Novel intramolecular base-pairing of the U8 snoRNA underlies a Mendelian form of cerebral small vessel disease

How mutations in the non-coding U8 snoRNA cause the neurological disorder leukoencephalopathy with calcification and cysts (LCC) is poorly understood. We report the first vertebrate mutant U8 animal model for interrogating LCC-associated pathology. Mutant U8 zebrafish exhibit defective central nervous system development and ribosomal RNA (rRNA) biogenesis, with tp53 activation which monitors ribosome biogenesis. Importantly, LCC patient fibroblasts demonstrate rRNA processing defects. Human precursor-U8 (pre-U8) containing a 3’ extension rescued mutant U8 zebrafish, indicating conserved biological function. Analysis of LCC-associated U8 alleles in zebrafish revealed that one null and one hypomorphic, but still functional, allele combine to cause LCC. Mutations involving any one of seven nucleotides within the human pre-U8 3’ extension, or 5’ region of U8, alter processing of pre-U8, and identify a novel base-pairing interaction between the 5’ end and 3’ extension of human pre-U8. Variants in these seven nucleotides, one of which is present on a single allele in almost all patients, act as hypomorphic mutations. Given that biallelic null U8 alleles are likely incompatible with human development, identification of hypomorphic mutations mediating viable embryogenesis furthers understanding of LCC molecular pathology and cerebral vascular homeostasis.

Multiple decay events target HAC1 mRNA during splicing to regulate the unfolded protein response

ABSTRACTIn the unfolded protein response (UPR), protein-folding stress in the endoplasmic reticulum (ER) activates a large transcriptional program to increase ER folding capacity. During the budding yeast UPR, the trans-ER-membrane kinase-endoribonuclease Ire1 excises an intron from the HAC1 mRNA and the exon cleavage products are ligated and translated to a transcription factor that induces hundreds of stress-response genes. HAC1 cleavage by Ire1 is thought to be the rate limiting step of its processing. Using cells with mutations in RNA repair and decay enzymes, we show that phosphorylation of two different HAC1 splicing intermediates by Trl1 RNA 5′-kinase is required for their degradation by the 5′→3′ exonuclease Xrn1 to enact opposing effects on the UPR. Kinase-mediated decay (KMD) of cleaved HAC1 3′-exon competes with its ligation to limit productive splicing and suppress the UPR, whereas KMD of the excised intron activates HAC1 translation, likely by relieving an inhibitory base-pairing interaction between the intron and 5′-untranslated region. We also found that ligated but 2′-phosphorylated HAC1 mRNA is endonucleolytically cleaved, yielding a KMD intermediate with both 5′- and 2′-phosphates at its 5′-end that inhibit 5′→3′ decay, and suggesting that Ire1 initiates the degradation of incompletely processed HAC1s to proofread ligation or attenuate the UPR. These multiple decay events expand the scope of RNA-based regulation in the budding yeast UPR and may have implications for the control of the metazoan UPR by mRNA processing.

Peptide nucleic acid-ionic self-complementary peptide conjugates: highly efficient DNA condensers with specific condensing mechanism

Peptide nucleic acid-ionic self-complementary peptide conjugates can induce efficient DNA condensation via base-pairing interaction and peptide association.