The in vivo dissection of direct RFX-target gene promoters in C. elegans reveals a novel cis-regulatory element, the C-box
ABSTRACT At the core of the primary transcriptional network regulating ciliary gene expression in Caenorhabditis elegans sensory neurons is the RFX/DAF-19 transcription factor, which binds and thereby positively regulates 13-15 bp X-box promoter motifs found in the cis-regulatory regions of many ciliary genes. However, the variable expression of direct RFX-target genes in various sets of ciliated sensory neurons (CSNs) occurs through as of yet uncharacterized mechanisms. In this study the cis-regulatory regions of 41 direct RFX-target genes are compared using in vivo genetic analyses and computational comparisons of orthologous nematode sequences. We find that neither the proximity to the translational start site nor the exact sequence composition of the X-box promoter motif of the respective ciliary gene can explain the variation in expression patterns observed among different direct RFX-target genes. Instead, a novel enhancer element appears to co-regulate ciliary genes in a DAF-19 dependent manner. This cytosine- and thymidine-rich sequence, the C-box, was found in the cis-regulatory regions in close proximity to the respective X-box motif for 84% of the most broadly expressed direct RFX-target genes sampled in this study. Molecular characterization confirmed that these 8-11 bp C-box sequences act as strong enhancer elements for direct RFX-target genes. An artificial promoter containing only an X-box promoter motif and two of the C-box enhancer elements was able to drive strong expression of a GFP reporter construct in many C. elegans CSNs. These data provide a much-improved understanding of how direct RFX-target genes are differentially regulated in C. elegans and will provide a molecular model for uncovering the transcriptional network mediating ciliary gene expression in animals.
SourceAvailable from: Jin G Park[Show abstract] [Hide abstract]
ABSTRACT: Background Tissue-specific RNA plasticity broadly impacts the development, tissue identity and adaptability of all organisms, but changes in composition, expression levels and its impact on gene regulation in different somatic tissues are largely unknown. Here we developed a new method, polyA-tagging and sequencing (PAT-Seq) to isolate high-quality tissue-specific mRNA from C. elegans intestine, pharynx, and body muscle tissues and study changes in their tissue-specific transcriptomes, and 3¿UTRomes.ResultsWe have identified thousands of novel genes and isoforms differentially expressed between these three tissues. The intestine transcriptome is expansive, expressing over 30% of C. elegans mRNAs, while muscle transcriptomes are smaller but contain characteristic unique gene signatures. Active promoter regions in all three tissues reveal both known and novel enriched tissue-specific elements, along with putative transcription factors, suggesting novel tissue-specific modes of transcription initiation. We have precisely mapped ~20,000 tissue-specific polyadenylation sites, and discovered that ~30% of transcripts in somatic cells use alternative polyadenylation in a tissue-specific manner, with their 3¿UTR isoforms significantly enriched with microRNA targets.ConclusionsPAT-Seq allowed for the first time to directly study tissue specific gene expression changes in an in vivo setting and compare these changes between three somatic tissues from the same organism at single-base resolution within the same experiment. We pinpoint precise tissue-specific transcriptome rearrangements and for the first time link tissue-specific alternative polyadenylation to miRNA regulation, suggesting novel and unexplored tissue-specific post-transcriptional regulatory networks in somatic cells.BMC Biology 01/2015; 13(1):4. DOI:10.1186/s12915-015-0116-6 · 7.43 Impact Factor
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ABSTRACT: Cilia play many essential roles in fluid transport and cellular locomotion, and as sensory hubs for a variety of signal transduction pathways. Despite having a conserved basic morphology, cilia vary extensively in their shapes and sizes, ultrastructural details, numbers per cell, motility patterns and sensory capabilities. Emerging evidence indicates that this diversity, which is intimately linked to the different functions that cilia perform, is in large part programmed at the transcriptional level. Here, we review our understanding of the transcriptional control of ciliary biogenesis, highlighting the activities of FOXJ1 and the RFX family of transcriptional regulators. In addition, we examine how a number of signaling pathways, and lineage and cell fate determinants can induce and modulate ciliogenic programs to bring about the differentiation of distinct cilia types.Development 04/2014; 141(7):1427-1441. DOI:10.1242/dev.074666 · 6.27 Impact Factor
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ABSTRACT: Cilia are found in many eukaryotic species and share a common microtubule architecture that can nonetheless show very diverse features within one animal. The genesis of cilia and their diversity require the expression of different specific genes. At least two classes of transcription factors are involved in ciliogenesis: the RFX family, essential for the assembly of most cilia and the FOXJ1 transcription factors that are key regulators of motile cilia assembly. These two different families of transcription factors have both specific and common target genes and they can also cooperate for the formation of cilia. In collaboration with cell type specific factors, they also contribute to the specialisation of cilia. As a consequence, the identification of RFX and FOXJ1 target genes has emerged as an efficient strategy to identify novel ciliary genes, and in particular genes potentially implicated in ciliopathies.Medecine sciences: M/S 11/2014; 30(11):968-75. DOI:10.1051/medsci/20143011010 · 0.52 Impact Factor