for the design and selection of novel DNA binding proteins . Most of these studies have used Cys ( 2 ) His ( 2 ) zinc finger proteins as a framework , and have focused on constructs that bind DNA in a manner similar to zif268 , with neighboring fingers connected by a canonical ( krüppel type ) linker . This linker does not seem ideal for larger constructs because only modest improvements in affinity are observed when more than three fingers are connected in this manner . Two strategies have been described that allow the productive assembly of more than three canonically linked fingers on a DNA site : connecting sets of fingers using linkers ( covalent ) , or assembling sets of fingers using dimerization domains ( non covalent ) . results : using a combination of structure based design and phage display , we have developed a new dimerization system for Cys ( 2 ) His ( 2 ) zinc fingers that allows the assembly of more than three fingers on a desired target site . Zinc finger constructs employing this new dimerization system have high affinity and good specificity for their target sites both in vitro and in vivo . constructs that recognize an asymmetric binding site as heterodimers can be obtained through substitutions in the zinc …

Mizip , a highly conserved , vertebrate specific melanin concentrating hormone receptor 1 interacting zinc finger protein. (2002 Sep)
mizip , a highly conserved , vertebrate specific melanin concentrating hormone receptor 1 interacting zinc finger protein . using the yeast two hybrid system a novel protein was identified from human brain that interacts with the C terminus of melanin concentrating hormone receptor 1 ( MCH R1 ) . This protein , characterized by a myeloid translocation protein 8 , nervy , deaf1 proteins ( MYND ) zinc finger domain , is termed MCH R1 interacting zinc finger protein , mizip . It is fully conserved in man , rat , mouse and highly conserved in xenopus and zebrafish , but not detectable in invertebrates . mizip gene organization in human ( six exons on chromosome 9q34 . 3 ) and mouse is highly conserved , yet in rodents an additional exon is generated giving rise to alternatively spliced mrnas . mizip is expressed in brain , testis and stomach , where expression of MCH and MCH R1 was previously reported . mizip interaction with MCH R1 was verified by overlay and pull down assays as well as by co transfection experiments in human embryonic kidney 293 cells . mizip is cytoplasmically localized but gets recruited to the plasma membrane when cells are co transfected with MCH R1 supporting the notion that mizip is involved in the function of MCH R1 .

Gene regulation in planta by plant derived engineered zinc finger protein transcription factors. (2005 Apr)
Gene regulation in planta by plant derived engineered zinc finger protein transcription factors . The ability to modify plant traits is of great commercial potential in agricultural biotechnology . To this end we have engineered plant based zinc finger protein transcription factors ( ZFP TFs ) that minimize the use of non plant DNA sequences . This novel architecture supports the use of tandem arrays of zinc finger DNA recognition domains such that the ZFP TF binds a contiguous DNA target site thus emulating the design of ZFP TFs described previously for mammalian gene regulation . We show that this plant based ZFP TF architecture supports high affinity DNA binding while allowing the specificity of the DNA protein interaction to be determined by the amino acid sequences of the recognition helices . This plant based backbone thus supports the use of previously characterized DNA recognition helices originally identified in a mammalian ZFP context without using mammalian DNA sequences . moreover , we show that plant based ZFP TFs employing this new architecture can up regulate endogenous ADH activity by 20 fold in transgenic arabidopsis . Thus plant based ZFP TFs are shown to be potent regulators of gene expression in vivo .

Identification of DNA recognition sequences and protein interaction domains of the multiple Zn finger protein Roaz. (1998 Nov)
identification of DNA recognition sequences and protein interaction domains of the multiple Zn finger protein Roaz . Roaz , a rat C2H2 zinc finger protein , plays a role in the regulation of olfactory neuronal differentiation through its interaction with the Olf 1 / EBF transcription factor family . An additional role for the Roaz / Olf 1 / EBF heterodimeric protein is suggested by its ability to regulate gene activation at a distinct promoter lacking Olf 1 / EBF binding sites . using an in vitro binding site selection assay ( selex ) , we demonstrate that Roaz protein binds to novel inverted perfect or imperfect repeats of gcaccc separated by 2 bp . We show that Roaz is capable of binding to a canonical consensus recognition sequence with high affinity ( Kd 3 nM ) . analysis of the structural requirement for protein dimerization and DNA binding by Roaz reveals the role of specific zinc finger motifs in the Roaz protein for homodimerization and heterodimerization with the Olf 1 / EBF transcription factor . The DNA binding domain of Roaz is mapped to the N terminal 277 amino acids , containing the first seven zinc finger motifs , which confers weak monomeric binding to a single half site and a stronger dimeric binding to the inverted repeat in a binding site dependent manner . Full length protein can form dimers on both the inverted repeat and direct repeat but not on a single half site . these findings …

The transactivation domain within cysteine / histidine rich region 1 of CBP comprises two novel zinc binding modules. (2000 Jun)
The transactivation domain within cysteine / histidine rich region 1 of CBP comprises two novel zinc binding modules . cAMP response element binding protein binding protein ( CBP ) is a transcriptional coactivator that interacts with a number of DNA binding proteins and cofactor proteins involved in the regulation of transcription . relatively little is known about the structure of CBP , but it has been noted that it contains three domains that are rich in cysteine and histidine ( CH1 , CH2 , and CH3 ) . The sequence of CH2 conforms to that of a leukemia associated protein domain ( PHD finger ) , and it has been postulated that this and both CH1 and CH3 may be zinc finger domains . This has not , however , been demonstrated experimentally . We have studied CH1 and show that it is composed of two novel zinc binding modules , which we term zinc bundles . Each bundle contains the sequence Cys X ( 4 ) Cys X ( 8 ) His X ( 3 ) Cys , and we show that a synthetic peptide comprising one zinc bundle from CH1 can fold in a zinc dependent manner . CH3 also appears to contain two zinc bundles , one with the variant sequence Cys X ( 2 ) Cys X ( 9 ) His X ( 3 ) Cys , and we demonstrate that this variant motif also undergoes Zn ( II ) induced folding . CH1 acts as …

Variations of the C2H2 zinc finger motif in the yeast genome and classification of yeast zinc finger proteins. (1997 Jul)
variations of the C2H2 zinc finger motif in the yeast genome and classification of yeast zinc finger proteins . The prosite pattern Zinc finger C2H2 was extended to permit the detection of all C2H2 zinc fingers and their parent proteins in the recently completed sequence of the yeast genome . additionally , a new computer program was written that extracts other zinc binding motifs ( non C2H2 fingers ) , overlapping with the classical zinc finger pattern , from the found set of yeast C2H2 fingers . The complete and correct detection of all fingers is a prerequisite for the classification of the yeast zinc finger proteins in functional terms . The detected 53 yeast C2H2 zinc finger proteins do not contain finger clusters with 10 or more repeats , as is frequently found in higher eukaryotes . Only three proteins contain four or more fingers in a cluster . moreover , nearly all 27 yeast proteins with tandem arrays of two or three finger domains can be classified into nine subgroups with high sequence conservation in their finger clusters , in particular of their DNA recognition helices . these results and application of the recently elaborated finger / DNA recognition rules suggest that the yeast proteins belonging to the same subgroup may recognize identical or very similar DNA sites .

From repression domains to designer zinc finger proteins : a novel strategy of intracellular immunization against HIV. (1996 Oct)
From repression domains to designer zinc finger proteins : a novel strategy of intracellular immunization against HIV . tissue specific gene regulation of eukaryotic organisms is to a large extent mediated by transcription factors that interact with genomic DNA sequences in a sequence specific manner . The purpose of this synopsis is to put forward the potential of designer zinc finger proteins in treating infections of human immunodeficiency virus ( HIV ) . artificial transcription factors containing designer zinc finger structures fused to activator or repressor domains have been designated transcription response modifiers ( TRMs ) . The principle of engineering TRMs has been derived from the analysis of human krüppel type zinc finger genes and their products . Our research efforts encompass two fascinating features that are displayed by the human krüppel type zinc finger protein KOX1 : 1 ) the krüppel type zinc finger domains display rules of sequence specific DNA recognition , and 2 ) the evolutionarily conserved krüppel associated box ( KRAB ) presents one of the strongest transcriptional repressors identified so far in mammalian organisms . The KRAB repressor activity is postulated to be mediated through co repressor molecules , such as silencing mediating protein 1 ( SMP 1 ) . Thus , the structural organization and functional analysis of zinc finger proteins revealed principles of zinc finger transcription factors that are applicable for reducing the viral load in individuals infected with HIV . In this article , a novel concept of generating therapeutic proteins …

Identification of a zinc finger encoding gene in onchocerca volvulus. (1994 Jan)
identification of a zinc finger encoding gene in onchocerca volvulus . eukaryotic transcription factors can be identified and classified according to conserved DNA binding motifs and conserved regulatory domains . The functional and structural analysis , and the conservation of these genes among metazoans emphasizes the importance of these DNA binding proteins in development and differentiation . In order to identify genes with common DNA binding motifs in the genome of onchocerca volvulus we have cloned a zinc finger encoding gene of the structure C2 H2 . using the caenorhabditis elegans sex determining tra 1 gene as a probe we have identified two tra 1 related genes in the genome of O . volvulus . The zinc finger region of one of these genes ( ovzf1 ) was subcloned and sequenced . The predicted protein has at least eight consecutive zinc fingers and each finger possesses the characteristic paired cysteine and histidine residues and the proper spacing of the amino acids between the conserved residues .

Self association of the erythroid transcription factor GATA 1 mediated by its zinc finger domains. (1995 Jun)
Self association of the erythroid transcription factor GATA 1 mediated by its zinc finger domains . GATA 1 , the founding member of a distinctive family of transcription factors , is expressed predominantly in erythroid cells and participates in the expression of numerous erythroid cell expressed genes . GATA binding sites are found in the promoters and enhancers of globin and nonglobin erythroid genes as well as in the alpha and beta globin locus control regions . To elucidate how GATA 1 may function in a variety of regulatory contexts , we have examined its protein protein interactions . Here we show that GATA 1 self associates in solution and in whole cell extracts and that the zinc finger region of the molecule is sufficient to mediate this interaction . This physical interaction can influence transcription , as GATA 1 self association is able to recruit a transcriptionally active but DNA binding defective derivative of GATA 1 to promoter bound GATA 1 and result in superactivation . through in vitro studies with bacterially expressed glutathione S transferase fusion proteins , we have localized the minimal domain required for GATA 1 self association to 40 amino acid residues within the C terminal zinc finger region . finally , we have detected physical interaction of GATA 1 with other GATA family members ( GATA 2 and GATA 3 ) also mediated through the zinc finger domain . these findings have broad implications for the involvement of GATA factors in transcriptional control . …

Zinc finger proteins in oncogenesis : DNA binding and Gene regulation. (1993 Jul)
Zinc finger proteins in oncogenesis : DNA binding and Gene regulation . conference proceedings . noordwijkerhout , The netherlands , september 6 9 , 1992 .

Treble clef finger a functionally diverse zinc binding structural motif. (2001 Apr)
treble clef finger a functionally diverse zinc binding structural motif . detection of similarity is particularly difficult for small proteins and thus connections between many of them remain unnoticed . structure and sequence analysis of several metal binding proteins reveals unexpected similarities in structural domains classified as different protein folds in SCOP and suggests unification of seven folds that belong to two protein classes . The common motif , termed treble clef finger in this study , forms the protein structural core and is 25 45 residues long . The treble clef motif is assembled around the central zinc ion and consists of a zinc knuckle , loop , beta hairpin and an alpha helix . The knuckle and the first turn of the helix each incorporate two zinc ligands . treble clef domains constitute the core of many structures such as ribosomal proteins L24E and S14 , RING fingers , protein kinase cysteine rich domains , nuclear receptor like fingers , LIM domains , phosphatidylinositol 3 phosphate binding domains and His Me finger endonucleases . The treble clef finger is a uniquely versatile motif adaptable for various functions . This small domain with a 25 residue structural core can accommodate eight different metal binding sites and can have many types of functions from binding of nucleic acids , proteins and small molecules , to catalysis of phosphodiester bond hydrolysis . treble clef motifs are frequently incorporated in larger structures or occur in doublets . present analysis suggests that the …

A novel zinc finger protein is encoded by the arabidopsis LSD1 gene and functions as a negative regulator of plant … (1997 Apr)
A novel zinc finger protein is encoded by the arabidopsis LSD1 gene and functions as a negative regulator of plant cell death . arabidopsis Isd1 mutants are hyperresponsive to cell death initiators and fail to limit the extent of cell death . superoxide is a necessary and sufficient signal for cell death propagation . Thus , LSD1 monitors a superoxide dependent signal and negatively regulates a plant cell death pathway . We isolated LSD1 via its map position . The predicted LSD1 protein contains three zinc finger domains , defined by cxxcxrxxlmyxxgasxvxcxxc . these domains are present in three additional arabidopsis genes , suggesting that LSD1 defines a zinc finger protein subclass . LSD1 is constitutively expressed , consistent with the mutant phenotype . alternate splicing gives rise to a low abundance mRNA encoding an extra five amino terminal amino acids . We propose that LSD1 regulates transcription , via either repression of a prodeath pathway or activation of an antideath pathway , in response to signals emanating from cells undergoing pathogen induced hypersensitive cell death .

Nuclear transport and phosphorylation of the RNA binding xenopus zinc finger protein XFG 5 1. (1993 Mar)
nuclear transport and phosphorylation of the RNA binding xenopus zinc finger protein XFG 5 1 . XFG 5 1 is a krüppel type xenopus zinc finger protein with specific RNA homopolymer binding activity in vitro . In the oocyte , the protein is distributed between nucleus and cytoplasm ; the nuclear fraction , not the cytoplasm , contains phosphorylated isoform ( s ) of XFG 5 1 . In vitro phosphorylation by use of oocyte / egg extracts or purified casein kinase II is specific to the amino terminal portion of the protein . The carboxy terminal zinc finger domain contains a signal sufficient for nuclear transport . overexpression of either full length XFG 5 1 or of the carboxy terminal portion alone , which maintains RNA binding and nuclear import activities , was achieved in xenopus embryos by mRNA injection . This treatment did not result in impaired regulation of development , suggesting that XFG 5 1 functions in a way distinct from the mode of action exemplified in the drosophila zinc finger protein krüppel .

Zinc finger like structure in U1 specific protein C is essential for specific binding to U1 snrnp. (1991 May)
Zinc finger like structure in U1 specific protein C is essential for specific binding to U1 snrnp . The U1 small nuclear ribonucleoprotein ( snrnp ) contains three specific proteins denoted 70K , A and C , in addition to the common proteins . specific functions of these proteins are not known although recently protein C was shown to be involved in the binding of U1 snrnp to the 5 splice site of a pre mRNA . unlike proteins A and 70K , U1 C lacks an RNA binding domain ( RNP 80 motif ) and does not appear to bind directly to U1 snrna . however , at the amino terminal end protein C contains a zinc finger like structure of the CC HH type found in transcription factor TF IIIA . several lines of evidence indicate that the zinc finger like structure is essential for the binding of protein C to U1 snrnp particles : i ) deletion analysis of protein C showed that the N terminal 45 amino acids are sufficient for binding to U1 snrnps , ii ) modification of the cysteine residues in the N terminal domain with N ethylmaleimide and iii ) single point mutations of the cysteines and histidines contributing to the putative zinc finger abolished binding of protein C to U1 snrnps . interestingly , unlike the proteins U1 A and U1 70K the U1 C protein is unable to bind to naked U1 snrna . On the other hand it is …

Solution structure of the focal adhesion adaptor pinch LIM1 domain and characterization of its interaction with the integrin linked kinase … (2001 May)
solution structure of the focal adhesion adaptor pinch LIM1 domain and characterization of its interaction with the integrin linked kinase ankyrin repeat domain . pinch is a recently identified adaptor protein that comprises an array of five LIM domains . pinch functions through LIM mediated protein protein interactions that are involved in cell adhesion , growth , and differentiation . The LIM1 domain of pinch interacts with integrin linked kinase ( ILK ) , thereby mediating focal adhesions via a specific integrin / ILK signaling pathway . We have solved the NMR structure of the pinch LIM1 domain and characterized its binding to ILK . LIM1 contains two contiguous zinc fingers of the CCHC and CCCH types and adopts a global fold similar to that of functionally distinct LIM domains from cysteine rich protein and cysteine rich intestinal protein families with CCHC and CCCC zinc finger types . Gel filtration and NMR experiments demonstrated a 1 : 1 complex between pinch LIM1 and the ankyrin repeat domain of ILK . A chemical shift mapping experiment identified regions in pinch LIM1 that are important for interaction with ILK . comparison of surface features between pinch LIM1 and other functionally different LIM domains indicated that the LIM motif might have a highly variable mode in recognizing various target proteins .

Sexual dimorphism of hepatic gene expression : novel biological role of KRAB zinc finger repressors revealed. (2003 Nov)
sexual dimorphism of hepatic gene expression : novel biological role of KRAB zinc finger repressors revealed .

Isolation and functional characterization of cDNA of serum amyloid A activating factor that binds to the serum amyloid A promoter. (1998 Dec)
isolation and functional characterization of cDNA of serum amyloid A activating factor that binds to the serum amyloid A promoter . serum amyloid A ( SAA ) , a plasma protein inducible in response to many inflammatory conditions , is associated with the pathogenesis of several diseases including reactive amyloidosis , rheumatoid arthritis , and atherosclerosis . We have previously reported an element of the SAA promoter , designated SAA activating sequence ( SAS ) , that is involved in the inflammation induced SAA expression , and a nuclear factor , SAS binding factor ( SAF ) , that interacts with the SAS element has been identified previously ( A . Ray and B . K . Ray , Mol . Cell . Biol . 16 : 1584 1594 , 1996 ) . To evaluate how SAF is involved in SAA promoter activation , we have investigated structural features and functional characteristics of this transcription factor . Our studies indicate that SAF belongs to a family of transcription factors characterized by the presence of multiple zinc finger motifs of the Cys2 His2 type at the carboxyl end . Of the three cloned SAF cdnas ( SAF 1 , SAF 5 , and SAF 8 ) , SAF 1 isoform showed a high degree of homology to MAZ / ZF87 / Pur 1 protein while SAF 5 and SAF 8 isoforms are unique and are related to SAF 1 / MAZ / ZF87 / Pur 1 at the zinc finger domains …