From de Bruijn Graph to Minimal Promoter

Another paper with similar theme as Mike White’s got published today (h/t: @genologos). The beauty of this work is their creative use of de Bruijn graphs to come up with a compact set of nucleotides covering all possible k-mers (k=6) as promoters. Quite surprisingly, they show few bindings of transcription factors leading to GFP expression (27 out of 4096) as opposed to ubiquitous binding in Mike White’s paper. We do not know where exactly the difference comes from, and requested Mike to help. BTW, where do people download the supplements from for Genome Biology papers? We do not see any link for supplementary materials on the sidebar.

[Edit. There is no conflict between Mike White’s finding and this one. Firstly, the ratio 27 out of 4096 stated above is wrong and should be 27/184 as pointed out by Juan in comment section. Secondly, even after that we are not sure whether that 27/184 is a fair comparison with the other paper, because total length of promoter is different as well as the content. Finally, this paper does GFP measurement, whereas Mike measured gene expression directly.]

Published in Nature Genome Biology

Background

Large-scale annotation efforts have improved our ability to coarsely predict regulatory elements throughout vertebrate genomes. However, it is unclear how complex spatiotemporal patterns of gene expression driven by these elements emerge from the activity of short, transcription factor binding sequences.

Results

We describe a comprehensive promoter extension assay in which the regulatory potential of all 6 base-pair (bp) sequences was tested in the context of a minimal promoter. To enable this large-scale screen, we developed algorithms that use a reverse-complement aware decomposition of the de Bruijn graph to design a library of DNA oligomers incorporating every 6-bp sequence exactly once. Our library multiplexes all 4,096 unique 6-mers into 184 double-stranded 15-bp oligomers, which is sufficiently compact for in vivo testing. We injected each multiplexed construct into zebrafish embryos and scored GFP expression in 15 tissues at two developmental time points. Twenty-seven constructs produced consistent expression patterns, with the majority doing so in only one tissue. Functional sequences are enriched near biologically relevant genes, match motifs for developmental transcription factors, and are required for enhancer activity. By concatenating tissue-specific functional sequences, we generated completely synthetic enhancers for the notochord, epidermis, spinal cord, forebrain and otic lateral line, and show that short regulatory sequences do not always function modularly.

Conclusions

This work introduces a unique in vivo catalog of short, functional regulatory sequences and demonstrates several important principles of regulatory element organization. Furthermore, we provide resources for designing compact, reverse-complement aware k-mer libraries.