PROJECT SUMMARY Kabuki-like syndrome is a neurodevelopmental disorder caused by mutations in the H3K4 demethylase, LSD1, and is characterized by a range of abnormal somatic phenotypes including intellectual disabilities, craniofacial abnormalities, and developmental delay. Despite their implication in Kabuki-like syndrome, and other neurodevelopmental syndromes, how mutations in histone modifying enzymes like LSD1 contribute to complex developmental phenotypes is unclear. To better understand how mutations in histone modifying enzymes affect somatic development we have developed a C. elegans model lacking SPR-5, the C. elegans ortholog of LSD1, in addition to the histone 3 lysine 9 (H3K9) methyltransferase MET-2. Without SPR-5 and MET-2, progeny ectopically express germline genes in somatic tissues leading to phenotypes that resemble those observed in neurodevelopmental syndromes, including muscle defects and developmental delay. Strikingly, the developmental delay and ectopic expression of candidate germline genes in spr-5; met-2 mutants can be rescued by knocking down the H3K36 methyltransferase, MES-4, which bookmarks germline genes for reexpression in the germline of the next generation. These data suggest that the developmental delay that we observe in the absence of SPR-5 and MET-2 is caused by ectopic expression of germline genes in somatic tissues. Among the MES-4 regulated germline loci that are ectopically expressed in the soma of spr-5; met-2 mutants, we detect PIWI-interacting RNAs, or piRNAs, and the genes that regulate their function. piRNAs are a large class of small non-coding RNAs that regulate development by functioning in several different pathways, including epigenetic programming, germline transcript silencing, RNA turnover, and translational control. Ectopic expression of piRNA genes has recently been implicated in somatic development and diseases, but how ectopic piRNAs function in somatic cells or if they contribute to abnormal developmental phenotypes poorly understood. Thus, our new C. elegans model provides a unique opportunity to molecularly demonstrate how ectopic germline gene and piRNA expression affects normal development. By combining cutting-edge large genomic experiments, classical developmental techniques, and single molecule fluorescent in situ hybridization we will use our C. elegans model in AIM 1 to determine how ectopic germline genes alter somatic gene expression, 2) define tissue specificity of ectopic germline gene expression, and 3) examine how distinct chromatin states contribute to misexpression of germline genes in somatic tissues. In AIM 2, we will employ similar approaches to examine how ectopic expression of piRNAs contributes to tissue specific somatic defects. Together, the proposed aims will provide mechanistic insight into how these ectopic transcriptional events, that occur when histone modifying enzymes are mutated, contribute to phenotypes that overlap with those seen in human neurodevelopmental disorders.

PROJECT NARRATIVE Data from mouse and human suggest a new disease paradigm where ectopic expression of genes and piRNAs when histone methylation is misregulated leads to developmental abnormalities. We have developed a C. elegans model that can be used to elucidate the mechanisms underlying this new disease paradigm. The proposed studies will take advantage of this new C. elegans model in which two histone modifying enzymes are mutated to determine how ectopic gene expression and ectopic piRNAs cause developmental defects.