Alternative splicing during motor neuron differentiation in Spinal Muscular Atrophy

Funding Type: 
Basic Biology II
Grant Number: 
RB2-01547
ICOC Funds Committed: 
$0
Stem Cell Use: 
Embryonic Stem Cell
iPS Cell
Cell Line Generation: 
Embryonic Stem Cell
iPS Cell
Public Abstract: 
Mutations in one of the duplicated Survival of Motor Neuron (SMN) genes lead to the progressive loss of motor neurons and subsequent development of Spinal Muscular Atrophy (SMA), a common, usually fatal, pediatric disease. Recent experimental evidence strongly supports the notion that reduced levels of SMN result in a general pre-mRNA splicing defect. The proposed study tests the hypothesis that low levels of SMN result in perturbed pre-mRNA splicing and mRNA transcript levels during motor neuron differentiation, growth, and sprouting, thus reducing the life span of motor neurons as is characteristic for SMA. To test this hypothesis, we plan to use high throughput sequencing technologies to evaluate gene expression and alternative splicing profiles throughout motor neurons differentiation of stem cells derived from normal or SMA patients. These genome-wide surveys will be complemented with experimental verification and computational analyses to gain insights into fluctuations of pre-mRNA splicing networks during the differentiation of stem cells into motor neurons. The results from the proposed experiments will provide expression profiles and an alternative splicing maps that will permit to determine how motor neuron differentiation is perturbed in SMA patients and perhaps other neurological diseases. The verification of the hypothesis will provide new insights into the biological processes that specify longevity of motor neurons and direct future research to identify alternative targets for SMA therapy.
Statement of Benefit to California: 
Recent experiments demonstrated that pre-mRNA splicing is required to establish gene expression profiles that dictate the longevity of developing motor neurons. The ability to differentiate human embryonic stem cells into motor neurons in cell culture permits an evaluation of gene expression and alternative pre-mRNA splicing throughout the differentiation process. This unique opportunity is available because the state of California has actively supported research using human embryonic stem cells. The results obtained from this proposal will establish gene expression and alternative splicing maps that link defined developmental events with gene expression and motor neuron differentiation. The resulting signature profiles will enable future studies investigating motor neuron longevity and they will likely identity new molecular targets to battle various motor neuron diseases that have in common premature motor neuron death. As such, the information gained from the proposed experiments would maintain California’s leading status in stem cell research and may provide the intellectual framework for the development of new therapeutic developments in the private sector. An additional benefit to California citizens could be the availability of cutting edge technologies in clinical trials carried out at California's research centers.
Progress Report: 
  • Public Summary of Scientific Progress:
  • The Hippo tumor suppressor pathway plays a major role in limiting tissue and organ growth. A major molecular function of the Hippo pathway is to inhibit the YAP oncoprotein, which promote tissue growth. Mutations in the Hippo pathway can result in tissue overgrowth and tumor formation. Inhibition of the Hippo pathway is also important for wound healing and tissue regeneration. We have investigated the mechanism of the Hippo pathway regulation and its role in stemness and differentiation. We found that the Hippo pathway is regulated by signals, such as cell contact. Our new data support a critical role of the Hippo pathway in maintenance of embryonic stem cell stemness and in vitro differentiation of stem cells. Therefore, activation of the Hippo pathway may promote differentiation while inhibition of this pathway may main stem cell population.
  • The Hippo tumor suppressor pathway was initially identified in the fruit fly to control tissue growth and organ size. Subsequent studies show that this pathway is highly conserved and also controls organ size in mammals. Hippo pathway controls organ size by regulating cell numbers. YAP and TAZ are two transcription factors that are inhibited by the Hippo pathway. Inhibition of YAP/TAZ represents the major functional output of the Hippo signaling. YAP and TAZ function to promote cell growth and organ size by promoting stem cell growth. Furthermore, mutation leading to dysregulation of the Hippo pathway is associated with cancer development. In fact, high YAP activity is frequently found in human cancer. Furthermore, TAZ has been shown to play important role in breast cancer stem cells. We have been investigating the mechanism of the Hippo pathway regulation and its role in stemness, differentiation, and tumorigenesis. We discovered the cell-cell contact and cell-matrix interaction play critical role in Hippo pathway regulation. In other words, the Hippo pathway can sense its neighbors and environment, and then relays these signals to tell the cell whether to proliferate, survive, or die. These functions are important for maintenance of both embryonic stem cells and tissue specific progenitor cells. Furthermore, we have isolated and characterized inhibitors that can modulate the Hippo pathway. These inhibitors are useful reagents for research and potential anti-cancer drugs. Finally, we have discovered new molecules that can either inhibit or activate the Hippo pathway. These new findings gain new insights into the Hippo pathway regulation and provide valuable scientific knowledge of targeting the Hippo pathway for therapeutic intervention.
  • The Hippo tumor suppressor pathway was initially identified in the fruit fly to control tissue growth and organ size. Subsequent studies show that this pathway is highly conserved and also controls organ size in mammals. Hippo pathway controls organ size by regulating cell numbers. YAP and TAZ are two transcription factors that are inhibited by the Hippo pathway. Inhibition of YAP/TAZ represents the major functional output of the Hippo signaling. YAP and TAZ function to promote cell growth and organ size by promoting stem cell growth. Furthermore, mutation leading to dysregulation of the Hippo pathway is associated with cancer development. In fact, high YAP activity is frequently found in human cancer. Furthermore, TAZ has been shown to play important role in breast cancer stem cells. We have been investigating the mechanism of the Hippo pathway regulation and its role in stemness, differentiation, and tumorigenesis. We discovered the cell-cell contact and cell-matrix interaction play critical role in Hippo pathway regulation. In other words, the Hippo pathway can sense its neighbors and environment, and then relays these signals to tell the cell whether to proliferate, survive, or die. These functions are important for maintenance of both embryonic stem cells and tissue specific progenitor cells. Furthermore, we have identified hormones that act through G-protein coupled receptor to modulate the Hippo-YAP activity. These results suggest possible physiological cues that may regulate YAP activity to control tissue stem cells, therefore to influence organ size and tissue regeneration. Finally, we have isolated and characterized inhibitors that can modulate the Hippo pathway. These inhibitors are useful reagents for research and potential anti-cancer drugs. Importantly, our study shows that some of the FDA approved drugs can potently inhibits YAP. These novel findings gain new insights into the Hippo pathway regulation and provide valuable scientific knowledge of targeting the Hippo pathway for therapeutic intervention.

© 2013 California Institute for Regenerative Medicine