1. CHMI 4237 E Special topics in Biochemistry Eric R. Gauthier, Ph.D. Dept. Chemistry-Biochemistry Laurentian University 1- Introduction 1CHMI 4237 E - Winter 2010

2. Central theme: Cellular Biochemistry This course aims at providing a detailed view of several cellular phenomena at the molecular level. I will also explain how different experimental methods can help scientists to shed light onto the molecular basis of various cellular processes. I will finally illustrate how alterations in normal molecular events can change cell behavior and lead to diseases. 2CHMI 4237 E - Winter 2010

3. Course outline (tentative) 1) Introduction: cell structure vs gene expression and its regulation; 2) Cell proliferation: cell cycle, mitosis, checkpoints ◦Diseases: cancer, viral infections 3) Cell survival and cell death: apoptosis, necrosis, autophagy; ◦Diseases: cancer; auto-immunity; Proteus syndrome 4) Cell response to stresses: heat shock, hypoxia, nutrient starvation: ◦Diseases: cancer; 3CHMI 4237 E - Winter 2010

4. Course outline (tentative) 5) Cell structure, adhesion and motility: cytoskeleton, intracellular trafficking ◦Diseases: metastasis, progeria 6) Cell differentiation: stem cells, cell fate selection ◦Diseases: hematopoietic diseases 7) Cell aging and lifespan: basic hypotheses, lifespan extension ◦Diseases: aging 8) Cells and development: morphogen gradients, limb patterning, control of organ size ◦Diseases: freaks 4CHMI 4237 E - Winter 2010

5. Cell structure – general features 5CHMI 4237 E - Winter 2010

6. Cellular organelles 6CHMI 4237 E - Winter 2010

7. Molecules: the basis of cell structure and function 7CHMI 4237 E - Winter 2010

8. The cytosol: a crowded place 8CHMI 4237 E - Winter 2010

9. Molecules of the cell 9CHMI 4237 E - Winter 2010

10. Molecules of the cell - sugars 10CHMI 4237 E - Winter 2010

11. Molecules of the cell - lipids 11CHMI 4237 E - Winter 2010

12. Molecules of the cell – amino acids 12CHMI 4237 E - Winter 2010

13. Molecules of the cell – amino acids 13CHMI 4237 E - Winter 2010

14. Molecules of the cell – proteins 14CHMI 4237 E - Winter 2010

15. Molecules of the cell – proteins 15CHMI 4237 E - Winter 2010

16. Molecules of the cell – nucleic acids 16CHMI 4237 E - Winter 2010 Wikipedia

17. Cell signaling 17CHMI 4237 E - Winter 2010 Input Receptor Intracellular molecules Phenotype Environment Detector Decoding / Transmitting Adaptation

18. Cell signaling 18 Input Receptor Intracellular molecules Phenotype Molecules Osmotic pressure Temperature VoltageLight Transmembrane Nuclear Cytosolic Proteins DNA RNA LipidsIons Sugars Proliferation Survival Differentiation Motility

19. Cell signaling 19 Proliferation Differentiation Motility Survival Modified cellular Response to stimuli Blood cell production Stress responseUnicellular  multicellular Memory / learning Wound healing Neurite extension Intestinal epithelium GrowthOrgan size Cellular contacts Immune response

20. Changes in phenotype  adaptation 20 Proliferation Differentiation

21. Changes in phenotype  adaptation 21 Motility Growth

22. Changes in phenotype  adaptation 22 Cellular contacts Survival

23. Changes in phenotype  adaptation 23 Stress response Memory / learning

24. Complexity of intracellular signalling 24

25. Cell signaling 25CHMI 4237 E - Winter 2010 SignalProcessingResponse

26. Central dogma of molecular biology 26CHMI 4237 E - Winter 2010 Wikipedia

27. Molecules of the cell – nucleic acids 27CHMI 4237 E - Winter 2010 Wikipedia

28. Turning gene expression ON 28CHMI 4237 E - Winter 2010 NOTE: The sequence of the mRNA is complementary to the template (non-sense, transcribed) strand and is identical to the non-template (sense, non-transcribed) strand on the DNA Weaver. Molecular Biology. 3rd edition. McGraw Hill. 2008.

29. Gene Promoters 29CHMI 4237 E - Winter 2010 Glucocorticoid response element 5’ AGAACAnnnTGTTCT 3’ 3’ TCTTGTnnnACAAGA 5’

30. Transcription factors 30CHMI 4237 E - Winter 2010

31. Transcription machinery 31CHMI 4237 E - Winter 2010 RNA polymerase II Transcription factors

32. - Heterochromatin: repressed (no transcription) - Euchromatin: active (transcription is going on) - BUT: 10 µm nucleus vs 2 m genomic DNA CHMI 4237 E - Winter 201032 Gene expression – In the nucleus

33. DNA is tightly compacted in the nucleus 33CHMI 4237 E - Winter 2010

34. Nucleosome: basic unit for nuclear DNA packaging 34CHMI 4237 E - Winter 2010 - Histone octamer: - 2 x Histone H2A + H2B - 2 x Histone H3 + H4 - DNA: wraps around histone octamer (1.75 turns – about 146 bp) x 2 « beads on a string » 

35. The nucleosome 35CHMI 4237 E - Winter 2010 Histone N-terminal tail: the site of several modifications 

36. Histone tails are chemically modified 36CHMI 4237 E - Winter 2010 Legend: -A = active chromatin -R = repressed chromatin -Numbers: amino acid targeted by the modification -Ac = acetylation (CH 3 -COO-) -P = phosphorylation (PO 4 -) - Me = methylation (CH 3 -) -So: H3K9me3 = histone 3 with three methyl groups on lysine 9

37. Histone modifications are reversible Transcription factor Histone acetylation Opens up chromatin Recruitment of other TFs and RNA pol II

38. Histone modifications are reversible 38CHMI 4237 E - Winter 2010 Acetylated Histone: active chromatin Deacetylated Histone: repressed chromatin HDAC = histone deacetylase SUV39H1: « polycomb group protein»  silences gene expression

39. Cell signaling 39CHMI 4237 E - Winter 2010 SignalProcessingResponse

40. RNA processing 40CHMI 4237 E - Winter 2010 5’ untranslated region (5’UTR): ◦located upstream of the ORF ◦Generally short (<150b) ◦Involved in the regulation of the initiation of translation Open reading frame (ORF): ◦The region of the mRNA encoding a specific protein ◦Sequence is well conserved across species 3’UTR: ◦Located downstream of the ORF ◦Generally quite long (>1000 b) ◦Involved in regulating mRNA stability as well as translation mRNA 5’UTRORF3’UTR AUG UAA UGA UAG

41. RNA processing 41CHMI 4237 E - Winter 2010 http://www.exonhit.com/alternativesplicing/pages/diagrams/figure2.htm

42. Translation: making proteins 42CHMI 4237 E - Winter 2010

43. Membrane-embedded and secreted proteins 43CHMI 4237 E - Winter 2010

44. Secretory pathway 44CHMI 4237 E - Winter 2010

45. Membrane/secretory proteins transit through the endoplasmic reticulum 45CHMI 4237 E - Winter 2010

46. Membrane/secretory proteins transit through the endoplasmic reticulum 46CHMI 4237 E - Winter 2010

47. Membrane/secretory proteins transit through the endoplasmic reticulum 47CHMI 4237 E - Winter 2010