Biology

Permanent link for this communityhttps://hdl.handle.net/2022/7910

Browse

Browsing Biology by Subject "3' untranslated region"

Filter results by typing the first few letters
Now showing 1 - 2 of 2
  • Thumbnail Image
    Item
    Global Patterns of Tissue-Specific Alternative Polyadenylation in Drosophila
    (Cell Reports, 2012) Smibert, P.; Miura, P.; Westholm, J.O.; Shenker, S.; May, G.; Duff, M.O.; Zhang, D.; Eads, B.D.; Carlson, J.; Brown, J.B.; Eisman, R.C.; Andrews, J.; Kaufman, T.; Cherbas, P.; Celniker, S.E.; Graveley, B.R.; Lai, E.C.
    We analyzed the usage and consequences of alternative cleavage and polyadenylation (APA) in $\textit{Drosophila melanogaster}$ by using >1 billion reads of stranded mRNA-seq across a variety of dissected tissues. Beyond demonstrating that a majority of fly transcripts are subject to APA, we observed broad trends for 3′ untranslated region (UTR) shortening in the testis and lengthening in the central nervous system (CNS); the latter included hundreds of unannotated extensions ranging up to 18 kb. Extensive northern analyses validated the accumulation of full-length neural extended transcripts, and in situ hybridization indicated their spatial restriction to the CNS. Genes encoding RNA binding proteins (RBPs) and transcription factors were preferentially subject to 3′ UTR extensions. Motif analysis indicated enrichment of miRNA and RBP sites in the neural extensions, and their termini were enriched in canonical cis elements that promote cleavage and polyadenylation. Altogether, we reveal broad tissue-specific patterns of APA in $\textit{Drosophila}$ and transcripts with unprecedented 3′ UTR length in the nervous system.
  • Thumbnail Image
    Item
    Primary and Secondary siRNAs in Geminivirus-induced Gene Silencing
    (Public Library of Science, 2012) Aregger, M.; Borah, B.K.; Seguin, J.; Rajeswaran, R.; Gubaeva, E.G.; Zvereva, A.S.; Windels, D.; Vazquez, F.; Blevins, T.; Farinelli, L.; Pooggin, M.M.
    In plants, RNA silencing-based antiviral defense is mediated by Dicer-like (DCL) proteins producing short interfering (si)RNAs. In $\textit{Arabidopsis}$ infected with the bipartite circular DNA geminivirus $\textit{Cabbage leaf curl virus}$ (CaLCuV), four distinct DCLs produce 21, 22 and 24 nt viral siRNAs. Using deep sequencing and blot hybridization, we found that viral siRNAs of each size-class densely cover the entire viral genome sequences in both polarities, but highly abundant siRNAs correspond primarily to the leftward and rightward transcription units. Double-stranded RNA precursors of viral siRNAs can potentially be generated by host RDR-dependent RNA polymerase (RDR). However, genetic evidence revealed that CaLCuV siRNA biogenesis does not require RDR1, RDR2, or RDR6. By contrast, CaLCuV derivatives engineered to target 30 nt sequences of a $\textit{GFP}$ transgene by primary viral siRNAs trigger RDR6-dependent production of secondary siRNAs. Viral siRNAs targeting upstream of the $\textit{GFP}$ stop codon induce secondary siRNAs almost exclusively from sequences downstream of the target site. Conversely, viral siRNAs targeting the $\textit{GFP}$ 3′-untranslated region (UTR) induce secondary siRNAs mostly upstream of the target site. RDR6-dependent siRNA production is not necessary for robust $\textit{GFP}$ silencing, except when viral siRNAs targeted $\textit{GFP}$ 5′-UTR. Furthermore, viral siRNAs targeting the transgene enhancer region cause $\textit{GFP}$ silencing without secondary siRNA production. We conclude that the majority of viral siRNAs accumulating during geminiviral infection are RDR1/2/6-independent primary siRNAs. Double-stranded RNA precursors of these siRNAs are likely generated by bidirectional readthrough transcription of circular viral DNA by RNA polymerase II. Unlike transgenic mRNA, geminiviral mRNAs appear to be poor templates for RDR-dependent production of secondary siRNAs.