This technology includes a system that allows for robust differentiation of human-induced pluripotent stem cells (iPSC) into motor neurons within a time frame of 7 to 10 days. To differentiate the iPSC, a stable transgene is inserted into the CLYBL safe harbor locus in the human genome using TALENs. The transgene allows for doxycycline-inducible expression of the transcription factors (NGN2, ISL1, and LHX3) that are needed for the cells to differentiate to motor neurons. The technology is described in detail in the protocol paper published by Fernandopulle et al, cited below.

This invention includes HeLa cells that are engineered to inducibly express a mutant form of ornithine decarboxylase that is targeted to the mitochondrial matrix and forms insoluble protein aggregates. The presence of unfolded proteins in the matrix causes the accumulation of the mitochondrial kinase PINK1 and the E3 ubiquitin ligase PARK2/Parkin. These proteins play a critical role in degrading the mitochondria where they are expressed, a process call mitophagy. Mutations in these two genes are associated with familial Parkinson disease.

The technology includes Pink1 knockout HeLa cells that were generated using CRISPR technology. Pink1 is the key master gene to trigger degradation of mitochondria, mitophagy, and is implicated in familial Parkinson Disease. Knocking out Pink1 allows us to study the roles of Pink1 in many aspects of mitophagy and to display Pink1-dependent or independent activity. To create the HeLa cells, two CRISPR gRNAs targeting exon 1 and exon 7 of the Pink1 genome were used for transfection with Cas9 and GFP-C1 reporter. Cells were sorted 2 days after transfection and plated out in 96-well plates.

This technology includes the generation and use of an antibody that can detect endogenous Neuroligin 4Y, NLGN4Y, a cell adhesion molecule on the X-chromosome. NLGN4Y is part of an X-Y pair with NLGN4X, which has been implicated with autism spectrum disorder (ASD) and intellectual disability. ASD has a sex bias etiology that is not well understood, affecting four times as many males as females. Previous work has revealed a potential pathogenic mechanism for male-bias based on mutations in NLGN4X and NLGN4Y. The use of the NLGN4Y antibody could be used to study potential mechanisms.

This technology relates to the generation and use of an antibody that recognizes the S1459 phosphorylated site of the GRIN2A gene, which encodes the GluN2A subunit of the NMDA receptor. This gene is widely accepted as an epilepsy-causative gene and has been implicated in autism spectrum disorder (ASD). The S1459 phosphorylation site was selected based on an identified mutation in an epilepsy patient. This antibody can be used to specifically visualize the localization of the phosphorylated version of the GRIN2A protein product in the brain.

This technology includes the creation and use of a mouse anti-Atlastin-1 monoclonal antibody (3194, IgG1). Mutations in Atlastin-1 are commonly found in hereditary spastic paraplegia, SPG3A. In addition, this protein is conserved in all eukaryotes, and it mediates fusion of endoplasmic reticulum tubules in cells, giving it its characteristic polygonal appearance. Thus, this protein is of interest to both those interested in disease pathogenesis and those studying basic cell biology.

This technology relates to the therapeutic use of antibodies to decrease the potential neurodegenerative effect of the HERV-K retrovirus. Previous work has shown that patients with Amyotrophic Lateral Sclerosis (ALS) can have HERV-K activation. In animal models, activation of HERV-K can lead to neurodegenerative symptoms similar to those exhibited by ALS patients. This neurodegenerative effect is thought to be caused by the release of HERV-K envelope proteins into the extracellular space. Work with monoclonal antibodies in vitro has neutralized the toxicity of this protein.

This invention includes human induced pluripotent stem cell (iPSC) lines that harbor a single copy dCas9-BFP-KRAB at the CLYBL safe harbor locus (mediating CRISPR inhibition of human gene expression) and/or a single copy of dox-inducible NGN2 at the AAVS1 locus (enabling the differentiation of the iPSCs into neurons). The CRISPR-mediated inhibition of human gene expression is maintained into the differentiated neurons, permitting functional studies of targeted genes in neurons.

This invention includes the generation and use of a polyclonal antibody for synapse-associated protein 102 (SAP102) and a polyclonal antibody that binds to mGluR7 when phosphorylated at Serine 862. Peptides of the sites were generated and injected into rabbits to create an immune response. Serum was collected from the rabbits that was then affinity purified. The specificity of the resulting polyclonal antibodies was then determined using biochemical techniques.

This invention includes the generation and use of monoclonal antibodies that specifically recognize either arginine vasopressin (AVP) or oxytocin (OT) when bound to neurophysins. The neurophysins (NPs) are a family of proteins that bind to hormones as they are released from the hypothalamus and make their way to the pituitary gland. Monoclonal antibodies were generated that specifically recognize vasopressin bound to a neurophysin (NP-AVP) or oxytocin bound to a neurophysin (NP-OT). Seven monoclonal antibodies were characterized.