f Adaptation to changes in extracellular tonicity is essential for cell survival. However, severe or chronic hyperosmotic stress induces apoptosis, which involves cytochrome c (Cyt c) release from mitochondria and subsequent apoptosome formation. Here, we show that angiogenin-induced accumulation of tRNA halves (or tiRNAs) is accompanied by increased survival in hyperosmotically stressed mouse embryonic fibroblasts. Treatment of cells with angiogenin inhibits stress-induced formation of the apoptosome and increases the interaction of small RNAs with released Cyt c in a ribonucleoprotein (Cyt c-RNP) complex. Nextgeneration sequencing of RNA isolated from the Cyt c-RNP complex reveals that 20 tiRNAs are highly enriched in the Cyt c-RNP complex. Preferred components of this complex are 5= and 3= tiRNAs of specific isodecoders within a family of isoacceptors. We also demonstrate that Cyt c binds tiRNAs in vitro, and the pool of Cyt c-interacting RNAs binds tighter than individual tiRNAs. Finally, we show that angiogenin treatment of primary cortical neurons exposed to hyperosmotic stress also decreases apoptosis. Our findings reveal a connection between angiogenin-generated tiRNAs and cell survival in response to hyperosmotic stress and suggest a novel cellular complex involving Cyt c and tiRNAs that inhibits apoptosome formation and activity.

Background: Regulation of stress-induced tRNA cleavage by angiogenin is not well studied. Results: tRNA fragment accumulation was higher during oxidative than hypertonic stress. Conclusion: tRNA cleavage is regulated by the availability of angiogenin and tRNA substrate, levels of RNH1, and the rates of protein synthesis. Significance: Stress-specific tRNA cleavage mechanisms and patterns will provide insights into novel stress signaling pathways.

Background: Protein synthesis control is important for ␤-cell fate during ER stress. Results: Increased protein synthesis during chronic ER stress in ␤-cells involves the transcriptional induction of an amino acid transporter network. Conclusion: Increased amino acid uptake in ␤-cells during ER stress promotes apoptosis. Significance: Induced expression of a network of amino acid transporters in islets can contribute to chronic ER stress-induced diabetes.

The N 1 -methyl-Adenosine (m 1 A58) modification at the conserved nucleotide 58 in the TCC loop is present in most eukaryotic tRNAs. In yeast, m 1 A58 modification is essential for viability because it is required for the stability of the initiator-tRNA Met . However, m 1 A58 modification is not required for the stability of several other tRNAs in yeast. This differential m 1 A58 response for different tRNA species raises the question of whether some tRNAs are hypomodified at A58 in normal cells, and how hypomodification at A58 may affect the stability and function of tRNA. Here, we apply a genomic approach to determine the presence of m 1 A58 hypomodified tRNAs in human cell lines and show how A58 hypomodification affects stability and involvement of tRNAs in translation. Our microarray-based method detects the presence of m

Over 100 chemical types of RNA modifications have been identified in thousands of sites in all three domains of life. Recent data suggest that modifications function synergistically to mediate biological function, and that cells may coordinately modulate modification levels for regulatory purposes. However, this area of RNA biology remains largely unexplored due to the lack of robust, high-throughput methods to quantify the extent of modification at specific sites. Recently, we developed a facile enzymatic ligation-based method for detection and quantitation of methylated 2′-hydroxyl groups within RNA. Here we exploit the principles of molecular recognition and nucleic acid chemistry to establish the experimental parameters for ligation-based detection and quantitation of pseudouridine (Ψ) and N6-methyladenosine (m6A), two abundant modifications in eukaryotic rRNA/tRNA and mRNA, respectively. Detection of pseudouridylation at several sites in the large subunit rRNA derived from yeast demonstrates the feasibility of the approach for analysis of pseudouridylation in biological RNA samples.