The recent identification of transcription factors (TFs) that can induce conversion of fibroblasts into pluripotent stem (iPS) cells makes it potentially possible to generate patient-specific neurons from fibroblasts. However, the neurons thus produced are difficult to obtain. The present project builds on preliminary results in which we demonstrate direct conversion of adult fibroblasts cells into neurons, referred to as 'induced neuronal cells'(iN cells), without an iPS intermediate. The resulting iN cells have the functional properties of neurons, including the ability to form functional synapses as assayed by electrophysiology. Thus, the iN cell technology provides a novel, more facile approach to generating and studying human neurons, and opens up a new avenue to investigating human disease processes. However, at this point the iN cell technology has only been developed for mouse fibroblasts, fundamental questions regarding the conversion process and the molecular identity of iN cells were not determined, the generation of iN cells from human fibroblasts has not yet been established, and most importantly, the feasibility of the iN cell technology to study diseases affecting neuronal function has not been demonstrated. In this project, we propose to address these important challenges in an interdisciplinary approach capitalizing from the combined expertises of the Wernig and Sidhof laboratories. We propose experiments that will systematically investigate the cellular and molecular identity of iN cells, and develop protocols to induce specific neuronal subpopulations from fibroblasts. These protocols will then be employed to model genetic diseases in mouse iN cells. Furthermore, we will extend our findings to human fibroblasts, with the long term goal to establish a cell model for neuropsychiatric diseases. Our goals will be pursued by a combination of tissue culture experiments with cells cultured from mice and humans, cell biology, molecular biology, and electrophysiology. We believe our proposed experiments have the potential to fundamentally change existing paradigms of cellular differentiation and epigenetic gene regulation, and could provide a novel platform to study human neurons from patients suffering from a variety of brain diseases.
This application will develop methods to generate neurons directly from non-neuronal cells, allowing the production of neurons from skin fibroblasts of human patients. These methods will then be used to test the effects of mutations associated with neuropsychiatric disorders on neuronal biology, with the long-term goal of establishing a better understanding of the pathomechanism of these diseases in human neurons.
|Ang, Cheen Euong; Wernig, Marius (2014) Induced neuronal reprogramming. J Comp Neurol 522:2877-86|
|Chanda, Soham; Ang, Cheen Euong; Davila, Jonathan et al. (2014) Generation of induced neuronal cells by the single reprogramming factor ASCL1. Stem Cell Reports 3:282-96|
|Chanda, Soham; Marro, Samuele; Wernig, Marius et al. (2013) Neurons generated by direct conversion of fibroblasts reproduce synaptic phenotype caused by autism-associated neuroligin-3 mutation. Proc Natl Acad Sci U S A 110:16622-7|
|Webb, Ashley E; Pollina, Elizabeth A; Vierbuchen, Thomas et al. (2013) FOXO3 shares common targets with ASCL1 genome-wide and inhibits ASCL1-dependent neurogenesis. Cell Rep 4:477-91|
|Yang, Nan; Zuchero, J Bradley; Ahlenius, Henrik et al. (2013) Generation of oligodendroglial cells by direct lineage conversion. Nat Biotechnol 31:434-9|
|Davila, Jonathan; Chanda, Soham; Ang, Cheen Euong et al. (2013) Acute reduction in oxygen tension enhances the induction of neurons from human fibroblasts. J Neurosci Methods 216:104-9|
|Lujan, Ernesto; Chanda, Soham; Ahlenius, Henrik et al. (2012) Direct conversion of mouse fibroblasts to self-renewing, tripotent neural precursor cells. Proc Natl Acad Sci U S A 109:2527-32|
|Pang, Zhiping P; Yang, Nan; Vierbuchen, Thomas et al. (2011) Induction of human neuronal cells by defined transcription factors. Nature 476:220-3|
|Marro, Samuele; Pang, Zhiping P; Yang, Nan et al. (2011) Direct lineage conversion of terminally differentiated hepatocytes to functional neurons. Cell Stem Cell 9:374-82|
|Ming, Guo-Li; Brustle, Oliver; Muotri, Alysson et al. (2011) Cellular reprogramming: recent advances in modeling neurological diseases. J Neurosci 31:16070-5|
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