Disclosed herein are methods and compositions for modulating expression of a PD1 gene.

Disclosed herein include methods, compositions, and kits suitable for robust and tunable control of payload gene expression. Some embodiments provide rationally designed circuits, including miRNA-level and/or protein-level incoherent feed-forward loop circuits, that maintain the expression of a payload at an efficacious level. The circuit can comprise a promoter operably linked to a polynucleotide encoding a fusion protein comprising a payload protein, a protease, and one or more self-cleaving peptide sequences. The payload protein can comprise a degron and a cut site the protease is capable of cutting to expose the degron. The circuit can comprise a promoter operably linked to a polynucleotide comprising a payload gene, a silencer effector cassette, and one or more silencer effector binding sequences.

The present invention relates to methods and compositions for reducing the immunogenicity of chimeric Notch receptors, and specifically to transcription factors useful for controlling gene expression delivered to tissues by such chimeric Notch receptors.

An adenovirus or adenoviral vector is described that includes a non-native nucleotide sequence capable of expressing a chimeric protein comprising an N-terminal nucleotide binding domain of transactivation response element DNA-binding protein (TDP-43), a C-terminal domain derived from a splicing repressor, and an autoregulatory element. Methods of using the adenovirus or adenoviral vector to treat degenerative diseases such as inclusion body myocytosis, amyotrophic lateral sclerosis, and frontotemporal dementia are also described.

In an embodiment, a device for modulating intracellular gene expression, the device having a calcium actuator component and a transcription reprogramming component. In another embodiment, a method for modulating intracellular gene expression, where the method includes inducing a system having a calcium actuator component and a transcription reprogramming component with at least one of light and a chemical, causing an increase in Ca.sup.2+, and translocating the transcription reprogramming component from cytosol to the nucleus. In a further embodiment, a method for modulating gene intracellular expression, where the method includes inducing a system having an NIR-stimulable optogenetic platform with at least one of light and a chemical, where the NIR-stimulable optogenetic platform facilitates Ca.sup.2+ release and the NIR-stimulable optogenetic platform is LOV2-SOAR, causing an increase in Ca.sup.2+, and translocating a calcium-responsive dCas9 fusion construct from cytosol to the nucleus, where the calcium-responsive dCas9 fusion construct is NFAT.sub.1-460-dCas9-VP64.

It is intended to provide a fusion polypeptide that regulates the transcription of a target gene. The present inventors have provided a fusion polypeptide comprising: a cell-penetrating peptide; a DNA-binding polypeptide; and a transcriptional regulator.

The present disclosure provides genome engineering proteins, e.g., nucleic acid binding domains and/or functional domains that have a net positive charge and are cell permeable and can be introduced into the cells without the use of a carrier such as micelles, vesicles, liposomes, and the like.

The present disclosure provides polypeptides, compositions thereof, and methods for suppressing expression of a target gene such as PDCD1, CTLA4, LAG3, or TIM-3. The polypeptides disclosed herein include a DNA binding domain (DBD) that binds to a sequence of the target gene and a transcriptional repressor domain that suppresses expression of the target gene. The transcriptional repressor domain may be a known transcriptional repressor or may be a novel transcriptional repressor disclosed herein. Also disclosed herein are novel transcriptional repressors that are conjugated to a heterologous DNA binding domain and mediate suppression of expression of a target gene bound by the DNA binding domain.

Engineered CRISPR-Cas9 nucleases with altered and improved PAM specificities and their use in genomic engineering, epigenomic engineering, and genome targeting.

The presently disclosed subject matter provides high-throughput assays, e.g., yeast three-hybrid assays, for detecting the presence of a molecule, e.g., therapeutic molecule. In certain embodiments, the presently disclosed subject matter provides genetically-engineered cells and assays for detecting tetracycline or derivatives thereof and kits for performing such assays.