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Transfer RNA Fragments (tRFs): a Novel Class of Non‐micro Short RNAs that Uses Ago1, 3 and 4 to Repress Specific Target RNAs Through 5′ Seed Sequences
Author(s) -
Dutta Anindya,
Kumar Pankaj,
Kiran Manjari,
Kuscu Canan
Publication year - 2016
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.30.1_supplement.1054.5
Subject(s) - rna , biology , dicer , microrna , small rna , transfer rna , genetics , non coding rna , gene , small interfering rna , microbiology and biotechnology
tRFs, 14–32 nt long single‐stranded RNA derived from mature or precursor tRNAs, are a recently discovered class of small RNA present at read counts comparable to miRNAs. The tRFs are precisely generated fragments present in all human cell lines, mice, flies, worms, yeasts and even some bacteria and originate from the 5′ end (tRF‐5) or 3′ end (tRF‐3) of mature tRNAs or from 3′ trailer sequences of primary tRNA transcripts (tRF‐1). Genes involved in generating canonical miRNAs or siRNAs (Dicer or DGCR8) are dispensable for tRF generation. tRF‐1s and tRF‐3s are more abundant in the cytoplasm than the nucleus, but tRF‐5s are enriched in the nucleus. Human Ago PAR‐CLIP data show that tRF‐5s and tRF‐3s associate with Ago‐1, ‐3, and ‐4 rather than Ago‐2 (unlike microRNAs), raising intriguing questions about how these single‐stranded RNA fragments are loaded on to Ago complexes and how the selectivity is determined for Ago‐1, ‐3 and ‐4 versus Ago‐2. tRF‐1s are not associated with Ago proteins. The locations of the U to C mutation caused by the cross‐linking of the thio‐uracil in the tRF or the target RNA to the Ago protein demonstrate that a 5′ seed sequence of 6–7 bases in tRF‐5 or ‐3 is used to recognize the target RNA in exactly the same way as used by a microRNA to recognize its target RNA. Human Ago‐1 CLASH data identify tens of thousands of chimeric tRF‐target molecules produced by ligation of specific tRFs to a paired target RNA in the Ago‐1 protein isolated from 293T cells. Surprisingly, tRF‐target chimeras are 2–3 fold more abundant than microRNA‐target chimeras suggesting that more tRFs than microRNAs are paired with targets in the Ago‐1 complex. The chimeras identify hundreds of tRF targets and demonstrate that tRF‐5s and tRF‐3s in the Ago‐1 complex use complementarity to their 5′ seed sequences to recognize the target RNAs. Expression of specific tRNAs predicted to produce the same tRF shows that not all tRNAs are equally proficient in producing a given tRF. tRF‐3s produced from transfected tRNAs load into Ago complexes and specifically repress reporter genes with complementarity to the tRFs in their 3′ UTRs. Mutations of the target sites in the reporter or in the generating tRNA show that complementarity to the seed sequence of the tRF is critical for repression. In conclusion, tRF‐5s and ‐3s are non‐micro‐short RNAs produced from specific tRNAs by Dicer‐independent pathways to regulate gene expression. Support or Funding Information NIH P01 CA104106