119 results about "Transactivation" patented technology

The present invention relates to non-steroidal ligands for use in nuclear receptor-based inducible gene expression system, and a method to modulate exogenous gene expression in which an ecdysone receptor complex comprising: a DNA binding domain; a ligand binding domain; a transactivation domain; and a ligand is contacted with a DNA construct comprising: the exogenous gene and a response element; wherein the exogenous gene is under the control of the response element and binding of the DNA binding domain to the response element in the presence of the ligand results in activation or suppression of the gene.

The invention provides chimeric proteins having at least two functional protein units, each containing the dimerization domain of a member of the steroid / thyroid hormone nuclear receptor superfamily. The chimeric proteins can fold under crystallization conditions to form functional entities. The functional entities optionally contain a novel flexible peptide linker of variable lengths between at least two of the protein units. In a preferred embodiment, the linker is designed to be increased in increments of 12 amino acids each to aid in preparation of variant chimeric proteins. The DNA binding characteristics of the invention functional entities differ from those of wild-type complexes formed between “monomeric” receptors and their binding partners. Some functional entities, e.g. dimers expressed as fusion proteins, transactivate responsive promoters in a manner similar to wild-type complexes, while others do not promote transactivation and function instead essentially as constitutive repressors. The invention further provides nucleotide sequences encoding the invention chimeric proteins, cells containing such nucleotide sequences, and methods for using the invention chimeric proteins to modulate expression of one or more exogenous genes in a subject organism. In addition, isolated protein crystals suitable for x-ray diffraction analysis and methods for obtaining putative ligands for the invention chimeric proteins are provided.

In accordance with the present invention, it has been discovered that various members of the steroid / thyroid superfamily of receptors can interact with the insect-derived ultraspiracle receptor, to form multimeric species. Accordingly, the interaction of at least one member of the steroid / thyroid superfamily of receptors with at least the dimerization domain of the ultraspiracle receptor modulates the ability of said member of the steroid / thyroid superfamily of receptors to transactivate transcription of genes maintained under hormone expression control in the presence of the cognate ligand for said member of the superfamily.

In accordance with the present invention, it has been discovered that various members of the steroid / thyroid superfamily of receptors can interact to form multimeric species comprising a complex of more than one receptor. Accordingly, the interaction of a first receptor species with a second receptor species modulates the ability of the first receptor species to trans-activate transcription of genes maintained under hormone expression control in the presence of the cognate ligand for said first receptor.

The present invention relates to non-steroidal ligands for use in nuclear receptor-based inducible gene expression system, and a method to modulate exogenous gene expression in which an ecdysone receptor complex comprising: a DNA binding domain; a ligand binding domain; a transactivation domain; and a ligand is contacted with a DNA construct comprising: the exogenous gene and a response element; wherein the exogenous gene is under the control of the response element and binding of the DNA binding domain to the response element in the presence of the ligand results in activation or suppression of the gene.

The present disclosure relates generally to novel methods and compositions for using engineered reprogramming factor(s) for the creation of induced pluripotent stem cells (iPSCs) through a kinetically controlled process. Specifically, this disclosure relates to establishing combinations of reprogramming factors, including fusions between conventional reprogramming factors with transactivation domains, optimized for reprogramming various types of cells. More specifically, the exemplary methods disclosed herein can be used for creating induced pluripotent stem cells from various mammalian cell types, including human fibroblasts. Exemplary methods of feeder-free derivation of human induced pluripotent stem cells using synthetic messenger RNA are also disclosed.

The present invention relates to non-steroidal ligands for use in nuclear receptor-based inducible gene expression system, and a method to modulate exogenous gene expression in which an ecdysone receptor complex comprising: a DNA binding domain; a ligand binding domain; a transactivation domain; and a ligand is contacted with a DNA construct comprising: the exogenous gene and a response element; wherein the exogenous gene is under the control of the response element and binding of the DNA binding domain to the response element in the presence of the ligand results in activation or suppression of the gene.

A transactivation system and method for producing a foreign polypeptide of interest in cells of a host plant is disclosed. The transactivation system has two components. It has genetically transformed cells of the host plant having integrated in their nuclear genome, an inactive or silenced foreign nucleic acid sequence, which is capable of encoding, upon its activation, the foreign polypeptide of interest; and a recombinant RNA viral vector capable of infecting the cells of the host plant and encoding therein a factor for activating or facilitating the expression of inactive or silenced foreign nucleic acid sequence.

A Tat-based vaccine composition comprising at least one antigen coupled to at least one immunostimulatory lentivirus trans-activator of transcription (Tat) molecule wherein the antigen is a cancer antigen an infectious disease antigen or a fragment thereof and methods to treat disease by administering the Tat-based vaccine composition. An additional Tat-based vaccine composition comprising immunostimulatory lentivirus Tat is provided.

The present invention is related to an antibiotic inducible / repressible genetic construct for controlling the transcription of a gene of interes: by a cell. The genetic construct comprises a bi-directional antibiotic controlled activator-responsive promoter / operator sequence which is located between the gene of interest and a cistron encoding a reverse antibiotic controlled transactivator and controls the transcription of the gene of interest and of the cistron.

This application describes the cloning of p63, a gene at chromosome 3q27-29, that bears homology to the tumor suppressor p53. The p63 gene encodes at least six different isotypes. p63 was detected in a variety of human and mouse tissue and demonstrates remarkably divergent activities, such as the ability to transactivate p53 reporter genes and induce apoptosis. Isotopes of p63 lacking a transactivation domain act as dominant negatives towards the transactivation by p53 and p63.

The invention provides for the use of oligodeoxynucleotide decoys for the prophylactic or therapeutic treatment of diseases associated with the binding of endogenous transcription factors to genes involved in cell growth, differentiation and signalling or to viral genes. By inhibiting endogenous trans-activating factors from binding transcription regulatory regions, the decoys modulate gene expression and thereby regulating pathological processes including inflammation, intimal hyperplasia, angiogenesis, neoplasia, immune responses and viral infection. The decoys are administered in amounts and under conditions whereby binding of the endogenous transcription factor to the endogenous gene is effectively competitively inhibited without significant host toxicity. The subject compositions comprise the decoy molecules in a context which provides for pharmacokinetics sufficient for effective therapeutic use.

The present invention provides recombinant nucleic acid molecules encoding a chimeric transactivator protein including a DNA binding domain of a DNA binding protein and a protein domain capable of transcriptional activation. The present invention also provides recombinant viral and non-viral vectors that are able to infect and / or transfect and sustain expression of a biologically active chimeric transactivator proteins in mammalian cells. Also provided are host cell lines and non-human transgenic animals capable of expressing biologically active chimeric transactivator proteins. In another aspect, compositions and methods for treating or preventing ischemic damage associated with hypoxia-related disorders are provided.

The present invention provides nucleic acid molecules, DNA constructs, plasmids, and methods for post-transcriptional regulation of gene expression using RNA molecules to both repress and activate translation of an open reading frame. Repression of gene expression is achieved through the presence of a regulatory nucleic acid element (the cis-repressive RNA or crRNA) within the 5′ untranslated region (5′ UTR) of an mRNA molecule. The nucleic acid element forms a hairpin (stem / loop) structure through complementary base pairing. The hairpin blocks access to the mRNA transcript by the ribosome, thereby preventing translation. In particular, in embodiments of the invention designed to operate in prokaryotic cells, the stem of the hairpin secondary structure sequesters the ribosome binding site (RBS). In embodiments of the invention designed to operate in eukaryotic cells, the stem of the hairpin is positioned upstream of the start codon, anywhere within the 5′ UTR of an mRNA. A small RNA (trans-activating RNA, or taRNA), expressed in trans, interacts with the crRNA and alters the hairpin structure. This alteration allows the ribosome to gain access to the region of the transcript upstream of the start codon, thereby activating transcription from its previously repressed state.

A method for regulating expression of a tet operator-linked gene in a cell of a subject is disclosed. In one embodiment, the method involves introducing into the cell a nucleic acid molecule encoding a tetracycline-controllable transactivator (tTA), the tTA comprising a Tet repressor operably linked to a polypeptide which directly or indirectly activates transcription in eucaryotic cells; and modulating the concentration of a tetracycline, or analogue thereof, in the subject. Alternatively, in another embodiment, the method involves obtaining the cell from the subject, introducing into the cell a first nucleic acid molecule which operatively links a gene to at least one tet operator sequence, introducing into the cell a second nucleic acid molecule encoding a tTA, to form a modified cell, administering the modified cell to the subject, and modulating the concentration of a tetracycline, or analogue thereof, in the subject. The first and second nucleic acid molecule can be within a single molecule (e.g., in the same vector) or on separate molecules.

This invention provides a novel pyrin-only protein (POP2), polypeptides, nucleic acids encoding them and methods for making and using them. The polypeptides of this invention have nuclear factor-κB (NF-κB) modulating activity. NF-κB is pivotal for transactivation of cell-cycle regulatory, cytokine and adhesion molecule genes and is dysregulated in many cancers, neurodegenerative disorders, and inflammatory diseases. Proteins with Pyrin and / or caspase recruitment (CARD) domains have roles in apoptosis, innate immunity, and inflammation. Many pyrin domain proteins modulate NF-KB activity as well as participate in assembling both the perinuclear “apoptotic speck” and the pro-IL1β / IL-18 converting inflammasome complex. ‘Pyrin-only’ proteins are attractive as negative regulators of pyrin domain-mediated functions and one such protein, POP1, has been reported. We teach a second Pyrin-only protein (POP2). POP2 is a 294 nt single exon gene located on human chromosome 3 encoding a 97 amino acid protein with sequence and predicted structural similarity to other pyrin domains. Highly similar to pyrin domains in CATERPILLER (CLR, NLR, NALP) family proteins, POP2 is less like the prototypic Pyrin and ASC pyrin domains. POP2 is expressed principally in peripheral blood leukocytes and displays both cytoplasmic and nuclear expression patterns in transfected cells. TNFα-stimulated and p65 (RelA) induced NF-KB-dependent gene transcription is inhibited by POP2 in vitro by a mechanism involving changes in NF-κB nuclear import or distribution. While colocalizing with ASC in perinuclear specks, POP2 also inhibits the formation of specks by the CLR protein CIAS1 / NALP3. Together these observations indicate that POP2 is a negative regulator of NF-KB activity that may influence the assembly of pyrin-domain dependent complexes.

The present invention provides a panel of transcriptional activator fusion proteins which comprises both tetracycline controlled transactivator proteins and reverse tetracycline transactivator proteins. These transactivators have novel phenotypes such as altered basal transcriptional activity in the absence of doxycycline, altered induced transcriptional activity in the presence of doxycycline, or differential induction by tetracycline and analogs of tetracycline.

The present invention relates to methods to use non-steroidal ligands in nuclear receptor-based inducible gene expression system to modulate exogenous gene expression in which an ecdysone receptor complex comprising: a DNA binding domain; a ligand binding domain; a transactivation domain; and a ligand is contacted with a DNA construct comprising: the exogenous gene and a response element; wherein the exogenous gene is under the control of the response element and binding of the DNA binding domain to the response element in the presence of the ligand results in activation or suppression of the gene.

The present technology relates to a method of treating cancer by sensitizing human tumours to DNA damaging therapies through activating FBXO32 expression. Transactivation of FBXO32 through the inhibition of EZH2, a histone methyltransferase, decreases p21 protein induction which results in the sensitization of human tumours to chemotherapy. The method further provides a prognostic method to determine if a combination treatment would be effective.

The present invention relates to non-steroidal ligands for use in nuclear receptor-based inducible gene expression system, and a method to modulate exogenous gene expression in which an ecdysone receptor complex comprising: a DNA binding domain; a ligand binding domain; a transactivation domain; and a ligand is contacted with a DNA construct comprising: the exogenous gene and a response element; wherein the exogenous gene is under the control of the response element and binding of the DNA binding domain to the response element in the presence of the ligand results in activation or suppression of the gene.

The present invention is directed to a method for screening and identifying molecules that transactivate a neurotrophin receptor and mediate neuronal cell survival in the absence of neurotrophins which uses one or a combination of three different assays. The assays involve detecting the phosphorylation of a neurotrophin receptor, detecting the phosphorylation of phosphotidylinositol 3′-kinase or Akt enzyme, and assessing neuronal cell survival in the absence of neurotrophins.

The present invention discloses development of a model live vaccine for HIV, using an attenuated strain of Salmonella engineered to surface express specific HIV proteins and testing of this vaccine in mice. There are provided two recombinant plasmids, containing the Lpp-OmpA genes required for surface exposure, followed by the genes for the HIV-1 proteins, Reverse Transcriptase or Transactivating protein (Tat). These plasmids are electroporated into an attenuated strain of Salmonella, and antigen expression is verified. These live vaccines are then used to orally inoculate mice and the vaccinated mice are tested for fecal IgA response and helper T cell response specific for the HIV antigens.

Methods for altering levels in plants of one or more phenolic compounds that are intermediates or final products of the plant phenylpropanoid pathway are provided. One method comprises transforming a plant cell with an expression construct comprising a nucleic acid which encodes a transactivator protein comprising the myb domain of the maize “ZmMyb-IF35” protein and an activation domain. Another method comprises transforming a plant cell with an expression construct comprising a transgene which encodes an antisense ZmMyb-IF35 RNA. The present invention also relates to expression constructs and vectors used in the present methods, transformed plant cells and transgenic plants prepared according to the present methods, and the seeds of such transgenic plants.

The presently claimed invention offers programmable and precise regulation of Cas9 functions by utilizing a set of compact Cas9 derivatives created by deleting conserved HNH and / or REC-C domains based on the structural information across variant class 2 CRISPR effectors. In addition, a novel strategy for engineering the dimeric gRNA-guided nuclease by splitting the mini-dSaCas9 and fusing the FokI domain right after the split point is claimed to increase the on-target DNA cleavage efficiency and potentially reduce the off-target effect because of a closer proximity of dimeric FokI nuclease to the target sequence. By combining the optimized and compact gRNA expression cassette and the downsized SaCas9 derivatives, the entire CRISPR / Cas system with different effector domains for transactivation, DNA cleavage and base editing is loaded into a single AAV virus. Such an all-in-one AAV-CRISPR / Cas9 system will be particularly appealing in biomedical applications that require safe and efficient delivery in vivo.

Provided is a mosaic mouse model for use in determining the potency of an shRNA in vivo for reducing survival of cancer cells of chemotherapy-resistant leukemia. The syngeneic mouse recipient is transplanted with tet-on competent leukemia cells carrying a bicistronic nucleic acid construct comprising a promoter operably linked to a fusion gene associated with chemotherapy-resistant leukemia, and a sequence encoding a reverse tet-transactivator protein, such that both coding sequences are co-expressed from the promoter. Also provided are methods of treating soft tissue cancers.

The present invention provides a method for screening for progesterone receptor isoform specific ligands as well as a first method for screening for tissue-selective progesterone receptor ligands, both methods comprising selecting progesterone receptor isoform A or progesterone receptor isoform B selective ligands by means of an assay involving cells stably transfected with plasmids expressing the progesterone receptor isoform A or B. Furthermore, the present invention provides a second method for screening for tissue-selective progesterone receptor ligands, comprising in vivo tests in desired target tissues. The present invention further relates to cell lines suitable for this transactivation assay, a respective assay kit and medical uses of the isoform-specific and / or tissue-selective progesterone receptor ligands according to the present invention.

The invention provides a novel high throughput screening model for hepatitis C virus (HCV) resistant drugs. The high throughput screening model can be applied to research and drug screening of human HCV infection mechanisms, and is a reporting system carrying out efficient response aiming at HCV infection. After the HCV invades cells, protease NS3 / 4A expressed by the HCV is about to cut and release a tetracycline-regulated transactivator (rtTA) anchored on a mitochondrial outer membrane by an MAVS (mitochondrial antiviral signaling) protein, expression of a reporter gene is activated in the presence of doxycycline or tetracycline, and obvious fluorescence signals are observed in a certain time; or cell death is caused in the presence of GCV (gancilovir). The system is simple to operate, has short experimental period, high efficiency and strong stability, can be widely applied to real-time quantitative observation of HCV infection, flow cell sorting, high throughput compound screening, high throughput host cell target gene screening and molecular mechanism research of HCV infection.

The present invention is directed generally to compositions and methods for expressing recombinant proteins in a mammalian host cell using a co-expressed transcriptional activator. In particular, the invention provides vectors, host cells, and methods of expressing at least one desired polypeptide by transfecting a mammalian host cell with cistrons encoding a transactivator, a desired polypeptide, and an apoptosis-protective protein.

The present invention discloses a Tat PTD-Endostatin recombination protein, which is a fusion protein comprising a protein transduction domain of human immunodeficiency virus (HIV) transactivation transduction protein Tat and human endostatin, wherein the amino acid sequence of the protein transduction domain of the HIV transactivation transduction protein Tat is represented by SEQ ID NO.1, and the amino acid sequence of the human endostatin is represented by SEQ ID NO.2. The protein of the present invention has the following advantages that: the function of the endostatin can be maintained, wherein the function of the endostatin is that the endostatin can inhibit angiogenesis; the protein has advantages of high transduction efficiency, easy blood-brain barrier crossing, and easy blood-ocular barrier crossing; the protein can overcome the limitation of the poor membrane spanning effect of the endostatin and play the angiogenesis inhibition effect well; the protein can be used for treatments of various diseases caused by angiogenesis, including ocular vascular proliferative diseases and various tumors, such as diabetes retinopathy, non-small cell lung cancer, and the like.

This invention relates to a method to modulate exogenous gene expression in which an ecdysone receptor complex comprising: a DNA binding domain; a ligand binding domain; a transactivation domain; and a ligand is contacted with a DNA construct comprising: the exogenous gene and a response element; wherein the exogenous gene is under the control of the response element and binding of the DNA binding domain to the response element in the presence of the ligand results in activation or suppression of the gene. The ligands comprise a class of ketones.