The ability to manipulate neural progenitor cell differentiation may facilitate the treatment of many developmental, acquired and neurodegenerative processes, and understanding neuronal lineage determination may provide valuable insight into the mechanisms underlying neural physiology and disease. Lineage commitment by progenitor cells is influenced by epigenetic signals such as the bone morphogenetic proteins (Bmps) and the Delta/Notch lig and/receptor complexes that activate specific sets of transcriptional activators and repressors. Whereas many studies have focused on the signaling cascades regulating these distinct pathways, not much is known regarding how these pathways may interact to coordinately regulate neural stem cell fates. Recent evidence suggest that Bmp and Notch signaling may both activate members of the Hes (named after the Drosophila homologues Hairy and Enhancer of Split) and Hes-related (Herp) families of transcription factors, and that physical interactions among members of these families influence lineage fate. The protein Herp3 is a member of the Herp family of transcription factors whose function in neuronal development has yet to be elucidated. The goal of this research plan is to explore the role of the Herp3 protein in neuronal cell fate determination in response to Bmp signaling. The primary hypothesis of the proposed research is that Herp3 is involved in Bmp-mediated neuronal differentiation, and that interactions between Herp3 and Hes family members mediate Herp3 function.
The Specific Aims of the proposed project are: 1) to identify the transcriptional targets of Herp3 and its role in neuronogenesis; 2) to analyze the function of the Herp3-Hes interaction; 3) to define the functional importance of Herp3 expression in response to Bmp signaling in vivo. Experimental design and methods to accomplish the first Aim include RNA inhibition (RNAi), transient transfection, and the chromatin precipitation (CHIP) cloning assay to identify target regulatory elements of Herp3 transcriptional regulation.
The second Aim will involve electophoretic mobility shift assays (EMSAs) and transient transfections to evaluate the role of the Hes-Herp3 interaction on DNA binding.
The third Aim will involve in situ hybridization and immunohistochemistry in normal mice and in mice with mutations of the Bmp receptor to examine normal Herp3 expression in vivo and Herp3 expression in response to Bmp signaling.
The third Aim will also involve viral mis-expression of Herp3 and of RNAi molecules of Herp3, to evaluate the in vivo function of the Herp3 protein. The experiments described in this research plan should allow a detailed account of the basic function of the Herp3 protein in neuronal lineage determination in response to Bmp signaling, thereby elucidating the mechanisms by which neuronal differentiation is deranged in neurological disease. ? ?
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| Bassuk, Alexander G; Wallace, Robyn H; Buhr, Aimee et al. (2008) A homozygous mutation in human PRICKLE1 causes an autosomal-recessive progressive myoclonus epilepsy-ataxia syndrome. Am J Hum Genet 83:572-81 |
| Jalali, Ali; Aldinger, Kimberly A; Chary, Ajit et al. (2008) Linkage to chromosome 2q36.1 in autosomal dominant Dandy-Walker malformation with occipital cephalocele and evidence for genetic heterogeneity. Hum Genet 123:237-45 |
