Many mental disorders are closely associated with problems that occur during brain development in early life. For instance, by 2 years of age, autistic children have larger brains than normal kids, likely due to, at least in part, excess production of neurons and support cells, the building blocks of the nervous system. In autistic brains, how neurons grow various thread-like processes also shows some abnormalities. The cause of autism is complex and likely involves many genetic factors. These developmental defects are also associated with mental disorders caused by single-gene mutations, such as Rett syndrome and fragile X syndrome, the most common form of inherited mental retardation, whose clinical features overlap with autism. However, what causes the developmental defects in brains of children with different mental disorders is largely unknown. In recent years, an exciting new regulatory pathway was discovered that may well contribute to the etiology of mental disorders. The major player in this novel pathway is a class of tiny molecules 21
California is the most populated state in the US and has a large number of patients suffering from various mental disorders. The proposed studies in this grant application will contribute to the mission of developing novel avenues through stem cell research for the diagnosis, prevention and treatment of mental disorders
SYNOPSIS: Dr. Fen-Biao Gao of the Gladstone Institute of Neurological Diseases proposes to study the role of microRNA in neuronal development, particularly miR-9a and miR-124a. They will establish a cell culture system where they can reproducibly induce neuronal differentiation by hESC, determine the role of miR-9a in the formation of neural progenitor cells, and examine the role of miR-124a on morphological differentiation of the cells. They will also examine the interaction of the FMR1 gene and miR-124a.
SIGNIFICANCE AND INNOVATION: The proposed experiments are innovative and significant.
The investigator will characterize two miRNAs in hES cells that he, and others, have shown using model systems are involved in neuronal development. The investigator has primarily worked in the Drosophila system and has just started to work with hES cells. A big positive with this proposal is that the investigator will use the CIRM seed money to begin hESC experiments in his lab.
The laboratory has extensive experience working with miRNAs and will be able to investigate the function of these genes using both hES cells and Drosophila. This is a very powerful dual approach that has the potential to quickly unravel the role miRNAs play in both mental disorders and neuronal development. This proposal will lay the groundwork for the eventual use of hES cells for the treatment of human CNS diseases.
The experiments outlined in this proposal may be able to be funded from other sources. However, reviewers at more traditional granting agencies may balk at funding this proposal because the PI’s laboratory does not yet have training working with hES cells.
STRENGTHS: The clear molecular and signalling focus of the experiments is a great strength of the proposal.
The applicant is an experienced and productive investigator who has long worked on protein-mRNA interactions in mammalian neuronal differentiation. In addition, he has worked on the role of miR-9a in neuronal development. He trained with Martin Raff for 2 years and then worked with Dr. Yuh Nung Jan developing an assay system for dendrite development and cell fate determination.
The proposed experiments are well within the capabilities of his laboratory
The CIRM seed grant will be used to test theories about the role miR9a and miR124a play in hES cells. These experiments build on observations that the principle investigator has made using Drosophila.
Aim 1 will establish hES cell culture conditions in the PIs laboratory while Aims 2 and 3 will characterize miR9a and 124a in hES cells. These three Aims should yield basic information on what role these miRNAs play in hES cells. Aim 4, an examination of a possible interaction between FMR1 and miR124a, will take advantage of the ability of hES cells to be differentiated into a large number of postmitotic neurons.
The experiments outlined in the proposal can be completed in the two-year time frame of the proposal.
WEAKNESSES: It is not clear if the overexpression of miR124a in postmitotic neurons, which already produce high levels of this miRNA, would result in the production of a phenotype since the targets of this miRNA may already be at very low levels (although overexpression of miR124a in fly neurons does appear to cause a phenotype). It might be more interesting to misexpress miR124a in undifferentiated hES cells to see if miR124 can cause these cells to become more neuronal-like (similar to the published experiments in which the misexpression of miR124 caused HeLa cells to assume a more neuronal-like mRNA expression profile).
The last part of Aim 4 will use the Gladstone Genomics Core to determine the expression pattern of miRNAs in hESCs and hESc-derived neural progenitor cells. This experiment will yield a list of miRNAs that will not be analyzed within the two-year scope of the CIRM Seed grant. While this is an experiment that will yield a lot of data it detracts from the rest of the grant, which is focused on the role two specific miRNAs may play in neuronal development.
DISCUSSION: This proposal is from a young investigator who will produce useful results. Reviewer 2 reviewed this from the perspective of miRNAs and found this to be the best of those reviewed. The PI doesn't have experience with ESCs and needs to interact with those who do since there is a lot of morphological variation in neurons. The environment of the Gladstone Institutes may be helpful in this regard since there is experience in miRNA work and a solid stem cell program there.