This is a new R21 application to study in live human neurons the function of a bipolar disorder susceptibility gene encoding a neuronal calcium channel. Calcium ion channels are expressed in all excitable and many non-excitable cells to transduce electric signals at the cell membrane into chemical potentials of calcium influx. Calcium is a second messenger whose intracellular concentration controls many cellular functions including muscle contraction, neurotransmitter release, cell survival and growth. Calcium ion channels are important drug targets for hypertension and neuropathic pain. Coding mutations in the CACNA1C calcium ion channel gene cause a rare hereditary disorder called Timothy Syndrome, affecting the heart, the brain, and other organs. Bipolar disorder is a chronic and severe mental illness with 1-2% lifetime prevalence in population of the United States. Individuals with bipolar disorder suffer from recurrent episodes of mania and depression, and current bipolar treatments, such as lithium, are often ineffective in many patients. Treatment options for bipolar disorder are unlikely to advance substantially until the underlying molecular processes in pathogenic risk are better understood. There is a clear heritable risk in bipolar disorder, but it has been difficult to establish clear risk associations in any particular gene wit bipolar disorder. Large-scale genome-wide association studies in three independent cohorts and a large meta-analysis have revealed a strong and consistent association between bipolar disorder susceptibility and a genomic locus on chromosome 12 in the CACNA1C gene. Interestingly, the same genomic region was found to be associated with schizophrenia and major depression susceptibility last year, making it one of the most replicable and consistent associations in psychiatric genetics. These exciting genetic discoveries present unique opportunities to study how genomic variations in the CACNA1C gene may influence neuronal calcium channel activity. Recently, mouse and human fibroblasts have been directly converted to functional neuronal cells through ectopic expression of three transcription factors (ASCL1, MYT1L, BRN2). This technological breakthrough makes human-specific in vitro neuronal models possible. Transcription activator-like effector nuclease (TALEN) technology is a molecular engineering tool that recognizes unique sequences in a genomic locus, introduces site-specific changes in the native genome, and helps elucidate genotype-phenotype relationships in an isogenic background. We will use genome editing, together with a combination of gene expression, biochemical, and electrophysiological analyses, to reveal the functional role of intronic single nucleotide polymorphism (SNP) variations of CACNA1C in controlling L-type calcium channel activity in human induced neurons. Our work has the potential to provide novel insights into the molecular mechanism of a genetic risk locus for psychiatric illness and to validate a systematic approach to study the functional consequences of genetic variations in complex brain disorders.
This R21 proposal will test the hypothesis that the CACNA1C gene peak SNP associated haplotype, implicated in susceptibility to bipolar disorder, regulates Cav1.2 calcium channel function in human induced neurons. We will use reprogramming and genome editing, together with a combination of gene expression, biochemical, and electrophysiological analyses, to reveal the functional role of intronic variations of CACNA1C in controlling L- type calcium channel activity in human induced neurons.
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