Abstract
One of the central goals of developmental neurobiology is to describe and understand the multi-tiered molecular events that control the progression of a fertilized egg to a terminally differentiated neuron. In the nematode Caenorhabditis elegans, the progression from egg to terminally differentiated neuron has been visually traced by lineage analysis. For example, the two gustatory neurons ASEL and ASER, a bilaterally symmetric neuron pair that is functionally lateralized, are generated from a fertilized egg through an invariant sequence of 11 cellular cleavages that occur stereotypically along specific cleavage planes. Molecular events that occur along this developmental pathway are only superficially understood. We take here an unbiased, genome-wide approach to identify genes that may act at any stage to ensure the correct differentiation of ASEL. Screening a genome-wide RNAi library that knocks-down 18,179 genes (94% of the genome), we identified 245 genes that affect the development of the ASEL neuron, such that the neuron is either not generated, its fate is converted to that of another cell, or cells from other lineage branches now adopt ASEL fate. We analyze in detail two factors that we identify from this screen: (1) the proneural gene hlh-14, which we find to be bilaterally expressed in the ASEL/R lineages despite their asymmetric lineage origins and which we find is required to generate neurons from several lineage branches including the ASE neurons, and (2) the COMPASS histone methyltransferase complex, which we find to be a critical embryonic inducer of ASEL/R asymmetry, acting upstream of the previously identified miRNA lsy-6. Our study represents the first comprehensive, genome-wide analysis of a single neuronal cell fate decision. The results of this analysis provide a starting point for future studies that will eventually lead to a more complete understanding of how individual neuronal cell types are generated from a single-cell embryo.
Highlights
When establishing the nematode C.elegans as a genetic model system, Sydney Brenner’s stated goal was the identification of genes required to build a nervous system [1,2]
We find that dsRNA directed against gfp barely reduces gfp expression levels in transgenic animals that drive a gfp reporter in the ASEL neuron (Figure 2A)
The RNAi screen revealed a number of genes that we would have predicted to result in ASE lineage defects, including genes involved in the segregation of early patterning cues and involved in a re-iteratively used binary fate decision system[7]
Summary
When establishing the nematode C.elegans as a genetic model system, Sydney Brenner’s stated goal was the identification of genes required to build a nervous system [1,2]. Mainly using the classic forward screening methods established by Brenner [1], have revealed genes that are required to generate a diversity of individual neuron types [3]. The advent of RNAi technology in C.elegans has opened an alternative path to classic forward analysis to study the genetic bases of various processes [4,5]. This approach has not been undertaken in C.elegans to comprehensively study neuronal development and fate determination. In this paper we utilize the RNAi approach to screen, at a genome-wide level, for factors involved in all aspects of neuronal development
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