- Created by: olivia hodges
- Created on: 21-01-13 15:18
Vacuole - increases cell surface area, provides storage for cell waste and crucial for maintaining the plant's rigidity. cytoskeleton - define centre, establish polarity of cell, provides routes for transportations, network of fibrious proteins, actin, intermediate filaments and microtubules
cystol - semi-fluid portion of cytoplasm a mix of water, various ion, small molecules and proteins. site of intermediary metabolism ( generation of precursors necessary for biosynthesis of the larger molecules).
Lysosomes - break down and recycle worn our cell components. Contain enzymes for breaking down complex molecules. role in defence against infection.
Extracellular matrix - support and anchorage for cells, separates tissues, regulated intercellular communication. Glycoproteins, proteoglycans and hyaluronic acid, collagen, fibrin, elastin, fibronectins, laminins, nidogens,
Golgi - modify, sort and pachage substances for cell secretion, creates lysosomes and transports lipids, composed of membrane bound sacs known as cisternae.
RER - site of synthesis of many proteins, complex interconnected series of flattened sacs or membranes. Proteins synthesised at ribosomes, packaged proteins bud off and transported out of cell.
Protein function - oxygen binding proteins
Ligands binding may involve conformational change - induced fit. Allosteric effects in multisubunit proteins ( shape change in all units when one changes).
Oxygen (final electron acceptor in ATP phosphorolation, can be bound to a heme prosthetic group. o2 is non-polar. Amino acid R-groups dont bind to o2 but transition metals such a iron and copper do. An iron prosthetic group in haemoglobin. only Fe(II) binds not Fe(III). Myoglobin has a single binding site for o2. All globins are based on 8 alpha helicies. The smaller the Kd number the tighter the ligand binds. Higher o2 pressure higher ligand binding saturation.
Arterial blood 96% saturated with 02, venous blood only 64%. 2 alpha 2 beta chains. Each chain has a single heme prosthetic group can carry 4 o2. hemo and myo have same tertiary structure with pocket for o2 with heme. Structural changes are triggered by o2 binding, iron is moved which pulls histodine sidechains, which break the salt bridges that stablise the T-state.
Hemoglobin tetramer can exist in low affinity and high affinity states. Co-operative binding where it binds to o2 in lungs (high p02) and releases it in tissue (low pO2). Myoglobin has much higher affinity for o2 than hemoglobin so is used for muscles which need endurance.
hemoglobin can also bind to CO2 to remove it from blood.
Bohr effect - low pH means high H and CO, shifts teh o2 binding curve which means in exercising muscles pH is lower which decreases hemoglobins affinity for O2 so it will release it more readily.
o2 binding is regulated by BPG, which binds to deoxy form of hb. One molecule binds per tetramer and decreases o2 affinity. when pO2 is low BPG synthesis increases. Athletes train at high altitudes so their bodies make more BPG as it shifts curve to right so o2 is picked up more easily in lungs and released more easily in the muscles.
Sickle cell anemia - one amino acid change, glutanamic acid to valine. blocks capilloriesas they bind together.
foetal hemo alpha2gamma2. placenta place of gas exchange not lungs. has higher affinity for o2 than mothers hemo so it can cross over placenta.
Immunoglobulins and molecular motors
antigen - any molecule or pathogen able to illicite an immune response.
epitope - particular molecular structure within an antigen which an antibody or T-cell receptor binds.
haptens - small molecule covalently coupled to carrier proteins to produce an immune system.
macrophages - ingest large particles and cells by phagocytosis.
B - cells produce and secrete antibodies
Cytotoxic T-cells - interact with infected host cells through receptors
Helper T-cells - interact with macrophages and secrete cytotoxins that stimulate other B and T cells to activate.
MHC proteins on outside of host cells, main way infected cells signal to T-cells. class I and class II versions. both have 2 polypeptide chains and a cleft for displaying antigens of foreign pathogens. class I on all nucleated cells, class II on specialist cells e.g macrophages
antigen enters cell, enzymes break it down and antigen pieces bind to MHC protein inside ER. This is then transported to the cell surface on cell membrane by golgi to present the antigen. Helper T-cells then bind to this and activate cytotoxic T-cells which divide by mitosis and kill infected cells. Effector B-cells then make antibodies and memory B and T cells remember it.
antibody - 2 heavy chains 2 light chains for antigen binding sites. Variable region at the ends, and 3 constand in heavy, 1 in light. Antigens bind very tightly to antibodies. 5 classes of immunoglobin and the most abundant is lgG.
can exploite anitbodies using immunoblots: coat surface with samples (antigens), block unoccupied sites with nonspecific proteins, incubate with primary antibody against specific antigen, incubate with anti-body-enxyme complex that binds to primary antibody, add substrate, formation of coloured product indicates prescence of specific antigen.
chymotrypsin - in stomach, separates amino acids. Hydrogen + electrostatic bonds in binding site. Some enzymes also contain cofactors e.g iron in hb. enzyme purification - homogenise broken cells to get cell extract, separate in column chromotography.
column chromotogrpahy - column of solid porous matrix. to equlibrate the column pour aqueous buffer solution -the same composition of that the proteins are disolved in. Load the column with sample containing many proteins. Then separate the proteins by eluting with buffer. Separation due to size, charge or ability to bind to different molecules.
size exclusion chromotography - porous beads allow small proteins in and big ones flow out.
Ion exchange chromotography - positively charged matrix, apply mixture of proteins and low NaCl concentration. elude - most positive come out first. Anion (negative) use cellulose with DEAE positive group. For Cations use CM negative group attached to cellulose or agarose. NaCl disrupts bonds meaning proteins are released so can actually flow out. Affinity chromotography - matrix with ligands attached, proteins which dont attach to the ligands flow through. After loose ligands are put through so proteins willl bind to them and flow through.
assay of enzyme activity - measure conc of product at regular time intervals. can use this to identify the fraction containing the enzyme of interest. initial velocity of reaction defines the conc of product formed per unit of time. specific activity = moles of substrate converted per unit time per mass of protein.
beer lambert law = A=ecl absorbance = extinction coefficient x conc x path length
enzyme graphs start proportionally, enzyme fixed substrate increased, then slows down and plateued to maximum velocity = vmax. Km (michaelis constant) is vmax/2. When rate is proportional to enzyme conc, vmax changes Km doesnt. competitive inhibition - Km is raised Vmax is unchanged. Uncompetitive inhibitor - Km is lowered and so is Vmax.
michealis-menton equation - velocity of enzyme catalised reaction = vmax[s]/km + [s]. when substrate conc is less than km velocity is proportional to s. when s is bigger than km, km is negligable and s is cancelled out so velocity = vmax so velocity is independant of substrate conc.
when enzyme conc is changed rate is proportional to enzyme conc and vmax changes but km doesnt.
Kcat = turnover number (times enzyme can complete catholitic cycle. Vmax = [e]total. so vmax = [E]total x Kcat. therefore the new michaelis-menton equation is = Kcat x [E]total x [S] / Km + [S]. Km and Kcat are charactoristics properties of enzyme at a particular ph and temp.
enzymes is health - colorimetric glucose sensors for diabeties. peroxidase enzyme for break down of glucose turns quinoeimine dye blue so colour change on *****. anemia can be detected by high levels of lactate dehydrogenase and heart attack by creatine kinase.
lineweaver-burk plot - slope = km/vmax. y intercept = 1/vmax and x intercept = -1/km. because cant find true vmax as dont have unlimited enzyme and substrate. when competitive inhibtor used the y intercept will be same for all but x intercept will change. more inhibitor means steaper gradient.
inhibitors = inhibit prostaglandins release by reveraibly inhibiting COX enzyme using ibuprofen. this is competitive inhibition where km is raised but vmax stays the same. uncompetitive inhibitions lowers both km and vmax.
energy realeased by reaction. enzymes lower activation energy.
regulation by covalent modification - glycogen is realeased by glycogen phosphorylase to give glycose-1-photsphate. the activity of enzyme is regulated by shape changes triggered by phosphorylation of a serine side chain. allosteric enzymes undergo conformaional changes in response to non-covalent binding of an allosteric modulator e.g asparatate transcarbamoylase.
oxidoreductases - transfer electron. transferases - group transfer reactions. hydrolases - hydrolysis reactions. lysases - addition of groups to double bonds or formation of double bonds by removing groups. isomerases - transfer of groups within molecules to yeild isomeric forms. ligases - formation of c-c, c-s, c-o and c-n bonds by condensation coupled to ATP cleavages.
3 kingdoms - Eubacteria - true bacteria, prokaryotes, cell wall contains peptidoglycan. Archaebacteria - ancient odd bacteria some able to withstand extreme environments, cell wall contains pseudo-peptidoglycan. Eukaryotes - protists, fungi, plants and animals, eukaryotes no peptidoglycan. prokaryotes have no membrane bound organelle.
bacteria - small, high surface/volume ratio, different morpholg. coccus, bacillus rod or spiral. bacillus can join together in chains. cell wall peptidoglycan has tetrapeptide attached to NAM and NAG molecules with glycocidic bonds and peptide cross-bridges.
Gram positive - Thick peptido 60-90% cell wall, interwoven teichoic acids, no lipids - purple. Gram negative - thin pepto 10-20%, has lipopolysacharide outer membrane and periplasm between - red. ethanol dehydrates the thick peptido walls trapping the crystal violet-iodine complex, but dissolves lipid outer membrane letting it escape from negative bacteria. Counterstain safranin stains negative cell walls pink.
Cell division - binary fission with bidirectional DNA-replication. one origin of relication. Growth is limited to lack of nutrients and production of inhibitory waste product. Lag pahse - exponential growth - stationary - death. growth rate = n/time. N=initial no. x 2n. g=time/n. no. generations = logN - logNo/0.301
nuclear lamins - intermediate filaments that line the inner facing of the nucleus, provide attachment for dna.
smooth ER - site of synthesis of lipids and membrane proteins. rough ER - ribosomes on surface, site of protein synthesis. when proteins leave the RER its transferred to golgi in vesicles and processed by lysosomes, peroxisomes and secretoryvessicles.
Golgi - main site of carb synthesis, package micromolecules for transport elsewhere.
lysosomes - contain hydrolytic enzymes, rare in plants. Peroxisomes - protection against toxic hydrogen peroxide, contain oxidative enzymes which break it into water and o2, white blood cells produce hydrogen peroxide to kill bacteria.
cytoskeleton - actin filaments, intermediate filaments and microtubules.
microscopy - light microscope = 2um, electron =2nm.
light = light focused onto specimen by lenses in condenser, light must be able to pass through specimen, lenses focus the image on the eye. ultimate limit to resolution - wavelength of visible light 400-700nm.
light - condenser lens focuses a cone of light rays onto each point of the specimen, objective lens collects a cone of light rays to create an image. phase -contrast uses interferenace of waves to obtain different degrees of contrast. works by first prsim splitting light into 0 and 90 then second prism combining them, brightening and darkening at diff points depends of path difference. Cell staining reduces amplification of light so structures can be seen.
electron - images closer than 0.2um. uses electrons to illuminate the object as have a smaller wavelength than light, magnetic coils instead of glass lenses to focus beam, specimen in vacuum and very thin on ultramicrotome, pu ton copper grid and stained with heavy metals. picture is collected on photographic plate or digitalised by CCD cameras.
anti-body and imuno-labelling to detect molecules in cells, primary - generated against protein. secondary - conjuagated to flurophore for flurescense microscope and colloidal gold for electron. secondary antibody attaches to primary which is attached to antigen.
epi- flourescence microscopes - detect flourescense emitted by illuminated specimens. diff things have diff excitation and emission wavelengths.
'skeleton' of cell, cell division, intracellular movement and transport, highly dynamic. contraction-muscle contraction move and changes shape as needed. 3 protein filaments: actin (in all), microtubules (in all) and intermediate filaments (not in all) and support for entire tissues.
actin - microvilli, lamelipodia and contractile bundles such as stress fibres. important for rapid reorganisation during mitosis. cytokinesis (division of cytoplasm at end of division. cell polarity and phagocytosis. 20% of mass in skeletal muscle 5% in non muscle. + end barbed = fast growing. - end pointed = slow growing. assembly and dissassmbly for movement enable reorganisation in the cytoskeleton. rapid depolymerisation of actin filaments in response to external stimuli, then assemble at new site. cell migration involes formation of lamelipodia and flipodia (thin stiff projections + end at membrane then adhesion of protrusions to the surface and contraction at rear end. intergrin achorage sites formed at front help to pull cell body forward. actin-myosin interactions result in shortening of actin bundle - rear contraction.
intermediate filaments - non-polar, prominant in cells subjected to mechanical stress e.g skin. nuclear and cytoplasmic. anchored at cell junctions - desmosomes. interconnect cells. keratins - in epithelial, neurofilaments - nerves, nuclear lamins - in all animal nuclei.
cytoskeleton - microtubules
cell motility, intracellular transport, cell shape, division, polarity, organisation and cilia and flagella. organises DNA into daughter cells. hollow tubes alpha and beta tubulin hetrodimers. polar +end fast growing - end slow growing. gamma-TuRC restricted to - end. head to tail binding of hetrodimers form protofilaments.
the centrosome nucleates and organises microtubules. they grow from gamma TuRC ring complexes in centrosome 9 triplets make wall. centrosome usually next to nucleus, - end are anchored to centrosome + end grow and shrink (dynamic instability).
Microtubules transport cargo due to being polarised, axons contain microtubules all - ends towards cell body + end towards terminal. use motor proteins to transport, ATP used for energy. Dyenin and Kinesin are the motor proteins, hold subrstrate and 'walk' them along. can change direction. Kinesin move towards +end, Dyneins move towards - end towards centrosome. both have globular ATP binding motor heads.
cell adhesion molecules hold cells together: glycoproteins with extracellular domain, transmembrane domain and cytoplasmic domain where they are attached to cytoskeleton with linker/adaptor proteins. 3 major families: Cadherins with calcium binding sites; Ig-superfamily (CAMs) related to immunoglobulins as recognise things but not antibodies; slectins with sugars.
Cadherins: most prevalent. homotypic binding of same type of cadherin. Ca dependant interactions - limp without Ca need it to aggregate removal causes dissociation. molecular link between cytoskeletons of neighbouring cells due to anchoring of actin cytoskeleton.
Immunoglobulin-like proteins: Ig like doamins, dont need Ca, homophilic interaction to other N-CAMs. immunoglobulin like repeats, have multiple isoforms due to post translational modification.
Lectin-like CAMs: bind weakly to oligosaccharides ('a few suagrs), heterotypic interactions. weak binding in single molecules, protein which binds to sugar side chains.
stable cell adhesions formed by clusters of multiple receptors. either combination of diff CAMS for specificity and variation. or multiple CAMS for more strength and increased local interactions + with cytosketelon so more signalling.
cell junction 2
epithelial layers - polarization in cell orientation and contacts. have basal side, apical side (contact with luminal spaces) + lateral contacts (cell-cell ) - forms barrier so conc. gradients of ions etc stay same.
cell - cell contacts: tight junctions, adherens, desmosomes and gap junctions. tight junctions - occludin + claudins, define tissue and membrane compartments, tight linkage of membranes, linked to actin cytoskeleton for signalling. Adherin junctions - mediated to cadherins Ca dependant, linked to actin. often for adhesion belt in apical parts of epithelium, can be contractile e.g embryo bone formation, contracts making embryo fold in onself. desmosomes - associated with intermediate filaments, electron dense proteins, add mechanical strength so in stress cells e.g skin. keratin filaments connect cytoskeleton via desmosomes. 3 major protein families - desmosomal caherins, armadillo family, plakin family of cytolinkers.
gap junctions - dimeric complex of connexin hexamers. forms a channel to allow selective transit of molecules. found in most animal tissue. direct connections between cytoplasms of 2 cells. prevent passage of proteins and nucleic acids. enables coupling of metabolic activity, electric response and transfer of signalling molecules. can be openned or closed in response to extracellular signals e.g dopamine.
contribute to structure of tissues, stability, modify reactions of cells, roles during development. 30-50% of all proteins in body are collagen. bones - mechanical stability, cartilage - elasticity, ligaments and tendons - tensile forces, connective tissue in organs - stabilization, basal laminae - separation of tissue compartments. ECM consist of collagens, glycoproteins and proteoglycans. mostly many comlex proteins which can be recognised by cell-surface receptors and interact complexly.
ECM protein synthesis - transription, translation, transport to ER, folding, transport via golgi, modifications, processing, secretion, intergration into ECM and modifications/processing. ECM proteins composed of domains (secondary structure content and hydrophobic core).
collagens contain repetitions of (gly-***-Yyy) sequences. have subunit monomers : left handed alpha chains, have unusual amino acid composition 30% glycine, high amounts of proline. 3 monomers (trimer) assmeble into a right handed triple helix which is unique, homotrimers identical chains, hetrotrimers diff chains. every 3rd aminoacid is a glycine. collagen synthesis = transcription/splicing: nucleus. translation: RER. assembly/modification: ER. processing/transport: golgi. processing/cleavage: membrane. fibril formation: EC space.
collagen and glycoproteins
collagens are post-transcriptionally modified. removal or terminal domains by proteolytic digestion. proline - hydroxyproline. lysine - hydroxylysin. covalent crosslinking. fibril formation has a staggered arrangement with co-ordinated aggregation and a crystal like structure. diseases associated with collagen: ehlers-danlos syndrome (rubber man) mostly mutations in collagen I. hyperelasticity. Osteogenesis imperfecta (brittle bone disease) mutations in collagen I. defects in assembly and degradation causes fragile bones. Scurvy, most common in sailors 19th century, lack of vitimin C, essential for proline hydroxylation, causes unstable collagens and degradation, teeth become loose, blood vessels are weak.
Glycoproteins are highly diverse. attachment of oligosaccharide side chains to either ser, thr, asn. variable oligosaccharide side chains branched or linear, mono or oligo. most secreted proteins are glycosylates. E.g fibronectin, monomer, multiple domains, formation of dimer, diffferential splicing, multiple variants, found in blood plasma, ECM and in tissues bound to cells, multiple functional domains - interaction with ECM comonents with serum isoform; cell-binding activity in the RGD loop in tissue isoform and formation of FN-polymers which are insoluble, covalently linked stretchable networks.
glycoproteins and proteoglycans
laminins - large family of proteins, hetrotrimers, found in basement membranes, basal lamina. functions: self aggregation to form polymers, major cell-binding sites, protein-protein interactions, essential component of all BMs. polymerisation in presence of Ca ions, non-covalent interactions.
proteoglycans. central protein core with glycosaminoglycan side chains and glycosylation (N-linked oligosaccharides). glycosaminoglycans are long unbranched polysacharide chains, repeating units of disaccharides - one sugar amino sugar other uronic acid. GAGs found either linked to proteins e.g keratan sulfate,or not - hyaluronic acid. highly hydrated and occupy large space. resistance to compression, have elasticity and conservation of shape so is movable and effect is enhanced by modifications e.g increased hydration or repulsion by charged groups. e.g perlecan (basla lamina, basement membranes. binding of growth factors and modulation of receptor functions, glycosylation. cell surface proteoglycans, transmembrane domain, interact with growth factors, ECM components, serum proteins, cell adhesion molecules and receptors.
ECM are composed by specialised alloys of multiple components for specific cellular reactions e.g differentiation, migration, survival and cell death and cell-cell interactions.
1) interstitial tissue - contains collagen 1 containing fibrils, elastic fibres. 2) cartilage - elastic matrices which provide stability against pressure and stress. collagen fibrils II, IX and XI, hyaluronic acid and proeoglycan/aggrecan and GAGs, coordination of large amounts of water for elasticity. Aggregates formed by aggrecan and water. 3) bone - stiff aswell as elastic components. collagen I,III and V, mineral deposits and glycoproteins. 4) basement membranes - 2D ECM aggregates, underlying tissues or surrounding cells. functions: compartmentalisation of tissues, barrier functions - inhibit migration, filter, stability, cell adhesion, induce polarisation/ differntiation. laminins, collagen IV and perlecan 5) elastic tissues - skin, blood vessels and lung ECM need to be stable and elastic to provide recoil ability. main component elastin.
ECM in plants - cellulose most abundant for tensile strength, cross linked glycans bound tightly to cellulose. pectin forms networkd for resistance to compression, lamella rich in pectin and cements one cell wall to another. no specific adhesion molecules, essential cell wall for resisiting osmotic pressue, communication by plasmodesmata.
ECM are supramolecular aggregates of a diverse set of specialised proteins outside of cells.
cells and ECM
cell adhesion is essential as need to bind to substrates for - stable anchoring, differentiation, proliferation (growth), migration, signalling and survival. adhesion is dependant on specific ligands/ receptors. cell adhesion is essential for cell spreading which causes flattening of cell bodies and involves actin in cytoskeleton. adhesion is essential for migration on substrate e.g muscle cells (myoblasts), they migrate on laminin but not fibronectin.
cells interact with ECM proteins in matrix. contact with collagen fibrils, fibronectin etc, and modifiy the matrix (degredation). cell-matrix interactions have 2 major structure - motile cells and cells in tissues use focal adhesions. cells of basal laminae and epithelial tissues use focal adhesions and hemidesmosomes. cell-matrix interactions are mediated by specific receptors. 3 major domains: extracellular domain - ligand binding. transmembrance domain - anchoring to cell surface. intracellular - linkage to cytoskeleton and signalling functions.
most important cell-matrix receptor are integrins: specific recognition of ECM proteins, specific adhesion to substraes and induction of signalling. hetrodimers with 2 diff subunits aplha and beta. binding site for ligands. is membrane bound. alpha unit defines specifiicty
cells and ECM 2
integrins - 2 types of binding. both subnits invlolved where ligand binds in cleft. only alpha unit involved and ligand binds to extra domain. integrins have huge family, similar ligands can be recognised by multiple receptors. RGD, collagen and laminin receptors. e.g integrin binding to fibronectin - binding dependant on specific RGD sequence motif. most be exposed and flexible to bind to aV integrin. integrin can be inactive and bent (low affinity) or active and extended (high affinity). interaction of integrin with ligand induces clustering of receptors and organisation of cytoskeleton and clustering of adaptor complexes which mediate contact b/tween cytoskel and receptors.
outside - in signalling where ligand binds to intergrin induces intracellular signalling causes conformational change, effect on adaptor proteins and signalling kinases and cellular response: proliferation, death, survival, gene transcription, differentiation. inside-out signalling crosstalk between integrin-independant signalling pathways which induces changes of ligand binding e.g changes affinity.
adhesion via hemidesomosomes - 'half desmosomes' as real ones dont have integrins. stable connection of epithelial cells to BM. electron dense areas found on basal side of epithelial cells. involve specialised proteins e.g ECM receptors (integrins), adaptors (linking receptors to cytoskeleton) and cytoskeleton (link to the keratin intermediate filaments). defects cause disruption of anchoring cells causing blistering diseases.
cells and ECM 3
other ECM receptors: dystrophin-dystroglycan complex in muscle cells, recognise laminins and are linked to ECM -cytoskeleton. essential connection between BM - laminin and actin. defect cause muscular dystrophy.
cell-surface proteoglycans - membrane-bound proteoglycans e.g syndecan. interact with growth factors, collagens, fibronectin and other ECM proteins. Glycosaminoglycan side chain domain. protein core.
endothelium - leukocyte interactions after inflammation. changes of endothelium ,induction of cell surgace receptors (selectins) ,firm binding of leukocytes via integrins : rolling> binding > migration. leads to extravation.
interaction of cells with ligands: mediated by multiple specific receptors. formation of multi-protein adhesion complexes, mediate linkage to cytoskeleton, induces signalling cascades.
different cell types : contain a variety of recepetor complexs, use combinations of receptor systems, adaption to cellular phenotype and induce diverse signalling cascades.
motility of cells
motor structure - flagellum and cillia. cytoskeletal changes - dependant on contact with surfaces. cilia - can propel single cell though liquid, combined action with multiple cilia, move fluid over surface of cells. vital for sweeping layers of mucus, dust and bacteria up to mouth in respiratory epothelium beats by repeated cycles of power and recovery strokes. Flagellum - found on sperm and protozoa, undulating motoin enbles swimming through liquid. longer - wave like motion from base to tip.
overall organisation - 2 central microtubules and 9 doublet microtubules connected by nexin, spokes nd dynein arms. anchored to cell body by basal body complex of 9x triplet microtubules. motor proteins cause the movement of microtubules. dyneien binds to microtubules which induces relative shift of microtubules. motor proteins: kinesin and dynein. bridging structures limit the sliding of microtubules causing the bending of flagellum. motor protein ciliary dynein generates the relative motion - sliding of MTS. dynein tail attached to MT heads interact with adjacent MT. ATP cause sliding.
movement of multicellular organisms - tissue organisation during development, wound healing (fibroblast migration to form connective tissue), angiogenesis (endothelial cells migrate to form vasculature), immune and inflatmmatory responses ( chemotaxis of leukocytes) and tumour metastasis (migration of tumour cells to surrounding tissues)
motility of cells
migration of cells during embryogenesis depend on - specific signals, attractans, location, cellular status and substances. when tumour cells spread to secondary site through blood stream called metastasis. how to measure cell migration - video time lapse microscopy. directionality of movement - motion is analogous to a diffusion process non-directed migration. chemotaxis - in prescense of a gradient of a 'chemo-attractant' substance. chemo-attractant can be ECM proteins, fragments, growth factors etc. depends on adhesive and stimulatory substrate.
migration is dependant on cytoskeleton and actin, myosin and acessory proteins. migration - formation of protrusions (filopodia and lamelipodia). Adhesion to surface. Pulling forward of cell body. Contraction at read end. migration on surfaces is dependant on substrate recognition. receptor integrins are concentrated at focal contacts which are linked to cytoskeleton which interact with substrate. changes of the actin cytoskeleton where force from contraction is translated to the substrate via focal contacts. focal adhesions - anchor bundled actin filaments at the sites of integrin clustering and ECM contact which are essential.
membrane structure and function
fluid-moissaic. 3 major kinds phospholipids, glycolipids and cholesterol - non covalent so can be fluid. phospholipid - derived from glycerol (3 carbon alchohol) - phosphoglyercides with glycerol backbones and 2 FA chains and a phosphorylated alcohol. spingosine backbone only has 1 FA chain added. double c=c bond important for fluidity.
glycolipids - sugar containing lipids e.g cerebroside - sphingosine backbone, FA unit linked via amide bond, primary hydroxyl group of sphingosine esterified to glucose or galactose. cholesterol - sterol present in eukaryotes but not pro. very important for fluidity, hydrophobic and philic OH end.
thin layer chromotography separates diff lipids based on relative affnities for a stationary phase or mobile phase. polar hydrophillic head and non-polar hydrophobic tails. potential micelle formation but only in lab due to fatty acyl chains too bulky. liposome's are used for drug delivery, have multiple layers. lipid bilayer actually happens - van der waals forces between hydrocarbon chains and electrostatic and hydrogen bonding attractions between polar head groups and water molecules stabilise structure. know this because x-ray diffraction. fluidity controlled by FA composition and cholesterol content. FA chains can be trans (rigid) or gauche (fluid). saturated reduces fluidity can stack, c=c is more fluid. chlolesterol prevents crystallisation of FA chains.
fluorescence microscopy with cell fusion experiments. fluorescence photobleaching recovery technique - bleach labelled lipid or protein and watch recovery of fluorescence. water soluble solutes cannot diffuse across memebrane. urea, glycerol and indole can. permeability can be measured using artificial liposomes and pathclamping where you can measure when ion channels open and close.
specific proteins carry out most membrane processes - transport, enzymatic reactions, recognition and communication. proteins are either peripheral - bound to membrane by electrostatic and hydrogen bnd interactions, or integral - embedded in and spanning bilayer, iteract with hydrocarbon chains in lipid. know this because freeze fracture/ electron microscopy and EM and x-ray crystallography.
because of hydrophobic interior membranes are semi-permeable barriers. selective membrane transport proteins. passive diffusion - gases and small uncharged polar molecules. membrane proteins - active transport and facilitated diffusion (ion channels) transporters, uni, anti and sym. ATP powered pumps: P class (H and Na/K), V-class (protons vascular membrane and endosomal and lysosomal), F-class (protons bacterial plasma membrane + inner mitochondrial membrane), ABC superfamily (small molecules)
uniporter e.g GLUT1 transports glucose into most mammalian cells, 2 comformational states. coupled transport symport - e.g 2Na/glucose. antiport e.g 3Na/1Ca. can use ihibitor of H/K ATPase to treat stomach ulcers. Inhibit kidney channel proteins - control hypertension. calcium channel blockers - control heart contraction. brain Na channel mutation - epilepsy. CFTR - cystic fibrosis.
hypo-osmotic medium liposomes swell. shrink in hyper-osmotic medium. aquaporin channels.
nuclear membranes and ER may have evolved through invagination of the plasma membrane. movement of proteins in cell - 1 transport through nuclear pores. 2 transport across membranes. 3 transport by vesicles. all require ATP and the protein remains folded in 1&3 but needs to be unfolded in 2. signal sequences (usually 15-60 amino acids) direct proteins to correct compartments e.g cytoplsmic proteins - default so none. secreted proteins - have ER import signal sequence. Cell surface membrane - ER import plus transmembrane domain. Nuclear proteins - nuclear localisation sequence.
nuclear envelope has 2 membranes, inner rests on nuclear lamina and outer is continious with ER. Nuclear pores comprises nuclear pore complex - allow passive diffusion of proteins, import of larger proteins require nuclear localisation signals. mechanism involves docking of cargo with specfifc importins that interact with NPC. similar mechaism exists for export -exportins which require nuclear export signals.
for proteins that are destined for ER, signal sequence and signal recognition protein directs the polypeptide from ribosome to ER. SRP binds to channel on ER and is passed onto translocation chanel to be recylced. protein then crosses ER membrane to ER lumen and signal peptide is cleaved by signal peptidase and degraded. when protein has second transmembrane domain sequence it gets 'stuck' in the membrane.
when intracellular material is moved between compartments, vesciles bud from one membrane and fuse with the other carry membrane components and protein contents, each compartment has an inner lumen topologically equivalent to the outside of cell. communication ebtween compartments is via vesicles (secretatroy vesciles, endosomes and lysosomes). Clathrin molecules form basketliek cages that shape the membrane into vesciles via endocytosis. also helps cpture molecules for onward transport to other compartments. adaptin forms links with clatherin with binds to cargo receptors and cargo to form vesicles, dynamin GTPase pinches off vesicle, vesicle is uncoated and fuses with membrane.
budding vesicles carry specific markers called vesicle v-SNAREs that bind to complementary target t-SNAREs on target membranes to help direct them. some vesicles undergo exocytosis as most secreted and transmembrane proteins are modified in ER/golgi first e.g glycosylation (post-translational modification). in specialised secretory cells, regulated and constitutive pathways of exocytosis diverge in the trans golgi network e.g beta-cells in pancreas. endocytosed material is delivered to endosom and if not recycled to cell surface, sent to lysosome for degredation. lysosomes main site of intracellular digestion containing hydrolytic enzymes and H pump to maintain acidic conditions.
meiosis - gives rise to egg and sperm, pre-meitotic cells are diploid (2n), starts with 1 round of DNA replication(4n) followed by 2 seperate cell divisions to give 4 haploid(1n) cells. mitosis - in diploid somatic cells ensure genetic identity of equal daughter cells. process simillar but only 1 round of DNA synthesis.
mitosis - mitotic phase of nuclear division followed by cytoplasmic division (cytokinesis). cell cycle - G1 (organelle duplication), S (DNA synthesis), G2 (cells expand and prepare for M), M phase (prophase, metaphase, anaphase, telophase and cytokinesis). essential processes of cell cycle (DNA replication, mitosis and cytokinesis) are triggered by cell cycle control systems - protonca genes and tumour suppressor genes. have multiple check points before each stage e.g is cell big enough, is environment favourable, is all DNA replicated.
CDK cyclin dependant kinases- enzyme accelerator for cell cycle. CDKI - inhibitor enzyme, stops cell cycle. Cyclin - protein affects activity of engine. enzymatic activity controlled by cdk phosphorylation cyclin binding.
phases of mitosis - prophase chromatin condenses, centrioles divide and migrate, kinetochore, and fibres and spindles form. prometaphase - nuclear envelope breaks down
cell division 2
metaphase - chromosomes are randomly places in motion but come to lie halfway between spindle poles in metaphase plate. Anaphase - paired chromosomes separate at their kinetochores and start to move along MTs to opposite poles. Telophase - daughter chromosomes arrive at poles, new nuclear envelope forms. actin and myosin filaments form contractile ring. microtubules form mitotic spindles.
The centrosome in an interphase cell duplicates to form the 2 poles - early prophase. 3 classes of microtubules make up mitotic spindles - aster microtubules which attach to spindle poles, kinetochore microtubules which physically attach to kinetochore, and interpolar microtubules wchih overlap kinesin and holds them together to provide extra push.
2 processes separate sister chromatids at anaphase. anaphase 1 - chromosomes are pulled poleward, shortening of kinetochore microtubules. anaphase 2 - poles are pushed and pulled apart, sliding force generated between interpole microtubules and pulling force directly on poles to move them apart.
cell cycle controlled by protein kinases composed of regulatory subunit (cyclin) and catalytic subunit (cdk) cyclin levels rise and fall during cell cycle.
hormone signalling - singalling cells - bloodstream- traget cell - receptor - response). diverse can be amino acids e.g adrenalin, polypeptides e.g EGF, insulin, derviatives of cholesterol (steroids) e.g oestrogen, modified FAs (ecisanoids) e.g protaglandins. bind to specific receptors on target cells, and evoke a response through signal transduction. control cell growth and differentiation (EGF), wound healing (platelet-dervived GF), response to infection (interleukins), electrical and mechaincal functions supporting locomotion (acetylcholine), digestion of food and nutritional support for tissues (insulin).
endocrine signalling - tissue A - bloodstream - tissue B . Pacrine signalling - not through blood to adjacent cell. Autocrine signalling - target sites on same cell, instructs self to grow/divide. Synaptic signalling - along a synapse. Signalling by plasma membrane attached proteins - membrane bound ligand binds directly to membrance recptor on adjacent cell.
contact dependant signalling controls nerve cell production. same signalling molecule can induce diff responses in diff target cells. e.g acetylcholine in heart, salivary gland and muscle cell. hormones can be classified based on their solubility and receptor location in cell. hydrophilic signal moleucle binds to cell surface receptor as cant get through membrane. Small hydrophobic signal molecule e.g steriods get to intracellular receptor.
glucose: complete oxidation to co2 and h20 proves -2,840kj/mol. high solubility and can be stored as high molecular weight polymer. can be transferred to storage as glycogen, starch and sucrose. can synthesise strcutural polymers e.g ECM and cell wall polysaccharides. oxidation via pentose phosphate pathway to ribose 5-phosphate for RNA and DNA synthesis. can by oxidised to pyruvate via glycolysis.
tumour cells are anaerobic so generate less ATP per glucose molecule, but tumour cells consume a lot of ATP and have high rates of glucose usage which can be used as a marker. Patients are fed fdG which reacts with hexokinase and traps flourescence in cells. and positron emission tomogrpahy (PET) identifies tissues which flouresce and therefore use a lot of glucose.
glycolysis is catabolic, gluconeogenesis is anabolic. not exact reversable as 3 steps of glycolysis have large -G so are irreversible. glucose+ ATP - glucose 6-phosphate+ ADP. Fructose 6 phosphate+ATP - fructose 1,6 bisphosphate+ADP. and phosphoenolpryuvate +ADP - pyruvate +ATP. PEP - pyruvate in glycolysis uses 2ATP and one enzyme pyruvate kinase. Pyruvate - PEP uses 2 ATP and 2 GTP and goes to oxaloacetate then PEP using pyruvate carboxylate and PEP carboxylate. these enzymes are in diff cellular compartments as NADH is lower in cystol.
pyruvate carboxylase and PEP carboxykinase provide bypass reactions. The other bypass reactions are catalysed by FBPase-1 and glucose-6- phosphatase which dont use up high-energy molecules. synthesis of glucose from pyruvate is energetically expensive uses 4ATP, 2GTP, 2NADH, 2H and 4H20. is required to ensure irreversibility of gluconeogenesis.
Enzymes are regulated using allosteric mechanisms to prevent both pathways working at the same time. gluconeogenesis in liver, kidney and small intestine to provide glucose for brain, muscles and erthrocytes.
pentose phosphate pathway - in the oxidative pathway, NADP is the electron acceptor. Major product of pentose-5-phosphate is RNA, DNA and coenzymes ATP, NADH, FADH and coenzyme A. important for rapidly dividing cells e.g bone marrow, skin, intestinal mucus and tumours. glucose 6 phosphate - 6-phosphogluconate - ribulose 5 phosphate - ribose 5 phosphate - nucleotides, RNA etc. 2 NADPH molecules produced and co2.
oxidation of fatty acids and amino acids
FA provide a lot of energy 80% in heart and liver. During oxidation of FA acetyl-coA is produced in the same 4-step B-oxidation. processing dietary lipids - bile salts in small intestine emulsify fat forming micelles. Intestinal lipases degrade triacylglyerols, FA are taken up by the intestinal mucosa amd converted to triacylglycerols. Triacylglycerols are incorportated with cholesterol and apolipoproteins into chlyomicrons. they move through lymphatic system to tissue. Lipoprotein lipases activated by apoC-II in capillary convert to FA and glycerol. FA enter cells and oxidised as fuel or reesterfied for storage.
low levels of glucose trigger glucagon to be released which releases cAMP in cells which activates lipase to release FAs from triacylglycerol to blood stream where B-oxidation happens and energy is released. The released glycerol is processed via glycolysis. fatty acyl-coAs are high energy compounds-hydrolysis to FA and CoA has large negative free-energy charges. Fatty acyl-coA esters formed in cystol can be used to synthesise membrane lipids. also transported into the mitochondiron and oxidised to produce ATP.
fatty acid oxidation = stage 1 - Long chain FA oxidised to give acetyl-CoA - process termed B-oxidation stage 2 - acetyl groups oxidised to co2 via citric acid cycle. Stage 3 - Electrons from stages 1 and 2 pass to o2 via respiratory chain, generating ATP.
fatty acid oxidation
B-oxidation - 1. dehydrogenation by FAD, oxidation by acyl-CoA dehydrogenase which forms double bond between C2 and C3. 2. hydration of the bond to form L-3-hydroxyacl CoA by enoyl CoA hydratase. 3. oxidation by NAD. converts hydroxyl group to keto group by L-3-hydroxyacl CoA dehydrogenase to form B-ketoacyl Coa. 4. Thiolysis. cleavage by thiol group of another coenzyme A molecule. acyl-CoA and acetyl-CoA are formed by B-ketothiolase. additional enzyme steps if FA is unsaturated (2) or has an odd number of C's (3). B-oxidation removes each acetly-CoA.
genetic defects in FA oxidation. muscle contraction suffers as triacylglycerols are cheif source of energy. mutation in gene encoding medium chain acyl-coa dehydrogenase (MCAD) if recognised early infants can be fed low fat high carb diet.
amino acid oxidation - occurs during normal synthesis and degredation of cellular proteins. When diet is protein rich and ingested amino acid exceeds need. During starvation or uncontrolled diabetes. important feature of AA degredation is to separate a-amino group from carbon skeleton. excess NH4 is excreted as ammonia in aquatic vertebrates, bony fish or amphibian larvae, urea terrestial vertebrates or uric acid birds and reptiles. carbon atoms in urea and uric acid are highly oxidised - most of available energy has been extracted
amino acid oxidation
amino groups are removed from glutamate to prepare them for excretion. Glutamate dehydrogenase converts molecule to a-ketoglutarate, which can feed into citric acid cycle or glucose synthesis. this releases NH4 which is transported to liver as amide nitrogen of glutamateor alanine (in skeletal muscle). Transport of ammonia must be carefully regulated as is toxic to brain. occurs mainly in mitochondira of hepatocytes and urea passes into blood and excreted through kidneys into urine.
carbon skeletons undergo oxidation to compounds that can be converted into either glucose or ketone bodies. amino acids are grouped into either glucogenic or ketogenic groups.
some serious human diseases can be traced to genetic defects in enzymes of amino acid catabolism. e.g albinism, mental retardation, vomiting/concussion, faulty bone development.
from lipids to hormones
cellular roles of lipids - store energy, major constituent of cellular membranes, sepcialised - pigments, cofactors, detergents, transporters, hormones, extra-intracellular messengers and anchors for membrane proteins. lipids are metabolised in mitochondria, ER and cystol, chloroplasts and perixosomes. Peptide and amine hormones act faster than steroid and thyroid hormones. when a single peptide hormones binds many molecules of 2nd messenger are produced.
hormonal regulation of metabolism - biosynthesis and degredation of triacylglycerols is regulated to take into account metabolic resources and requirements and rate is altered by several hormones e.g insulin. which stimulates vonversion of dietary carbs and proteins to fat. glucose and amino acids to acetly-coa - FA - triacylglycerols and ketone bodies are produced. ketone bodies - acetone, acetoacetic acid and beta-hydroxypentanoate are by-products when FAs are broken down by liver and are used in brain during fasting and muscle to produce atp and have distinguished smell do can diagnose diabetes. uncontrolled diabetes results in diminished FA synthesis and higher production of ketone bodies. Conc of glucose in blood plamse should be 60-90mg/100ml. <40 constitutes sevre hypoglycemia.
lipids to hormones
when well fed - pancreas releases insulin - glucose in brain to ATP and CO2. glucose in liver released and turned to glycogen. glucose - pyruvate - acetyl-coA to ATP and CO2 in cyclic cycle. TAG in adipose tissue and lymphatic system. FAs in muscle to ATP and CO2. Amino acids moved through blood vessels form intestine to liver where urea is produced and protein synthesis occurs.
when fasting - glucagon produced by pancreas - liver when gluconeognesis occurs. Produced glucose and ketone bodies given to brain which produces ATP and CO2. TAGs in adipose tissue break down to glycerol and FAs. FAs to ATP, ketone bodies and CO2. Glycerol, pyruvate and glycogen go from glucose - 6- phosphate to glucose. in muscle - ketone bodies broken down to ATP and CO2 and proteins are broken down in liver to amino acids and pyruvate.
metabolism under control = pathways are unidirectional, though individual steps may be reversable. Net G of any metabolic pathway is negative under physiological conditions. Cells have evolved mecahisms that couple unfavourable reactions to strongly exergonic processes. cells have to do 'work; so combined processes are exergonic overall. energy from breakdown of biomolecules used to provide energy for biosynthesis of others.
ATP catabolic reaction is exergonic where ATP is made from stored nutrients, ingested food and solar photons. Then broken down in anabolic reaction (endergonic) where complex biomolecules are synthesised and need energy from ATP. also sed in other cellular work, mechanical work and osmotic work. Levels of ATP and AMP are senstive indicators of cells energy status. during anabolic pathways ATP is used up and most forms AMP. 10% decrease in ATP conc to 600% increase in AMP. nearly constant levels of ATP and NADH maintained in cells and glucose in blood.
Intermediary (central) metabolism refers to the combined activities of all pathways that interconvert precurrsors and molecules of low molecular weight. metabolic pathways that are fundamental to cells are relatviely few. Close relationship between metabolism and bioenergetics. Enzymes are regulated using allosteric mechanisms to prevent both pathways working at same time called futile cycles.
major principles - stop futile cylces to maximise efficiency of fuel utilization. Partition metabolites between alternative oathways. Shut down biosynthetic pathways when products build up. changes in metabolite conc will lead to altered enzyme activity. In multi-step processes majority of regulation is at reactions that arent equilibriums. In humans hormones regulate metabolism.
cell communication 2
relay of information across the plasma membrane to cell interior and nucles can cause - activation of enyme, change in cytoskeletal organisation, change in ion permeability, initiation of DNA synthesis in nucleus - cell division, actication of specific gene transcription in nucleus - transcription factors or cell death. hormonal effects are mediated by secondary messengers e.g cAMP, cGMP and calcium ions or kinases/phosphatases which can also act as signalling 'switch' mechainsms e.g kinase activates, phosphatase inactivates.
3 main types of cell surface receptors - ion channel linked e.g neurotransmittors, G-protein linked e.g adrenalin, glucagon, catalytic or enzyme linked e.g insulin and EGF. one group is intracellular/nuclear - nuclear hormone receptors e.g oestrogen. enzyme linked - receptor has intrinsic enzyme domain such as kinase/phosphatase. enzyme is activated by ligand binding. e.g receptor tyrosine kinase. receptor activation dimerisation and trans-autophophorylation - phosphorylation and recruitment of intracellular substrates - transfer of signal thorugh intermediate protein - cascade - activation of rtanscription factors - specific changes in gene expression.
amplification of signals occurs by an intracelular cascade mecahinsm.
plant progenitor - autotrophic (photosynthesise to make own carbon source) and have cell wall. endosymbiont theory for origin of mitochondria and chloroplasts - many single-celled eukaryotes host symbiotic bacteria, M and C are similar size to bacteria and are similar in structure and composition to bacterial ribosomes and are sensitive to similar antibiotic drugs.
arabidopsis is a model organism in plant biology because : member of brassicacea family, smallest plant genome, short life cycle, controlled crossing, produces large nmbers of seeds, molecular and genetic maps, large mutant pop, transformation tecnology. can investigate gene expression in A/thailana by seeing which is expressed in white light and dark grown seedlings by probing with green and red fluorescent dye. rice is another model organism as most important cereal crop, stable food for 3b people, diploid genes.
plant life cycle - seed - embryogenesis - seed development (germination) - mature plant sporophyte - flower - petals produce carpels and stamens. stamens produce pollen grains (male gametophytes), carpels produce ovule with eff cell and embryo sac (female gametophyte) - come together in pollination to produce seeds. plant embryogenesis - establishes root and shoort polarity (primary meristems), estabilishes radial tissue patterning, embryo is determinate whereas subse
plant embryogenesis - establishes root and shoort polarity (primary meristems), estabilishes radial tissue patterning, embryo is determinate whereas subsequent development is indeterminate and cell fate is deterined by position rather than cell lineage (cell extrinsic information). whereas animal embryos - mophogenic changes confined to a breif embryonic phase, embryogensis generates rudimentary minature scale models of adults and lineage and mobility important in determining and maintiaing cell fate (cell intrinsic information).
2 cell stage - basal cell and terminal cell. octant stage - apical cells, central cells, hypophysis, suspensor, basal cell of suspensor. heart stage - shoot apical meristem. Early seedling - cotyledons, embryonic root, columella root cap.
plant anatomy - leaves, stem and root. all cells originate from meristems that generate 3 major tissue systems - dermal, ground and vascualr tissue. embryo is determinate, subsequent development is indeterminate. cell fate determined by position not cell linage. cell fate determined by transcription factors that regulate control of specific genes. Plant cells are totipotent. defined plane of cell division gives rise to specific cell files.
plant cell 3
meristemic activity in roots produces longitudinal files of cells. maturation zone - division and elongation ceased, differentiation has completed, radial pattern obvious. elongatoin zone - region of cell elongation, little cell division. meristemic zone - behind root cap, produces root only, no lateral apendages, max rate of cell divis. root cap - protects apical meristem, perceives gravity, secretes mucopolysaccharides.
DNA in nucleus, chloroplasts and mitochondria. plant nuclear genomes have introns. mRNA requires extensive post-transcriptional processing - capping, polyadenylation and splicing. leaves originate from primordia generated on the flanks of SAM. palisade and mesophyll cells contain chloroplasts. photosynthesis - plants convert solar energy to chemical energy in chloroplasts. stroma contain enzymes that catalize C fixation (light independant reaction). Thylakoid membrane contain intergral membrane proteins of the light dependant reaction - light absorbed by PSII produces strong oxidant and weak reductant electron passed to next acceptor. Strong PSII oxidant oxidises water (regains lost electron). Light absorbed by PSI produces weak oxidant and strong reducatant which reduces NADP to NADPH.
adjacent cell walls are cemented through the middle lamella. cellulose microfibrils are synthesised at cell surface. hemicelluloses and pectin are synthesised in golgi and desposited at cell surface. CMS are coated with hemicelluloses. Pectin control wall porosity
plant cell communication
the vacuole membrane is tonoplast. plant cells are connected through plasmodesmata. makes them symplastic as can mix cytoplasms. ER continuous between cells. spoke proteins regulate size exclusion limit between cells - can be varied to allow passage of larger molecules e.g RNA and virus'. plants communicate over long distances using vascular system. signalling molecules include RNA and proteins. Seive element cells in phloem lack nuclei and plasma membrane is continuous between neighborouing cells. phloem is living system that is primary mechanism for comunication over long distances.
xylem is dead cells, everything inside was digested apart from cell wall due to 'programmed cell death'. tracheids and vessel elements contain extensive secondary cell walls (lignin). unidirectional movement unlike phloem. e.g abiotic signalling - nutrient deficiency in roots generates signal - activates response in leaves - adjusts genetic programme controlling nutrient uptake.
vegetative meristems stem cells divide slowly. stem cells divide more rapidly in floral meristem to generate distinct organ types. aradopsis flowers have 4 distinct organ whorls. 4 sepals green at maturity, 4 petal (white), 6 stamens (4 long 2 short) and gynoecium (ovary with 2 carpels, style and stigma.
plant cell communication 2
positive and negative regulatory pathways controlling flowering in arabidopsis. FT forms complex with transcription factor FD. giberellin pathway. energy pathway with sucrose. low temp and leaf number in autonomous and vernalization pathways and light pathway. vernalization - cold treatment to hydrated seeds promotes flowering. makes SAM competant to undergo transition to a floral meristem. requires active metabolism, DNA replication and cell division. blocks expression of flowering locus C by an epigenetic mechanism.
leaves generate signal to induce flowering - 'florigen' production. plants flower under short day conditions, SD ;leaf grafted onto LD plant can induce flowering. SD leaf re-gradted induces. LD leaves from plants in flower do not. Floral apex grafted onto non-induced plant doenst either.
mobile signalling promoting floweing may be an mRNA molecule encoding FT. CO protein accumulation in the phloem during light is required for flowering. CO expression is controlled by circadian clock. Co encodes a transcription factor of many genes including FT and AGL20. FT mRNA is translated at the SAM and interacts with a second protein FD which activate floral meristem identity genes. FT and/or FTmRNA are transported through phloem. FT in SAM interact w/ FD to trigger Interflurorescent Meristem formation.
plant cell communication 3
proteins that are translocated through plasmodesmata are called NCAP - non cell-autonomous proteins. RNa molecules can also move through plasmodesmata and are mediated by rna biding proteins and specific chaperones. e.g the KNOTTED1 transcription factor. is a NCAP that can move between cell layers in the leaf controls cell fate in SAM. KNOTTED1 mRNa only detected in L2 layer in monocots and L2 and L3 in dicots but the protein is in all 3. KNOTTED1 activity is depednant on binding its mRNA to form a ribonucleoprotein complex. the RNP is translocated through PD which activates KNOTTED1 post-tranlationally. Which activates its target gene.
chaperones also assist non-cell autonomous proteins to PD docking sites which induces dilation of PD channels allowing them through. e.g genetic control of root development. mutations in SHR and SCR genes result in slow growing roots that lack distinct endodermins and cortex. both genes encode GRAS transcription factors. SHR stimulates SCR expression. further illustrates importance of intracellular communication in determining plant cell fate.
root apical meristem also comprises of slow dividing stem cells, plane of division is strictly controlled (anticlinal-periclinal-anticlinal) mutations in SCR and SHR stop periclinal divis.
cell communication 4
can anaylse SHR gene transcription and protein localisation using GFP fused with promotorSHR and SHR to construct recombinant genes. shows SHR is transcribed in cells of vascular cyclinder and shows protein is also located in ajacent endodermal cell layer and quiescent center. the SCR was fused with GFP and showed the protein was expressed in a single cell layer in root - the endodermis and was dependant on SHR expression. shows importance of extrinsic information in determining plant cell fate.
models for the role of SHR in endodermal cell fate - SHR synthesised in stele cell moves in both directions across nuclear pore complex. Post translational modifications of SHR when it enters the endodermal cell prevents further transport through PD. SHR activation of SCR initiates endodermal cell development.