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A global in vivo Drosophila RNAi screen identifies NOT3 as a conserved regulator of heart function.

ABSTRACT: Heart diseases are the most common causes of morbidity and death in humans. Using cardiac-specific RNAi-silencing in Drosophila, we knocked down 7061 evolutionarily conserved genes under conditions of stress. We present a first global roadmap of pathways potentially playing conserved roles in the cardiovascular system. One critical pathway identified was the CCR4-Not complex implicated in transcriptional and posttranscriptional regulatory mechanisms. Silencing of CCR4-Not components in adult Drosophila resulted in myofibrillar disarray and dilated cardiomyopathy. Heterozygous not3 knockout mice showed spontaneous impairment of cardiac contractility and increased susceptibility to heart failure. These heart defects were reversed via inhibition of HDACs, suggesting a mechanistic link to epigenetic chromatin remodeling. In humans, we show that a common NOT3 SNP correlates with altered cardiac QT intervals, a known cause of potentially lethal ventricular tachyarrhythmias. Thus, our functional genome-wide screen in Drosophila can identify candidates that directly translate into conserved mammalian genes involved in heart function.

SUBMITTER: Neely GG

PROVIDER: S-EPMC2855221 | biostudies-literature | 2010 Apr

REPOSITORIES: biostudies-literature

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A global in vivo Drosophila RNAi screen identifies NOT3 as a conserved regulator of heart function.

Neely G Gregory GG Kuba Keiji K Cammarato Anthony A Isobe Kazuya K Amann Sabine S Zhang Liyong L Murata Mitsushige M Elmén Lisa L Gupta Vaijayanti V Arora Suchir S Sarangi Rinku R Dan Debasis D Fujisawa Susumu S Usami Takako T Xia Cui-ping CP Keene Alex C AC Alayari Nakissa N NN Yamakawa Hiroyuki H Elling Ulrich U Berger Christian C Novatchkova Maria M Koglgruber Rubina R Fukuda Keiichi K Nishina Hiroshi H Isobe Mitsuaki M Pospisilik J Andrew JA Imai Yumiko Y Pfeufer Arne A Hicks Andrew A AA Pramstaller Peter P PP Subramaniam Sai S Kimura Akinori A Ocorr Karen K Bodmer Rolf R Penninger Josef M JM

Cell 20100401 1

Heart diseases are the most common causes of morbidity and death in humans. Using cardiac-specific RNAi-silencing in Drosophila, we knocked down 7061 evolutionarily conserved genes under conditions of stress. We present a first global roadmap of pathways potentially playing conserved roles in the cardiovascular system. One critical pathway identified was the CCR4-Not complex implicated in transcriptional and posttranscriptional regulatory mechanisms. Silencing of CCR4-Not components in adult Dro ...[more]

PMID: 20371351

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Project description:Glioblastoma multiforme is the most common and most aggressive type of primary brain tumor. The brain-infiltrative character of glioblastoma makes complete surgical removal of the tumor impossible and neither radiation nor current chemotherapy provide cure. Recent evidence shows that glioblastoma multiforme consists of heterogeneous cell populations which differ in tumor-forming potential. Enriched tumor-initiating capacity has been linked to poorly differentiated glioblastoma cells sharing features with neural stem cells. Thus, these cells are important targets for new therapeutic strategies. We aim to identify novel targets controlling maintenance and differentiation in glioblastoma-initiating cells through high throughput screening. To this end, we utilized libraries of small chemical compounds and small interference RNAs in combination with automated imaging and data analysis. Patient-derived glioblastoma cells were expanded and characterized using neural stem cell conditions. In culture, the cells showed low differentiation but expression of neural stem cell markers such as Nestin and Sox2. Upon intracranial injection into SCID mice these cells gave rise to tumors displaying the hallmarks of the human disease. Differentiation of glioblastoma-initiating cells (for example elicited through bone morphogenetic protein, BMP) was associated with strong morphological changes. Hence, cellular morphology, as well as markers specific for differentiation or death were used as screen readout. Lentiviral RNA interference-based screening yielded several gene knockdowns leading to ‘forced’ differentiation of glioblastoma-initiating cells. For example, knockdown of TRRAP (transformation/transcription domain associated protein) led to strongly increased differentiation and loss of proliferative and self-renewing capacity in these cells. TRRAP is an adapter protein implicated in oncogenic transformation through c-MYC transcription activation, also participating in chromatin remodeling and DNA repair. Glioblastoma-initiating cells with reduced TRRAP displayed increased apoptosis upon treatment with the genotoxic agent temozolomide. In vivo, Trapp knockdown cells were not able to give rise to glioblastoma upon transplantation into the brain of SCID mice. Together, these findings support a crucial role for TRRAP in maintenance and tumorigenicity of glioblastoma-initiating cells and might offer future therapeutic options. Two treatments compared to control: two different shRNA sequences for TRRAP were compared to a control shRNA sequence in their effects on global transcription in brain tumor initiating cells

Dataset Information

A global in vivo Drosophila RNAi screen identifies NOT3 as a conserved regulator of heart function.

Publications

A global in vivo Drosophila RNAi screen identifies NOT3 as a conserved regulator of heart function.

Similar Datasets

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