BCOR 103 1st Edition Lecture 18 Outline of Last Lecture I. PKA ActivationII. Second MessengersIII. Kinases and Signaling ProteinsOutline of Current Lecture I. ApoptosisII. CaspasesIII. Intrinsic and Extrinsic PathwaysIV. Cell CycleCurrent Lecture- Apoptosis- programmed cell deatho Balances cell division and cell death to maintain constant cell numbers in tissues- homeostasiso Defense mechanisms in response to pathogen infections, cell damage, and DNA damageo Elimination of unwanted cells during development- Cellular Effects of Apoptosiso DNA fragmentation occurs, chromatin condenses, fragmentation of the nucleus, then fragmentation of the cell- Apoptosis in C. elegans requires Ced-3o During development, 131 cells die through apoptosiso Results in exactly 959 somatic cells in adult wormso In mutant worms that have Ced-1 There is no engulfment of apoptotic dead cells Ced-1 is required for signaling to engulf apoptotic cells-receptors- Caspases Function in Apoptotic Protein Degradation These notes represent a detailed interpretation of the professor’s lecture. GradeBuddy is best used as a supplement to your own notes, not as a substitute.o Caspases: proteases with cysteine (C) resides at their active sites that cleave afteraspartic acid residues in substrate proteinso At least seven caspases in mammalian cells; only 1 (Ced-3) in C. eleganso >100 target proteins are destroyed- Functions of Caspaseso Inhibitor of DNase (ICAD)  DNA fragmentationo Nuclear lamins  fragmentation of nucleuso Cytoskeletal proteins, actin, myosin, α-actinin, tubulin, vimentin  cytoskeletal disruption, cell fragmentation, membrane bubblingo Golgi Matrix Proteins  fragmentation of Golgi- Caspases are activated by Proteolytic Caspase Cascadeso All caspases are activated by proteolytic cleavage, which is catalyzed by other caspaseso Caspases can be either initiator or effector caspases o Initiator caspases: activated in response to primary apoptotic signalo Effector caspases: activated by initiator caspases; they cleave target proteins- Intrinsic Pathway of Apoptosiso Two major pathways by which signals can trigger apoptosiso Intrinsic pathway: cell damage leads to release of cytochrome c from mitochondria a.k.a activated T-cell autonomous death (ACAD)o IAPs: inhibitor of Apoptosis proteins Suppress apoptosis by inhibiting caspase activity- Extrinsic Pathway of Apoptosiso Extrinsic pathway: secreted polypeptides activate cell death receptors, a.k.a activation-induced cell death (AICD)o Polypeptides belong to tumor necrosis factor (TNF) familyo TNFs bind to TNF receptors Leads to activation of initiator caspases (e.g. caspase 8) Ex: Fas receptor: functions in immune cell death- Apoptosis and Bacterial Pathogenso Bacterial pathogens and HIV can either inhibit or induce apoptosis through eitherintrinsic or extrinsic apoptosis pathways- Cell Survival Signaling requires PIP3o PIP2 (PI 3-Kinase) PIP3o PIP3(PTEN)PIP2- Cell Survival Signaling: Anti-Apoptosiso PI 3-Kinase and Akt: human oncogeneso PTEN: human tumor suppressor gene- Eukaryotic Cell Cycle and its Regulatorso Cell division cycle: process by which eukaryotic cells grow and divide- Cell Division Cycle must occur with High Fidelityo DNA must be duplicated without errorso Chromosomes must segregate equally to daughter cellso Centrioles (centrosomes) must be duplicated and distributed to daughter cells during mitosiso Transcriptional programming must be maintained or reprogrammed in response to environmental cueso Mitochondria and other cellular organelles must be distributed to daughter cellso Cell division must be carefully controlled during development and in tissueso Cell division must be integrated with apoptosis- Phases of Eukaryotic Cell Cycleo Mitosis (M phase): nuclear division; separation of daughter chromosomeso Interphase: period between mitoses; growth and DNA replication occurs G1 phase: gap between mitosis and DNA replication; cells are metabolically active S phase: DNA replication G2 phase: gap between DNA replication and mitosis; cells continue to synthesize proteins G0 phase: quiescent stage- Cells are metabolically active but do not proliferate- Extracellular signals can induce cells to re-enter G1 phase- Embryonic Cell Cycleso Very rapid cell divisionso No G1 or G2 phases- cells do not grow!o Only M and S phases- very short S phase- Dependency Mechanisms vs. Clock Mechanismo Cells integrate duplication of cell material with growtho Stage specific events vs. continuous processes Events depend on previous event being completedo Progress monitored at checkpointso DNA replication occurs immediately after mitosis and doesn’t depend on mitosis being completedo Suggest a clock mechanism for control- Techniques used to study Cell cycleo Biochemical approaches: Examine enzyme activities or protein levels that fluctuate during the cell cycle Determine cellular DNA content using fluorescent dye (propidium iodide) that binds to DNAo Cytological approaches: Morphology of yeast cells can distinguish stages of cell cycle Fluorescent dyes stain actin, tubulin, etc.o Genetic approaches: Isolate mutants with defects in cell cycle- cell division cycle (cdc) mutants- Most cdc mutants affect essential cell cycle events and are lethal events- must use conditional lethal mutants - Comparison of Yeast and Mammalian Cell Cycleso Genetic (yeast) and biochemical (mammal) approaches converged to show that cell cycle regulation is fundamentally the same for all eukaryotes*Transit though the cell cycle is regulated by phosphorylation/dephosphorylation and