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Potential entry points for loss of function analysis

Potential entry points for loss of function analysis. Potential entry points for loss of function analysis. Potential entry points for loss of function analysis. Dicer inactivation-in c elegans. Dcr1+/- X Dcr1+/-. Dcr-/-. Dicer inactivation-in c elegans. Dcr1+/- X Dcr1+/-. Dcr-/-. WT.

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Potential entry points for loss of function analysis

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  1. Potential entry points for loss of function analysis

  2. Potential entry points for loss of function analysis

  3. Potential entry points for loss of function analysis

  4. Dicer inactivation-in c elegans Dcr1+/- X Dcr1+/- Dcr-/-

  5. Dicer inactivation-in c elegans Dcr1+/- X Dcr1+/- Dcr-/- WT Dcr-/- (Science, 293, 2269)

  6. Dicer inactivation-in c elegans Dcr1+/- X Dcr1+/- Dcr-/- Dcr-/- WT (G & D, 15, 2654)

  7. Dicer inactivation-in c elegans • Dcr1 is required for germ line formation • Dcr1 is not essential for later development • Behaviors? • Amplification? (maternal species)

  8. Dicer inactivation-in c elegans • Dcr1 is required for germ line formation • Dcr1 is not essential for later development • Behaviors? • Amplification? (maternal miRNAs) Lin4 and Let7 phenotype

  9. Dcr1+/- X Dcr1+/- Dcr-/- Dicer inactivation-in zebra fish

  10. Dcr1+/- X Dcr1+/- Dcr-/- Dicer inactivation-in zebra fish Dcr1 not required for early gastralation/specifications events, but needed for later events. miRNA production?

  11. Dicer inactivation-in zebra fish (dead end: germ line depletion) (Mir430) (Science, 308, 833)

  12. Dicer inactivation-in zebra fish (Science, 308, 833)

  13. Dicer inactivation-in zebra fish (Science, 308, 833)

  14. Dicer inactivation-in zebra fish (Science, 308, 833)

  15. Dicer inactivation-in zebra fish (Science, 308, 833)

  16. Dicer inactivation-in zebra fish (Science, 308, 833)

  17. Dicer inactivation-in zebra fish Dicer inactivation-in zebra fish • miRNAs: • “mostly” dispensable for cell type specification, but critical for morphogenesis--intercellular communication. • Probably not required for activation of cell fate specification genes Zebrafish and c. elegans: Differential effect on germ line formation, or non-autonomous effect?

  18. Dicer inactivation-in zebra fish Piwi RNA in germ line niche Dicer function for mouse germ line formation: WT Dicer-/- Spindle and DNA (G &D: 61,644)

  19. Dicer inactivation-in zebra fish Piwi RNA in germ line niche Dicer function for mouse germ line formation: (G &D: 61,644)

  20. Dicer inactivation-in mouse

  21. Dicer inactivation-in mouse T expression Oct 4 expression (Nature Genetics, 35, 215)

  22. Dicer inactivation-in mouse T expression Oct 4 expression What cells are missing? (Nature Genetics, 35, 215)

  23. Dicer inactivation-in mouse Conditional alleles: Yeast, C elegans, Drosophila, Zebrafish, Mammals,

  24. Dicer inactivation-in mouse Conditional alleles: Yeast, C elegans, Drosophila, Zebrafish, Mammals: integrases Cre/LoxP, FRT/Flpe, PhiC31 PLoS ONE. 2: e162

  25. Dicer inactivation-in mouse Conditional alleles: (G &D: 19, 489)

  26. Dicer inactivation-in mouse What do you expect? (G &D: 19, 489)

  27. Dicer inactivation-in mouse What do you expect? (G &D: 19, 489)

  28. Dicer inactivation-in mouse miRNAs: required for differentiation (G &D: 19, 489)

  29. Dicer inactivation-in mouse

  30. Functional comparison between TFs and miRNA miRNA vs transcription factors Mode of action post-transcription transcription

  31. Dicer inactivation-in mouse Heterochromatin formation affected in Dicer-/- ES cells ? Methylation pattern miRNA ? Dnmts (G &D: 19, 489)

  32. Dicer inactivation-in mouse Heterochromatin formation affected in Dicer-/- ES cells ? Methylation pattern miRNA ? Dnmts (G &D: 19, 489)

  33. Dicer inactivation-in mouse Specific functions: • 110, 000 miRNA molecules/ES cell. • More than half belongs to four clusters, related with cell cycle • more than 300 different types (PNAS: 104, 18097)

  34. Dicer inactivation-in mouse Zp3- Cre to produce oocyte-specific deletion

  35. Dicer inactivation-in mouse MyoD-Cre for muscle (DB: 311, 359)

  36. Dicer inactivation-in mouse (DB: 311, 359)

  37. Dicer inactivation-in mouse (DB: 311, 359)

  38. Dicer inactivation-in mouse (PNAS: 103, 2209)

  39. Dicer inactivation-in mouse (PNAS: 103, 2209)

  40. Deletion of individual miRNAs Only two examples so far in mouse: Mir208 and Mir155

  41. Deletion of individual miRNAs Only two examples so far in mouse: Mir208 and Mir155 Accompanied by abnormal immune cell deposition +/+ -/- Science: 316, 604 and 608)

  42. Deletion of individual miRNAs Only two examples so far in mouse: Mir208 and Mir155 Science: 316, 604 and 608)

  43. Deletion of individual miRNAs Only two examples so far in mouse: Mir208 and Mir155 Survival: Pathogen challenge Science: 316, 604 and 608)

  44. Deletion of individual miRNAs Only two examples so far in mouse: Mir208 and Mir155 IG production: Pathogen challenge Science: 316, 604 and 608)

  45. Deletion of individual miRNAs Only two examples so far in mouse: Mir208 and Mir155 T-cell proliferation Science: 316, 604 and 608)

  46. Redundancy or the functional mode of miRNA?

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