Talk abstracts
Talk on Friday 04:15-04:30pm submitted by Christina Akirtava
Unraveling the roles of 5’ transcript leaders in gene regulation
Christina Akirtava (Carnegie Mellon University, Biological Sciences), Hunter Kready (Carnegie Mellon University, Biological Sciences), Lauren Nazzaro (Carnegie Mellon University, Biological Sciences), Matt Agar-Johnson (Carnegie Mellon University, Biological Sciences), Gemma E May (Carnegie Mellon University, Biological Sciences), C. Joel McManus (Carnegie Mellon University, Biological Sciences)
Abstract:
Translation initiation is regulated by sequences surrounding the start codon, the “Kozak context” (1), cis.-acting sequences and structures in the 5’ transcript leader (TL), and corresponding trans.-acting factors. Previous work evaluating the in vitro. translation efficiency of 96 native yeast TLs showed that changes of 50-200nt in TL lengths varied translation up to 100-fold (2). Larger-scale in vivo. reporter studies of fixed-length synthetic TLs identified upstream AUGs as major repressors of gene expression (3-5). However, the in vivo. regulation of translation by native yeast TLs has not been systematically studied. To investigate cis.-regulatory translational control by native TLs in vivo., we assayed gene expression from ~11,000 endogenous TLs of S. cerevisiae. and S. paradoxus. using Fluorescence-Activated Cell Sorting and high-throughput sequencing (FACS-seq) (6). Additionally, we tested all Kozak variants surrounding AUG start codons in S. cerevisiae. We find Kozak context influences expression over a ~20-fold range, with the expected strong preference for -3 A. Our results show that alternative transcription start sites that change leader length by as little as 10 nucleotides can impact gene expression as much as ~16 fold. Finally, we trained a machine learning model on native 5’ TLs to identify regulatory features that increase sequence-based predictions of translation. Although Kozak strengths explain much of the variance in expression, our model quantitates the influence of upstream open reading frames, mRNA folding around the 5’ cap, and other structures that regulate the rate of translation initiation in vivo.. Thus, our results identify the range and relative regulatory impacts of Kozak context and other cis.-acting sequences and structures on translation from native yeast TLs in vivo.
References:
1)Kozak, M. (1984). Compilation and analysis of sequences upstream… NAR, 12(2), 857–872.
2)Rojas-duran, M. F., & Gilbert, W. V. (2012). Alternative transcription start site selection leads to large differences in translation activity in yeast, 2299–2305.
3)Cuperus et al., (2017). Deep Learning Of The Regulatory Grammar Of Yeast 5′ Untranslated Regions From 500,000 Random Sequences. BioRxiv, 163(2), 1–10.
4)Dvir et al.,(2013). Deciphering the rules by which 5’-UTR sequences… PNAS USA, 110(30), E2792–E2801
5)Sample et al., (2019). Human 5′ UTR design and variant effect prediction from a massively parallel translation assay. Nat. Bio., 37(7), 803–809.
6)Noderer et al., (2014). Quantitative analysis of mammalian translation initiation sites by FACS-seq. Molecular Systems Bio., 10(8), 748–748.
Keywords: translation, modeling, UTR