The Role of Ultraconserved Non-Coding Elements in Mammalian Genomes

Ultraconserved elements are stretches of DNA that are 100% identical in mouse, rat, and human genomes. In order to qualify as an ultraconserved element, the length has to be greater than 200 bp. This eliminates most sequences that might be identical by chance.

The most interesting elements are those that fall outside of coding regions. These ultraconservative elements are most likely to be involved in regulating gene expression or some other essential feature of non-coding DNA. The fact that they are identical in species who last shared a common ancestor 100 million years ago is powerful evidence of adaptation.

Ahitiv et al. (2007) set out to test this hypothesis by selecting four examples of ultraconservative elements for further analysis. They discovered that the elements function as tissue specific enhancers in a test designed to look at how they control expression of a maker gene in mouse embryos. The results are shown in Figure 1 (left) of their paper, which was just published in the open access journal PLoS Biology.

The figure shows the genomic location of the four ultraconserved elements; uc248 (222 bp), uc329 (307 bp), uc467 (731 bp), and uc482 (295 bp).

Of these, uc467 is the most remarkable because it is 731 bp in length and resides in the last intron of the DNA polymerase alpha 1 gene (POLA1) on the human X chromosome. The enhancer trap experiment shows that this segment of conserved DNA directs expression of the marker gene in embryonic brain cells (shown as the dark blue area in the embryo above the 467 site). This is usually taken as evidence of specific regulatory sequences that bind transcription factors.

Ahituv et al. then deleted the four ultraconserved sequences from the mouse genome using standard knockout technology. Mice that were homozygous for the knockouts showed no evidence of any defect compared to wild-type mice. In other words, the ultraconserved elements seemed to be completely dispensable—a result that is not consistent with their extreme conservation.

THEME:
Junk DNA
What are the possible explanations? It's possible that the authors missed a phenotype that can only be detected outside the laboratory. It's also possible that the sequences really aren't conserved because they perform an important function but for another reason. Here's how the authors explain their results,

Based on the compelling evidence that ultraconserved elements are conserved due to functional constraint, it has been proposed that their removal in vivo would lead to a significant phenotypic impact [7,8]. Accordingly, our results were unexpected. It is possible that our assays were not able to detect dramatic phenotypes that under a different setting, for instance, outside the controlled laboratory setting, would become evident. Moreover, possible phenotypes might become evident only on a longer timescale, such as longer generation time. It is also possible that subtler genetic manipulations of the ultraconserved elements might lead to an evident phenotype due to a gain-of-function-type mechanism. All four elements examined in this study demonstrated in vivo enhancer activity when tested in a transgenic mouse assay (Figure 1) [6], which would suggest regulatory element redundancy as another possible explanation for the lack of a significant impact following the removal of these specific elements. Just as gene redundancy has been shown to be responsible for the lack of phenotypes associated with many seemingly vital gene knockouts, regulatory sequence redundancy [22] can similarly provide a possible explanation for the lack of a marked phenotype in this study. While our studies have not defined a specific need for the extreme sequence constraints of noncoding ultraconserved elements, they have ruled out the hypothesis that these constraints reflect crucial functions required for viability.

[UPDATE: Ryan Gregory at Genomicron discusses the same paper with a more thorough coverage of the background information and the relevance to junk DNA (Ultraconserved non-coding regions must be functional... right?). R. Ford Denison at This Week in Evolution has some thoughts on the paper (If it's junk, can we get rid of it?")]

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Ahituv, N,, Zhu, Y., Visel, A., Holt, A., Afzal, V., Pennacchio, L.A., and Rubin, E.M. (2007) Deletion of Ultraconserved Elements Yields Viable Mice. PLoS Biol 5(9): e234 doi:10.1371/journal.pbio.0050234.