CHMI 4237 E Special topics in Biochemistry Eric R. Gauthier, Ph.D. Dept. Chemistry-Biochemistry Laurentian University 1- Introduction 1CHMI 4237 E - Winter 2010
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
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
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
Cell structure – general features 5CHMI 4237 E - Winter 2010
Cellular organelles 6CHMI 4237 E - Winter 2010
Molecules: the basis of cell structure and function 7CHMI 4237 E - Winter 2010
The cytosol: a crowded place 8CHMI 4237 E - Winter 2010
Molecules of the cell 9CHMI 4237 E - Winter 2010
Molecules of the cell - sugars 10CHMI 4237 E - Winter 2010
Molecules of the cell - lipids 11CHMI 4237 E - Winter 2010
Molecules of the cell – amino acids 12CHMI 4237 E - Winter 2010
Molecules of the cell – amino acids 13CHMI 4237 E - Winter 2010
Molecules of the cell – proteins 14CHMI 4237 E - Winter 2010
Molecules of the cell – proteins 15CHMI 4237 E - Winter 2010
Molecules of the cell – nucleic acids 16CHMI 4237 E - Winter 2010 Wikipedia
Cell signaling 17CHMI 4237 E - Winter 2010 Input Receptor Intracellular molecules Phenotype Environment Detector Decoding / Transmitting Adaptation
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
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
Changes in phenotype adaptation 20 Proliferation Differentiation
Changes in phenotype adaptation 21 Motility Growth
Changes in phenotype adaptation 22 Cellular contacts Survival
Changes in phenotype adaptation 23 Stress response Memory / learning
Complexity of intracellular signalling 24
Cell signaling 25CHMI 4237 E - Winter 2010 SignalProcessingResponse
Central dogma of molecular biology 26CHMI 4237 E - Winter 2010 Wikipedia
Molecules of the cell – nucleic acids 27CHMI 4237 E - Winter 2010 Wikipedia
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
Gene Promoters 29CHMI 4237 E - Winter 2010 Glucocorticoid response element 5’ AGAACAnnnTGTTCT 3’ 3’ TCTTGTnnnACAAGA 5’
Transcription factors 30CHMI 4237 E - Winter 2010
Transcription machinery 31CHMI 4237 E - Winter 2010 RNA polymerase II Transcription factors
- Heterochromatin: repressed (no transcription) - Euchromatin: active (transcription is going on) - BUT: 10 µm nucleus vs 2 m genomic DNA CHMI 4237 E - Winter Gene expression – In the nucleus
DNA is tightly compacted in the nucleus 33CHMI 4237 E - Winter 2010
Nucleosome: basic unit for nuclear DNA packaging 34CHMI 4237 E - Winter 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 »
The nucleosome 35CHMI 4237 E - Winter 2010 Histone N-terminal tail: the site of several modifications
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
Histone modifications are reversible Transcription factor Histone acetylation Opens up chromatin Recruitment of other TFs and RNA pol II
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
Cell signaling 39CHMI 4237 E - Winter 2010 SignalProcessingResponse
RNA processing 40CHMI 4237 E - Winter ’ 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
RNA processing 41CHMI 4237 E - Winter
Translation: making proteins 42CHMI 4237 E - Winter 2010
Membrane-embedded and secreted proteins 43CHMI 4237 E - Winter 2010
Secretory pathway 44CHMI 4237 E - Winter 2010
Membrane/secretory proteins transit through the endoplasmic reticulum 45CHMI 4237 E - Winter 2010
Membrane/secretory proteins transit through the endoplasmic reticulum 46CHMI 4237 E - Winter 2010
Membrane/secretory proteins transit through the endoplasmic reticulum 47CHMI 4237 E - Winter 2010