Function of chromatin modifiers in cranial neural crest development

University Of Colorado Denver, Aurora CO

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Abstract

? DESCRIPTION (provided by applicant): There is a fundamental gap in our understanding of how defects in chromatin remodeling proteins, methyltransferases and acetyltransferases are causative for human craniofacial phenotypes. This represents an important problem, because craniofacial defects occur frequently in the human population, 1 in every 1000 live births annually in the U.S. (CDC, 2011) and many are associated with epigenetic regulators of the genome. Our long-term goal is to better understand the function of chromatin remodelers during cranial neural crest (cNCC) development. The objective of this application is to determine the mechanism by which two families of epigenetic regulators, KAT2a lysine acetyltransferase and PRDM lysine methyltransferases that regulate each other and act to modify the same H3K9 residue on histone 3, function in zebrafish and mouse cNCC development. We will use two excellent developmental model systems and combine genetic tools with live cell imaging of zebrafish and mouse cNCC behaviors and transcriptional studies to tackle the question of why mutations in Kat2a and Prdms lead to craniofacial abnormalities. The overall hypothesis is that these chromatin modifying enzymes act as opposing transcriptional regulators and function cell autonomously to regulate cNCC proliferation and migration. The rationale for this research is that understanding the mechanism of how KAT2a and PRDMs regulate cNCC development will have the potential to translate into a better understanding of the pathogenesis of craniofacial defects due to mutations in epigenetic regulators, including cleft lip and palate and various syndromes such as Kabuki and SBBYSS that affect the human population. From strong preliminary data, we have designed 3 specific aims: 1) Determine the developmental function of KAT2A and PRDMs in cranial neural crest development, 2) Examine the genetic interaction and regulation of gene targets by KAT2A and PRDMs, and 3) Determine the enzymatic regulation and chromatin state of KAT2A and PRDMs target genes. Under the first aim, we have determined that Prdm1, Prdm3, Prdm16 and Kat2a have craniofacial defects in both mouse and zebrafish. We have the tools and expertise to determine the specific craniofacial defects and to define abnormalities in proliferation and migration of cNCCs. For the second aim, we have generated and obtained most of the zebrafish and mouse strains, and performed transcriptional profiling in both zebrafish and mouse, demonstrating feasibility. For aim three, we have shown analysis of acetylation and methylation states in both tissue and biochemically. Our approach is conceptually innovative in testing a novel hypothesis and technically innovative in the use of live cell imaging and the interplay between two species that model human craniofacial development. The proposed research is significant because it is expected to advance an understanding of how cNCCs form the craniofacial skeleton, which has the potential to inform the treatment of neural crest associated birth defects and craniofacial syndromes.