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RNA localization

RNA localization. mRNA can be localized to subcellular compartments by actin or tubulin -dependent processes. Examples :. Xenopus : Vg1 mRNA (TGF b ) to vegetal pole. Drosophila : nanos, oskar mRNA (posterior) and bicoid (anterior). (requires mRNA binding protein staufen).

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RNA localization

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  1. RNA localization mRNA can be localized to subcellular compartments by actin or tubulin-dependent processes Examples: Xenopus: Vg1 mRNA (TGFb) to vegetal pole Drosophila: nanos, oskar mRNA (posterior) and bicoid (anterior) (requires mRNA binding protein staufen) prospero (into ganglion of mother cells; neuroblast TF) (requires staufen and miranda) Yeast: Ash1 mRNA to daughter cell

  2. lamellipodia staining perinuclear staining in myotubes

  3. Bertrand et al., Mol Cell (98) 2:437-445

  4. SUMMARY 2 I. mRNA decay - regulated and non-regulated turn-over - ordered pathway (deadenylation, decapping, exonucleolytic degradation) - NMD: recognition of premature stop codons II. Cytoplasmic mRNA localization - ZIP code in 3’ UTR - both actin and tubulin-mediated - yeast mating type switch as a model: Ash1 mRNA localization (via 3’ UTR, She2/3, Myo4 and actin cables)

  5. ER translocation & vesicular transport

  6. 3min 7min 37min 117min from: Jamieson and Palade

  7. In vitro reconstitution of ER translocation: - Sec61 complex: conserved translocation channel Sec61 subunits (a, b, g) Sec62/63 TRAM (translocating chain-assoc. membrane protein) - phospholipids (proteoliposomes) and luminal chaperones (BIP) - SRP/SRP receptor only required for co-translational translocation not for post-translational translocation (e.g pre-pro-alpha factor). - energetics of translocation: protein conducting channel (cotranslational) molecular ratcheting (posttranslational)

  8. Probing of translocation intermediates with fluorescent peptides From: Liao and Johnson Cell (97)

  9. The Sec61 complex forms a channel Menetret et al. Mol Cell (2000)6:1219

  10. From: Beckmann et al. Cell (2001)Vol 107, 361-372

  11. From: Beckmann et al. Cell (2001)Vol 107, 361-372

  12. From: Van den Berg et al. Nature (2004)427, 36-44

  13. From: Van den Berg et al. Nature (2004)427, 36-44

  14. Topology of membrane-spanning proteins

  15. Type I membrane proteins have a cleavable signal sequence

  16. Type II membrane proteins have internal signal sequence

  17. Type III membrane proteins have internal signal sequence

  18. Type II+III membrane proteins have internal signal sequences

  19. From: Beckmann et al. Cell (2001)Vol 107, 361-372

  20. Translocation of proteins with multiple membrane spanning domains

  21. From: Van den Berg et al. Nature (2004)427, 36-44

  22. Formation of a glycosylphosphatidylinositol (GPI)-anchor

  23. ER function - Proper folding of proteins (chaperones, lectins, petidyl-prolyl-isomerases) - Formation of disulfide bonds (PDI) GSH prevents oxidation in cytosol GS-SG + NADPH + H+<=> 2 GSH + NADP+ - Proteolytic cleavages - Addition & processing of carbohydrates - Assembly into multimeric proteins - Ca2+ storage - Lipid synthesis - Detoxification (liver!)

  24. Folding of Influenza hemagglutinin (HA)

  25. Ser/Thr

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