cytochrome-c-t and angiogenin

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. Next-generation 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.

Topics: Animals; Apoptosis; Apoptosomes; Apoptotic Protease-Activating Factor 1; Base Sequence; Caspase 3; Caspase 9; Cell Survival; Cells, Cultured; Cytochromes c; Electrophoretic Mobility Shift Assay; Fibroblasts; Mice; Mitochondria; Neurons; Osmotic Pressure; Ribonuclease, Pancreatic; Ribonucleoproteins; RNA Cleavage; RNA, Transfer; Sequence Analysis, RNA

Angiogenin is a 14 kDa protein originally identified as an angiogenic protein. Recent development has shown that angiogenin acts on both endothelial cells and neuronal cells. Loss-of-function mutations in the coding region of the ANG gene have recently been identified in patients with amyotrophic lateral sclerosis. Angiogenin has been shown to control motor neuron survival and protect neurons from apoptosis under various stress conditions. In this article, we characterize the anti-apoptotic activity of angiogenin in pluripotent P19 mouse embryonal carcinoma cells. Angiogenin prevents serum withdrawal-induced apoptosis. Angiogenin upregulates anti-apoptotic genes, including Bag1, Bcl-2, Hells, Nf-kappab and Ripk1, and downregulates pro-apoptotic genes, such as Bak1, Tnf, Tnfr, Traf1 and Trp63. Knockdown of Bcl-2 largely abolishes the anti-apoptotic activity of angiogenin, whereas the inhibition of Nf-kappab activity results in a partial, but significant, inhibition of the protective activity of angiogenin. Thus, angiogenin prevents stress-induced cell death through both the Bcl-2 and Nf-kappab pathways.

Topics: Animals; Apoptosis; Carcinoma, Embryonal; Caspases; Culture Media, Serum-Free; Cytochromes c; Gene Expression Regulation, Neoplastic; Mice; NF-kappa B; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-bcl-2; Recombinant Proteins; Reverse Transcriptase Polymerase Chain Reaction; Ribonuclease, Pancreatic; Tumor Cells, Cultured