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Cellular and extracellular tRNA-derived fragments demonstrate distinct signatures in cellular stress

ABSTRACT: Cellular and extracellular tRNA-derived fragments demonstrate distinct signatures in cellular stress

PROVIDER: PRJNA727096 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR14416500_1.fastq.gz	Fastqsanger.gz
	SRR14416500_2.fastq.gz	Fastqsanger.gz
	SRR14416501_1.fastq.gz	Fastqsanger.gz
	SRR14416501_2.fastq.gz	Fastqsanger.gz
	SRR14416502_1.fastq.gz	Fastqsanger.gz

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Cellular and extracellular tRNA-derived fragments demonstrate distinct signatures in cellular stress

Project description:The cellular response to stress is an important determinant of disease pathogenesis. Uncovering the molecular fingerprints of distinct stress responses may yield novel biomarkers for different diseases, and potentially identify key signaling pathways important for disease progression . tRNA and tRNA-derived small RNAs (tDRs) comprise one of the most abundant RNA species in cells and have been associated with cellular stress responses. However, systematic characterization of tDRs have been challenged by the technical difficulties in accurately profiling tDRs with normal small RNA sequencing techniques due to the presence of RNA modifications. Here we use AlkB-facilitated methylation sequencing (ARM-seq) to uncover a comprehensive landscape of cellular and extracellular tDRs expression in a variety of human and rat cells during common stress responses, including nutrition deprivation, hypoxia, and oxidative stress. We found that extracellular tDRs have a distinct fragmentation signature with a predominant length of 31-33 nts and a highly specific termination position when compared with intracellular tDRs, and comprise signatures that improve discrimination of different cellular stress responses compared to extracellular miRNAs. Distinct extracellular tDR signatures for each profiled stressor are elucidated in 4 different types of cells. This distinct extracellular tDR fragmentation pattern is also noted in plasma extracellular RNA from patients on cardiopulmonary bypass with significant overlap with the signatures of nutrition depravation and oxidative stress in our cellular models, providing preliminary in vivo correlation of our findings. Future application of our findings to human disease models may have promise in yielding novel biomarkers.

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tRNAArg-derived fragments mediate protein arginylation [tRNA]

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The Lupus autoantigen La prevents mis-channeling of tRNA fragments into the human microRNA pathway [PAR-CLIP]

Project description:The Lupus autoantigen (La) is a single-stranded RNA-binding protein that stabilizes RNA polymerase III (pol III) transcripts and supports RNA folding. In addition, La has been implicated in different steps of the mammalian small RNA pathway. Here, we have analyzed effects of La depletion on the Argonaute (Ago)-bound small RNAs in human cells. We find that in the absence of La, distinct tRNA fragments are loaded into Ago protein complexes and our data suggests that La prevents the production and loading of such tRNA fragments. However, one specific isoleucine tRNA escapes this regulation and produces both a functional tRNA as well as a microRNA (miRNA). We demonstrate that fully complementary 5' leader and 3' trailer of the pre-tRNA-Ile form a double-stranded RNA molecule that has low affinity to La. Instead, Exportin 5 (Xpo5) recognizes it as miRNA precursor and transports it into the cytoplasm for Dicer processing and Ago loading. Thus, La functions as gatekeeper to ensure correct tRNA generation and to protect the miRNA pathway from potentially functional tRNA fragments.

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Project description:Parental dietary conditions can influence the metabolic traits of offspring. In mice, paternal consumption of low protein diet alters cholesterol and lipid metabolism of progeny. Here, we examine RNA species expressed in male reproductive tissues of mice. Protein restriction leads to altered levels of multiple small RNAs in mature sperm, as well as throughout the male reproductive tract, with decreased levels of let-7 family members and increased levels of 5â?? fragments of tRNA-Gly isoacceptors. Intriguingly, tRNA fragments are scarce in the testis, but their levels increase in sperm during posttesticular maturation in the epididymis. We find that epididymosomes â?? extracellular vesicles which fuse with sperm during epididymal transit â?? exhibit RNA payloads closely matching those of mature sperm, and can deliver tRNA fragments to immature sperm in vitro both in mouse and in bull. Finally, we show that tRNA-Gly-GCC fragments play a role in repressing genes associated with the endogenous retroelement MERVL, both in ES cells and in preimplantation embryos. Our results shed light on small RNA biogenesis during post-testicular sperm maturation, and link tRNA fragments to regulation of endogenous retroelements active in the early embryo. E14 mESCs were transfected with LNA-containing oligos antisense to tRF-GG or GFP-esiRNA as control, then either total RNA was isolated or 80S ribosomes were isolated from Rnase-digested whole cell lysate, and footprints between 26-32nt were collected for ribosome profiling library construction using a kit-free protocol based on Heyer et al. 2015 NAR.

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Biogenesis and function of tRNA fragments during sperm maturation and fertilization in mammals (smallRNA-seq)

Project description:Parental dietary conditions can influence the metabolic traits of offspring. In mice, paternal consumption of low protein diet alters cholesterol and lipid metabolism of progeny. Here, we examine RNA species expressed in male reproductive tissues of mice. Protein restriction leads to altered levels of multiple small RNAs in mature sperm, as well as throughout the male reproductive tract, with decreased levels of let-7 family members and increased levels of 5â fragments of tRNA-Gly isoacceptors. Intriguingly, tRNA fragments are scarce in the testis, but their levels increase in sperm during post-testicular maturation in the epididymis. We find that epididymosomes â extracellular vesicles which fuse with sperm during epididymal transit â exhibit RNA payloads closely matching those of mature sperm, and can deliver tRNA fragments to immature sperm in vitro both in mouse and in bull. Finally, we show that tRNA-Gly-GCC fragments play a role in repressing genes associated with the endogenous retroelement MERVL, both in ES cells and in preimplantation embryos. Our results shed light on small RNA biogenesis during post-testicular sperm maturation, and link tRNA fragments to regulation of endogenous retroelements active in the early embryo. Small RNA (<40nt) profile of various tissues and cells was examined by deep sequencing

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