The integrity of the U12 snRNA 3′ stem–loop is necessary for its overall stability

Disruption of minor spliceosome functions underlies several genetic diseases with mutations in the minor spliceosome-specific small nuclear RNAs (snRNAs) and proteins. Here, we define the molecular outcome of the U12 snRNA mutation (84C>U) resulting in an early-onset form of cerebellar ataxia. To understand the molecular consequences of the U12 snRNA mutation, we created cell lines harboring the 84C>T mutation in the U12 snRNA gene (RNU12). We show that the 84C>U mutation leads to accelerated decay of the snRNA, resulting in significantly reduced steady-state U12 snRNA levels. Additionally, the mutation leads to accumulation of 3′-truncated forms of U12 snRNA, which have undergone the cytoplasmic steps of snRNP biogenesis. Our data suggests that the 84C>U-mutant snRNA is targeted for decay following reimport into the nucleus, and that the U12 snRNA fragments are decay intermediates that result from the stalling of a 3′-to-5′ exonuclease. Finally, we show that several other single-nucleotide variants in the 3′ stem-loop of U12 snRNA that are segregating in the human population are also highly destabilizing. This suggests that the 3′ stem-loop is important for the overall stability of the U12 snRNA and that additional disease-causing mutations are likely to exist in this region.

U1 snDNA chromosomal mapping in ten spittlebug species (Cercopidade, Auchenorrhyncha, Hemiptera)

Spittlebugs, which belong to the family Cercopidae (Auchenorrhyncha, Hemiptera), form a large group of xylem-feeding insects that are best known for causing damage to plantations and pasture grasses. The holocentric chromosomes of these insects remain poorly studied in regards to the organization of different classes of repetitive DNA. To improve chromosomal maps based on repetitive DNAs and to better understand the chromosomal organization and evolutionary dynamics of multigene families in spittlebugs, we physically mapped the U1 snRNA gene with fluorescence in situ hybridization (FISH) in 10 species of Cercopidae belonging to three different genera. All the U1 snDNA clusters were autosomal and located in interstitial position. In seven species, they were restricted to one autosome per haploid genome, while three species of the genus Mahanarva showed two clusters in two different autosomes. Although it was not possible to precisely define the ancestral location of this gene, it was possible to observe the presence of at least one cluster located in a small bivalent in all karyotypes. The karyotype stability observed in Cercopidae is also observed in respect to the distribution of U1 snDNA. Our data are discussed in light of possible mechanisms for U1 snDNA conservation and compared with the available data from other species.