hide
Free keywords:
-
Abstract:
Genetic robustness can be achieved via several mechanisms including transcriptional adaptation (TA), a sequence similarity-driven process whereby mutant mRNA degradation products modulate, directly or indirectly, the expression of so-called adapting genes. To identify the sequences required for this process, we utilized a transgenic approach in Caenorhabditis elegans, combining an overexpression construct for a mutant gene (act-5) and a fluorescent reporter for the corresponding adapting gene (act-3). Analyzing a series of modifications for each construct, we identified, in the 5' regulatory region of the act-3 locus, a 25-base pair (bp) element which exhibits 60% identity with a sequence in the act-5 mRNA and which, in the context of a minimal promoter, is sufficient to induce ectopic expression of the fluorescent reporter. The 25 nucleotide (nt) element in the act-5 mRNA lies between the premature termination codon (PTC) and the next exon/exon junction, suggesting the importance of this region of the mutant mRNA for TA. Additionally, we found that single-stranded RNA injections of this 25 nt element from act-5 into the intestine of wild-type larvae led to higher levels of adapting gene (act-3) mRNA. Different models have been proposed to underlie the modulation of gene expression during TA including chromatin remodeling, the inhibition of antisense RNAs, the release of transcriptional pausing, and the suppression of premature transcription termination, and our data clearly show the importance of the regulatory region of the adapting gene in this particular act-5/act-3 TA model. Our findings also suggest that RNA fragments can modulate the expression of loci exhibiting limited sequence similarity, possibly a critical observation when designing RNA based therapies.
Author summaryTranscriptional adaptation (TA) is a recently identified process that modulates the transcriptome of a cell in response to a mutation in a gene, independent of the mutation's effect on the encoded protein. Depending on the identity of the gene(s) whose expression is modulated, TA can lead to functional compensation or a worse phenotype. TA requires mutant mRNA degradation, and the current hypothesis is that the resulting RNA fragments, or their derivatives, modulate the expression of the so-called adapting genes. A major unresolved question about TA is how the mutant and adapting gene(s) are linked, and it has been proposed that sequence similarity plays an important role. Here, we link a short DNA sequence in the regulatory region of an adapting gene (act-3) to a corresponding sequence in the mutant gene (act-5), and also show that small RNAs containing this act-5 sequence element can lead to higher levels of adapting gene (act-3) mRNA. Because the sequence similarity between these two elements is lower than expected, these results are significant as they suggest that the number of potential adapting genes for a given mutant gene is currently underestimated.
