Abstract
BackgroundMature miRNAs can often be classified into large families, consisting of members with identical seeds (nucleotides 2 through 7 of the mature miRNAs) and highly homologous ∼21-nucleotide (nt) mature miRNA sequences. However, it is unclear whether members of a miRNA gene family, which encode identical or nearly identical mature miRNAs, are functionally interchangeable in vivo.Methods and FindingsWe show that mir-181a-1, but not mir-181c, can promote CD4 and CD8 double-positive (DP) T cell development when ectopically expressed in thymic progenitor cells. The distinct activities of mir-181a-1 and mir-181c are largely determined by their unique pre-miRNA loop nucleotides—not by the one-nucleotide difference in their mature miRNA sequences. Moreover, the activity of mir-181a-1 on DP cell development can be quantitatively influenced by nucleotide changes in its pre-miRNA loop region. We find that both the strength and the functional specificity of miRNA genes can be controlled by the pre-miRNA loop nucleotides. Intriguingly, we note that mutations in the pre-miRNA loop regions affect pre-miRNA and mature miRNA processing, but find no consistent correlation between the effects of pre-miRNA loop mutations on the levels of mature miRNAs and the activities of the mir-181a-1/c genes.ConclusionsThese results demonstrate that pre-miRNA loop nucleotides play a critical role in controlling the activity of miRNA genes and that members of the same miRNA gene families could have evolved to achieve different activities via alterations in their pre-miRNA loop sequences, while maintaining identical or nearly identical mature miRNA sequences.
Highlights
A large number of animal miRNAs have been identified and the genes encoding many of these small RNAs have been shown to play diverse functional roles in animals [1]
These results demonstrate that pre-miRNA loop nucleotides play a critical role in controlling the activity of miRNA genes and that members of the same miRNA gene families could have evolved to achieve different activities via alterations in their pre-miRNA loop sequences, while maintaining identical or nearly identical mature miRNA sequences
We used T cell development as a functional readout to determine the nucleotides and structural domains that are required for the function of mir-181 genes
Summary
A large number of animal miRNAs have been identified and the genes encoding many of these small RNAs have been shown to play diverse functional roles in animals [1]. Each miRNA gene produces at least three small RNA species, including a long primary miRNA transcript (pri-miRNA), an intermediate ,60-nt precursor miRNA (pre-miRNA), and a ,21-nt mature miRNA, through sequential endonucleolytic maturation steps [2]. Mature miRNAs can often be classified into large families, consisting of members with identical seeds (nucleotides 2 through 7 of the mature miRNAs) and highly homologous ,21-nucleotide (nt) mature miRNA sequences It is unclear whether members of a miRNA gene family, which encode identical or nearly identical mature miRNAs, are functionally interchangeable in vivo
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