Cell Bio Module V
Cell Bio Module V BIOL 541
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This 12 page Study Guide was uploaded by . Notetaker on Tuesday August 11, 2015. The Study Guide belongs to BIOL 541 at Kansas State University taught by Dr. Stella Lee in Fall 2014. Since its upload, it has received 81 views. For similar materials see Cell Biology in Biology at Kansas State University.
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Date Created: 08/11/15
Chapter 12 Endomembrane lumen is on outside because they eventually go to exterior of cell a b Cisternae membrane bound sacs space inside lumen involves lipid synthesis Rough er ribosomes subdomain transitional elements forms vesicles that shuttle lipids to Golgi i Carbs form glycoproteins folding polypeptides then it assembles multimeric protein complexes also removes misfolded proteins 1 ERAD degrades incorrect proteins Smooth er tubular structures synthesis of secretory proteins cells that have lots of steroid hormones have lots of smooth ER i Function detoxes drugs by hydroxylation adds hydroxyl groups to drugs to increase solubility then they can be easily excreted 1 Catalyzes cytochrome p450 ii Carb metabolism breakdown of glycogen producing glucose6 phosphate iii Calcium storage in sarcoplasmic reticulum of muscle cells pumped by ATPdependend Ca ATPases iv Steroid biosynthesis large amounts in ER in adrenal gland hormones share 4 ring structure differ in of arrangments of c chains Biosynthesis membranes ER primary source of membrane lipids i Peroxisomecholesterol mitochondiraphosphatodelthmine ii Fatty acids cytoplasm amp incorporated into ER iii Flippases phospholipid translocators bilayer transfers Exchange proteins move phospholipids from ER to mitochondira chloroplast peroxisome nonendomembrane Golgi processes vesicles from ER packages for transport i Cisternae 38 golgi stack ii Cis vesicle from ER iii Trans vesicle transported out iv Medial cisternae between TGN amp CGN processing occurs v Biochemical polarity speci c proteins unique to each portion of network vi Stationary golgi is architectural structure vs cisternal maturation golgi makes itself from scratch Centrifugation isolation amp puri cation a b c d Tra a b Differential separates organelles based on sizedensity Sedimentation rate movement of particles thru solution depends on sizedensity Sedimentation coefficient how rapidly the particle sediments Density gradient sample placed thru layer on top of gradient solute nsport Anterograde movment of matrial toward plasma membrane retrograde flow of vesicles from golgi cisternae back to ER Roles of ER and golgi in glycosylation i N linked addition of oligosaccharide to nitrogen of asp residues ii 0 linked addition of oligosaccharide to oxygen atom on hdroxl group of certain serine or threonine residues iii Glycosylation occurs in ER all carb side chains have common oligosaccharide core consisting of two N acetylglucosamine nine mannose untis and three glucose iv Steps Begins as dolichol phosphate an oligosaccharide carrier Inserts into ER membrane CIcNA and mannose groups are added to phosphate group Growing oligo is transocated to ER by ippase Once inside lumen mannose amp glucose are added and is transferred from dolichol to Asp residue v Cotranslational gycosyation usually oligosaccharide is added to recipient protein as polypeptide is being synthesized 1 Parts a Calnexin or calreticluin binds monoglucosylated proteins b ERp57 disul te bond formation c Glucosidase ll removes glucose d Glucosyl transferase adds back glucose 2 Further gycoyation occurs in golgi proteins move from cis to trans vi Terminal gycoyations variable and create diversity 1 ER amp golig contain hundred sof gycosy tranferases c Protein traf cking once protein reaches destination i Prevents from leaving each protein contains tag AA sequence hydrophobic domain or an oligo side chain 1 Liquid tags help vesicles reach destinations attached to 345 of membrane phosphatidyl inositol 2 Stays in ER contains RXR sequence argxarg 3 Retrieval tags short cterminal sequences such as KDEL lys asp glu leu for luminal proteins or KKXX for transmembrane receptor proteins bring proteins to ER ii Golgi speci c proteins integral membrane proteins with hydrophobic membrane spanning domains 1 Lengths of domains may determine into which cisternae is conroporated 2 Thickness of cell membranes increases from ER 5nm to plasma membrane 8nm IV Lysosomal proteins to endosomes a Lysosomal enzymes in ER i Undergo Nglycoylation followed by removal of glucose and mannose units 1 Mannose residues on sid ehcains are phosphorylated within goli complex forming mannose6phosphate a First enzyme responsible phosphotransferase that adds phosphate to mannose and the second wewwe removes CIcNA leaving behind mannose6 phosphoate receptors ii Lysoosmal enzymes transported from TGN to organelles known at late endosomes iii Dissociated of lysosomal enzymes at low pH prevents retrograde movement 1 Late endosomes natures to form new lysosome 2 Genetic disorder I cell disease lysoosmal enzymes are misdirected a Results from defective phosphotransferase that is needed to add mannose6phosphate to oligosaccharide chains on lysosomal enzymes iv Secretory pathways 1 Move proteins from ER to Golgi to secretor vesicles and secretory granules these discharge to exterior of cell 2 Constitutive secretion after budding from TGN vesicles move directly to cell surface and immediately fuse with plasma membrane one example is mucus secretion by intestinal lining a Not a default pathway 3 Regulated secretion neurotransmitter release V Exocytosis a Animals secrete hormones mucus milk proteins digestive enzymes b Plantsfungal enzme and structural proteins c Luminal membrane vesicle becomes part of outer surface i Mechanism not clear vesicle movement stops when cells treated with colchicine movement of microtubules d Polarized secretion exocytosis of speci c proteins is limited to a speci c surface of cell Vl Endocytosis plasma membrane pinches off forming viesicle of stubstance a Phagocytosis engulf foregin material large particles 5 um acquire food specialized cells called phagocytes neutrophils and macrophages i Receptor mediated ligand complexes diffuse laterally and encounter regions called coated pits sites for collection adapter protein clathrin can bind 20 surface are has pits 1 Dynamin binds to vesicle to nally pinch it off 2 Variations cell becomes less responsive to desensitization also LDL receptors are constitutively concentrated and internalized indpenedinng of ligand binding 3 Early endosomes sites for sorting and recycling materials a Ph 5965 early endosome pH gradually lowers ligand and receptors allow receptors to be returned to membrane and ingested 4 Late endosomes mature from TGN which then develop to lysosomes VII VIII 5 Alternate fates some go to lysosome some carried to TGN some transported by vesicles to plasma membrane where they are secreted transcytosis 6 Fluid phase endocytosis pinocytosis does not concentrate ingested material contents routed to early endosomes Lysosomal storage diseases a an Type II glycogenesis accumulatin of excessive glycogen a 1 4 glucosidase Hunter accumulation of glycosaminoglycans Taysachs accumulation of a ganglioside in nervous system Human neuronal ceroid lipfuscinoses has 13 enzmymes that are responsible i CLN5 lysosomal lumen ii CLN8 ER Peroxisomes bounded by single membrane not dereived from ER a b c Not part of endomembrane degrades H202 Plant peroxisomes consist of ctrystaliin urate oxidase Essential roles i Oxidation of fatty acids ii H202 metabolism iii Detox of harmful compounds iv Metabiolim of nitrogen compounds Oxidases generate H202 in peroxisomes i Hydrogen peroxide is detoxi ed by catalase ii Catalase can function as a peroxidase iii Either way hydrogen peroxided is degraded while still inside peroxisome Detox harmful i Detoxify reactive oxygen species 02 Oxidation fatty i Enzymes for Boxidation of fatty acids 1 Aminals oxidizel long chain fatty aicds once fewer than 16 carbons rest occurs in mito 2 Plants completely oxidized in peroxisomes Metabolisms of nitrogen i Require urate oidase to oxidize urate thaen aminotransferases catalyze transfer of amino groups from amino acids to a ketoacids Catabolism unusual substances i Break down xenobiotics compounds foreign to living organism ii Also deal with D amino acids Disorders i Xlinked adrenoleukodstrophy 1 Transporting long chain fatty acids into peroxisome for B oxidation and don39t move out Biogenesis i Occurs by divison of preexisting peroxisomes peroxins required for this process 1 Membrane components matrix enzymes cofactors are incorporated into peroxisomes 2 Some may obtain materials or form de novo from vesicles derived from ER k Signals i Targets at lease some proteins to peroxisomes is called PTSl on cterminus ii Second PTS2 found on Nterminus Chapter 22 l Posttranslational processing a Methionine at Nterminus of often removed b Signal sequence of lumen proteins are removed once they across the membrane c Some undergo protein splicing i Sequences called interins are removed and remaining sequences called exteins are spliced together d Cotranslational import to ER i Signal hypothesis b blocel and Sabatini 1 Proteins move into ER during synthesis possess an intrinsic signal signals usually 1530 amino acids long a Positively charged nterminal region hydrophobic region and polar region near cleavage from mature protein will take place proteins contain ER signal referred to as preproteins 2 Contact with ER mediated by signal recognition particle SRP binds ER signal sequence SRP binds signal sequence and blocks further translation SRP brings in ER membrane called translocon a Composed of SRP receptor binds SRP b Ribosome receptor binds ribosome c Pore protein channel for peptide to enter d Signal peptidase remove ER signal sequence 5 Next GTP binds SRP unblocking translation a Central channels open b GTP is hydrolyzed and SRP released c Polypeptide elongates passes into ER lumen and released into lumen and ribosome detaches from ER membrane ii Protein folding 1 Chaperone Hsp70 called BiP binds regions of polypeptide chains and prevents aggregation of polypeptides a Often have disul de bonds 2 Unfolded two responses a Unfolded proein response UPR defect misfolded sensors actiate pathways that enhance need for foliding and degradation PS b ER associated degradation ERAD removes proteins and degraded by proteasomes iii Integral membrane protein signal sequence and stoptransfer sequcne 1 Ctermini facing lumen internal starttransfer sequcne 2 Multiple membrane spanning alternating patter of start transfer and stoptransfer sequnces e Posttranslational import to nonendomembrane system organelles i Nuclear interior mito chloroplast peroxisome ii Nuclear 1 Nuclear localization signals target proteins for transport thru pores iii Mitochondria amp chloroplast 1 Trasit sequence nterminus then transit peptidase removes sequence 2 Once sequcne binds to receptor polypeptide tranlocatd across outer membrane thru TOM or TOC 3 Transport complexes a TOM translocase of outer mito membrane b TlM translocase of inner mito membrane 4 Chloroplast TOC amp TIC 5 Polypeptides transported in unfolded state a Hsp70 class chaperones bind newly forming polypeptide chaperones released ATP hydrolyzes then mito Hsp70 amp60 help to fold polypeptides in matrix 6 Many proteins require multiple signals a Thylakoid membrane signal Chapter 15 amp16 l Cytoskeletal system a Microtubules hollow tube with 3 proto laments outer 25nm inner 15nm a amp b tubulin ends cytoplasmic like animcal cell shape chromosome movemnts intracellular transport traf cking movment of organelles i Axonemal er cell motility b Micro laments two interwinded chains of F actin diameter 7 nm g actin ends fucntions contractions cell locomotion streaming cytokinesis cell shapte transporttraf cking c lntermediate eight proto laments joined end to end with staggered overlaps 812 nm no known polarity struc support cell shapte formation nuclear lamina strength of nerve cell axons keeps muscle bers in register d Bacteria i Bacteria amp archaea have polymer systems that function similarly to euk cytoskeleton 1 Actinlike MreB protein involved in DNA segregation 2 Tubulinlike FtsZ protein involved in regulating division 3 Intermediate lamentlike crescentin is regulator of cell shape e Microscope techniques i Fluorescence directly bind to cytoskeletal proteins 1 Ex show bundles of actin laments ii Live cell uorescents into living cells 1 Ex tubuin dimers incorporated into microtubules iii Computer enhanced digital video increase contrast and remove background 1 Ex microtubuoles in detail iv Electron resolve individual laments 1 Ex bundles of actin micro laments f Motile systems i Occurs in the following 1 Tissue muscle contraction micro laments 2 Cellular ciiated protozoa sperm agella 3 Subcellular separaton of chromosomes microtubules vesicle transport 4 To generate movement microtubules and laments provide scafford twith motor proteins or mechanoenzymes which interact with cytoskeleton to move g Microtubules i Largest of cyto components and have two types Cytoplasmic a Maintain axons b Formation mitotic and meiotic spindles c Maintain cell shape d Placement and movement of vesicles axonemal a Cilia b Flagella c Basal bodies to which ciia and agella attach d The axoneme the central shaft of cilium is highly ordered bundles of MTs ii Tubuin heterodimers building blocks 1 MT are straight and consist of 13 longitudianl arrays called proto aments a Basic subunit tubulin one alpha and one beta b Bind to form aB heterodimer iii Structure 1 Alpha amp beta have similar structure but only have 40 same amino acid identity each has N terminal GTP binding domain central domain and C ternial 2 All dimers oriented same way 3 Polarity two ends differ both chemically and structurally iv lsoforms 1 Cytoplasmic MT simple tubes or singlet MR with 13 proto laments vi vii Viii X xi 2 Some have doublet or triplet MT which contain 1 13 tubule A and one or two additional incomplete rings B and C of 10 amp 11 proto laments Addition of tubulin dimers 1 MT form by reversible polymerization of tublin dimers in GTP and Mg 2 Aggregate into oigomers serve as nuclei a Called nucleation addition is called elongation 3 When mass of MT reaches where tubulin is diminished reaches plateau phase Critical concentration 1 Tubulin concentration where MT assembly is exactly balanced by disassembly is called critical concentration 2 Two ends of MT can grow or shrink much faster than the other because they have different critical concentrations 3 Treadmilling a If tubulin subunits is above critical concentration for plus end but below of minus end treadmiing will occur Drugs 1 Nocodazole effects easily reversed than those of cochicine 2 Taxol binds to MT and stabilizes them depletes tubulin subunits causing dividing cells to arrest during mitosis 3 Drugs called antimitotic useful for cancer treatment GTP hydrolysis 1 Tubulin binds 2 GTP one a and one B 2 GTP bound to B can be hydrolyzed to GDP 3 GTP needed to promote heterodimer interactions and addition to MT but its hydrolysis is notrequired to MT assembly Dynamic instability model one population of MT grows by polymerization at plus while another population shrinks by depolymerizaiton 1 Growing MT have GTP at plus ends and shrinking have GDP a The GTP Gap at plus ends prevents subunits removal b If GTP is high it is added to MT creating large GTP tubulin cap c Concentration falls rate of tublin addition decreases 2 Low GTP rate of hydrolysis exceeds rate of subunit addition and cap shrinks Catastrophe and rescue 1 If GTP cap disappears MT becomes unstable 2 Switch from growth to shrinkage is called catastrophe 3 Sudden switch back is called microtubule rescue Origins 1 MT originate from microtubuleorganizing center MTOC a Centromere in center in aminal its associated with two centriolesIsurrounded by pericentriolar material 2 Centriole a Walls formed by 9 pairs of triplet microbubules b Oriented at right angles amp involved in basal body formation for cilia amp agella c Y tubulin i Large ring shaped complexes which contain gamma tubulin L MTOC 1 Nucleate and anchor MT grow outward with xed polarity the minus ends are anchored in MTOC xiii MAPS microtubule associated proteins 1 Bind at regular intervals along microtubule wall a Tau cases MT to form tight bundles in axons b MAP2 formation of looser bundles in dendrites c Protiens that promote depolarization i Stathmin catastrophins katanins h Kinesin amp dynein i Traf c towards minus ends of MT considered inbound toward plus end is outbound ii Kinesins toward plus ends 1 Three parts a Globular head attaches to MT b Coiled helical region c Light chain involved in attaching kinesins to other proteins or organelles d Kinesins look like they have two legs walking iii Dyneins toward minus ends 1 Cytoplasmic moves toward minus ends of MT 2 Associated with dynactin helps link it to cargo 3 Axonemal dyneins 4 different types iv Organelles moving along MT 1 Fast axonal transport involves movement of vesicles and organelles along MT 2 Kinesin 1 involved ATP dependent transport toward plus end called anterograde axonal transport 3 Cytoplasmic dynein moves particles in opposite retrograde i Structure cilia and agella i Each outer doublet of axoneme consists 1 MT A and one imcomplete MT B 1 A 13 310 or 11 ii Central pair are complete in tubules contain tubulin and tektin 1 Tektin related to IF proteins Aamp B tubules share wall in which tektin is major component iii Tubule structure 1 A has sidearms that project from outer doublets consists of axonemal dynein a Dynein involved in sliding of MT vs each other bends the axoneme i Dynein arms occur in pairs one inner and one outer iv Axoneme structure 1 Adjacent doublets joined by interdoublet links by poretin called nexin limit the extent of relative movment of doublets 2 Radial spokes project inward toward center a Important in translating the sliding of MTs into the bending of axoneme v Sliding microtubule model sliding of MTs relative to each other is converted into localized beinding because doublets are connected to central pair 1 Takes form of wave that begsin at base and proceeds up 2 Resistance provided by spokes that connect doublets to central piar nexin crosslinks between doublets connections convert sliding to bending vi lntrafalagellar transport 1 Tubulin subunits shuttled to and from frowing agellum tip by both plus and minus ends a Kinesins move material to tips of agella and dynein bring material back to base j Intermediate lamnets i Abundant in keratin tissue speci c ii Fibrous rather than globular dimer tetramer proto laments then intermediate ll Micro lament a Smallest develop and maintain cell shape just beneath the cell cortex structure core of microvilli b Actin abundant once synthesized folds into globularshaped molecule that can bind ATP or ADP gactin i G molecules polymerize to form micro lamnets Factin ii Aactins muscle speci c actins iii Nonmuscle B actins Y actins 1 B at apical 2 Y at basal iv Polarity 1 Plus end called barbed end and minus end called pointed end 2 Polarity re ected in more rapid addition or loss of G actin at the plus end thatn the minus end a After G actin assemble onto micro lament ATP bound to them is slowly hydrolyzed b Most composed of ADP actin 3 Drugs a Cytochalasins prevent addition of new monomers to existing MF block polymerization b Latrunculin A toxin equesters actin monomers and prevents their addition to MF blocks polymerization c Phallodin prevents their depolymeraiztion 4 Assembly into structures a Crawl have lamellipodia and lopodia at their leading edge allows them to move b Cells that adhere tightly to the underlying substratum have organized bundles called stress bers 5 Polymerization regulation a Concentration of ATP bound G actin is high MF will assemble until G actin is limiting i Thymosin B4 binds ADP bound G actin whereas pro ling exchanges G actin ADP to ATP ii Pro ling competes with thymosin iii Capping binds end of lament to prevent further loss of subunits 1 Gelsolin breaks actin and caps the newly exposed plus ends b Proteins that crosslink actin laments i Filamin is important in formation of these networks 1 Acts as splices joining two MF together where they intersect 2 Long molecule consisting of two identical polypeptides joined head to head actin binding site at each tail c Link actin to membranes i Indirect connection to membrane requires linking proteins spectrin and ankyrin d Actin branching i Actin form dendritic network ii Arp23 complex nucleates new branches on sides of laments v Microvilli core consists of right bundle MF with plus ends pointed toward tip 1 Connected to membrane by crosslinks made of myosin calmodulin 2 Bundle tightly bound by mbrin and villin 3 Base of microvillus extends network called terminal web a Stabilized by spectrin vi Cell signaling 1 lnositol phospholipids a PlP2 binds to pro ling CapZ ezrin i Recruits these proteins to membrane and regulates their interactions with actin 2 Rho family small G proteins a Rho Ra cdc42 i Rho stimulated by guaninenecleotide exchange factors thru exchange of GDP for GTP ii GAP inactivate Rho b Activation of rho pathway results in formation of stress bers c Rac activation results in extension of lamellipodia d Cdc activation results in lopodia c Nonmuscle cell movement Vl vii Viii Actin based lamellipodia or filopodia attaches to substrate generations of tension which pulls cell forward Protrusions at leading edge actin grows at tip Retrograde ow actin moves forward which pushes cytoplasm back to move 1 Cytoplasm attaches to membrane with integrins 2 Contraction due to actinmyosin interactions is under Rho activates myosin ll Chemotaxis moves towards chemical gradient Ameobas gelatins Cytoplasmic sreaming actin amp myosin process called cyclosis Actin based 1 Myosins ATP dependent have heavy amp light chains 2 Type II two heavy chains a Kinesisns operate in small numbers myosins large numbers Myosin based 1 Thick filament mosin thin lamentactin 2 Thin filament factin intertwined with tropomyosin amp troponin a Calcium sensitive switch
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