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Regulated Unproductive Splicing

This study investigates the mechanisms of alternative splicing in human genes and its role in generating isoforms that may be subjected to nonsense-mediated decay (NMD). By analyzing genomic DNA sequences and mRNA transcripts, we find that alternative splicing significantly influences gene expression and protein diversity, with a notable percentage of isoforms containing premature stop codons. The role of key proteins, such as Upf1, Upf2, and Upf3, in the regulation of mRNA stability and translation is also explored, highlighting the complex interplay between splicing and decay pathways.

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Regulated Unproductive Splicing

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  1. Regulated Unproductive Splicing Liana Lareau Brenner Group* *Labs aren’t carpeted. Groups are. Or so I’m told.

  2. Alternative Splicing Genomic DNA Sequence Transcription pre-mRNA Intron Intron Intron Exon Exon Exon Exon Exon RNA Processing Gm AAAAA Gm AAAAA mRNA mRNA

  3. How much alternative splicing occurs in humans?What is its role?

  4. Alternative Isoform Inference

  5. Inferred Isoforms 0 10000 2000 4000 6000 8000 5970 Alternative isoforms 9527 All isoforms, including canonical 3557 genes with alternative spicing, from 12,000 mRNAs

  6. Premature stop codon 2078 (35 % of 5970) Inferred Isoforms 0 10000 2000 4000 6000 8000 5970 Alternative isoforms 9527 All isoforms, including canonical

  7. Nonsense-Mediated mRNA Decay Translated normally ORF Gm mRNA AAAAAAAAA Degraded by NMD ORF Gm mRNA AAAAAAAAA >50 nt

  8. Alternative Splicing Can Yield Isoforms Differentially Subjected to Decay DNA DNA pre-mRNA pre-mRNA mRNA mRNA Nonsense mediated decay Premature termination codon

  9. Eukaryota

  10. Eukaryota human, mouse, fly yeast

  11. Eukaryote sequencing projects 1 3 6 1 16 1 8 5

  12. The Players Upf1aka RENT1 (human), SMG-2 (C. elegans) Binds to downstream Upf2 and Upf3 during translation termination Promotes mRNA decapping and degradation Upf2aka SMG-3 (C. elegans) Binds to Upf3 after nuclear export Upf3aka SMG-4 (C. elegans) Associates with exon junction complex in nucleus

  13. Upf1

  14. Upf2

  15. Upf3

  16. Eukaryota Plasmodium, Cryptosporidium human, mouse, fly, mosquito, worm yeast, S. pombe, Aspergillus, Neurospora

  17. Upf3: Yeast vs. Human BLAST results: Score = 26.9 bits (58), Expect = 9.8 Identities = 30/131 (22%), Positives = 48/131 (36%), Gaps = 16/131 (12%)

  18. transcriptional regulation Gene locus transcription pre-mRNA productive splicing productive mRNA regulated unproductive splicing translation Protein

  19. ORF SC35 pre-mRNA SC35 Auto-regulation Sureau et al. (2001) EMBO J20:1785-96

  20. ORF SC35 pre-mRNA Gm AAAAA SC35 mRNA SC35 Auto-regulation Sureau et al. (2001) EMBO J20:1785-96

  21. ORF SC35 pre-mRNA Gm AAAAA SC35 mRNA SC35 SC35 protein SC35 Auto-regulation Sureau et al. (2001) EMBO J20:1785-96

  22. ORF SC35 pre-mRNA Gm AAAAA SC35 mRNA SC35 SC35 SC35 protein SC35 pre-mRNA SC35 Auto-regulation Sureau et al. (2001) EMBO J20:1785-96

  23. ORF SC35 pre-mRNA Gm AAAAA SC35 mRNA SC35 SC35 SC35 protein SC35 pre-mRNA Gm AAAAA SC35 Auto-regulation SC35 mRNA (with premature termination codon) Sureau et al. (2001) EMBO J20:1785-96

  24. ORF SC35 pre-mRNA Gm AAAAA SC35 mRNA SC35 SC35 SC35 protein SC35 pre-mRNA Gm AAAAA SC35 Auto-regulation SC35 mRNA (with premature termination codon) Sureau et al. (2001) EMBO J20:1785-96

  25. SC35 Auto-regulation SC35 Locus transcription SC35 pre-mRNA alternative splicing splicing Productive SC35 mRNA translation SC35 protein

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