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BIOL 213: EXAM 2
extracellular matrix, what is the extracellular space |
it influences how cell interacts w/ other cells & its environment, applies to mostly animal cells and some protists that have extract. components but plant cells & fungi have cell walls, which is a box |
what are the types in the extracellular matrix |
glycocalyx (just out of membrane), and extracellular matrix (ECM, farther out of membrane) an example is basal lamina boundary between different tissues or types of cells |
glycocalyx |
associated with cell surface in animals and some protists, short carbohydrate chains, include cell coat (carbohydrates attached to proteins on outer membrane surface, trans membrane proteins), and fuzzy layer ( carbohydrates attached to cell coat by weak molecular interactions), rubinian stains shows glycocalyx, microvilli increase SA, microfilaments in cytosol reinforcement on plasma membrane |
glycocalyx functions |
can be removed w/out permemant damage to cell, if removed cell will regenerate it, mediates cell-cell substrate recognition/interactions, mechanical protection for surface, surface barrier (directs cell response and reinforce plasma membrane coat) |
ECM (extracellular membrane) |
its an organized 3D network of extracellular material beyond immediate vicinity of the membrane, more than packing material can be major influence on cell shape & activity, can be highly structured or loosely organized, (ex. basal lamina) ( connective tissue has small volume with cells, its more extra.cell. material like in ears or nose), (in dermis has collagen, fiber blast is like slug secrete collagen and creates densely packed), (chondrocytes lay down dense extra,cell. matrix has proteoglycans (nose, ears, makes cartilage) |
ECM charactertistics |
non living secreted, proteins not globular but fibrous, prominent in connective tissue , cartilage, bones, tendons etc, cell occupy small volume of tissues, it determines tissue characteristics, analogies ( scaffolds, girders, glue, wires, springs etc) |
basal lamina (basement membrane) |
specialized form of ECM, sheet tightly packed, under basal layer of epithial tissue (skin, digestive systems lining, respiration system) and inner lining of blood vessels |
basal lamina functions |
mechanical support cells, cell polarity (basal (bottom) to apical (top)), substrate for cell mitosis & cell migration /path, compartmentalizes from adjacent tissues, barrier to macromolecules & some cells, simples structured etc, inside out, up down |
what happens if cell are in touch with the basal lamina |
if cells touch it can divide if not touching than cannot, epithilial cells are stuck to the lamina ( epidermis then lamina then dermis) |
what are the molecular components of the ECM |
examples are collagens, proteoglycans, and (fibronectin, laminin, other ECM proteins) |
collagens |
large family of proteins atleast 19 different types, high tensile strength, fibrous proteins, insoluble makes an animal an animal, produced mostly by fibroblasts, smooth muscles cells and epithelial cells , common structural features are all trimers (3 polypeptides in alpha helix in 4 degree array), 3 chains wound around each other to make rodlike triple helix (side by side), collagen is leather, skin, tough durable material its highly processed and in jello, well organized tight arrays |
collagen properties |
insoluble framework determines matrix 3D organization and mech. properties of tissue, tendons (fibrous collagen type 1,2,3, assemble into rigid parallel cables, cornea (short fibrous collagens arranged into precise arrays to be tough but transparent), basal lamina (type 4 collagen non fibrillar arranged into thin flexible sheets,cross links made b/c damaged covalent bonds by ex. UV light (older you are more you have, causes wrinkles) smooth skin means not many cross links |
cornea transplant |
if cornea infected then white blood cells get into it and kill infection but they stay on cornea and cause cloudyness b/c collagen just gets packed on top of one another instead of being well organized so have to get transplant, fibroblasts can be seen at 1 mm they crawl around and leave collagen, big cables are type 4, type9 stick end to end or to others on their sides |
proteoglycans |
many ECMs contain proteogylcans (proteins with polysaccharides associated), each protein molecules has short polysaccharides glycosaminoglycans (GAGs) attached along its length, hydrophilic 3D volume like sponge, resist compression, GAGs contain modified amino sugars & sulfate sugars, protein/polysaccharide chains are attached to a polysaccharide core ( hyaluronic acid) to make huge gel like hydrophilic complex, proteoglycans are caterpillar like make cartilage, (like lobe of ear & shell is proteoglycans) |
fibronectin |
large protein w/ modular construction of specific domain shapes (domains 5 or 6 functional units per molecules); each unit has specific binding function for cells & other ECM components; specific fibronectin provide markers/pathways for specific cells in development (ex migration paths defined for certain cell types) |
what do different fibronectin have |
they have different sequences in domain and have cell binding domain (RGD loop) is what cell recognized on fibronectin; provide cross linking and info for cell;marker for fibronectin around connective tissue; cells in between fibronectin (they like it there)(fibronectin provide linkage, part binds to collagen and other part forms attachment site for cell) |
Laminins |
am of extract. gylcoproteins; 3 polypeptides covalently connected by disulfide bonds into cross shape; make molecular markers for cell in development migration, growth, and differentiation; bind lightly to other ECM components (ex contributes to strength of basal lamina); cross link and bind to each other and molecules can sit on them |
Other ECM molecules |
less common specialized components for certain specific ECM types |
cell detection of ECM |
cell needs surface receptors to detect differential signals of ECM; receptors are integrins cell can't detect if doesn't have receptor[ lrg fam of proteins on surface, heterodimer(are of alpha polypeptide and beta polypeptide held together by nonequivalent links); 18 dif. alpha types, 8 dif beta types (24 dif integrins) , need Ca ions to work, each subunit spans plasma membrane] |
integrin functions |
adhesion of cells to ECM or to other cells; transmem proteins, active only when bound w/ divalent cations and dimerized; signal transmission from exterior to interior, signal transmitted through mem. by conformational change of integrin; signal may affect cell motility, growth and survival; binds w/ whatever there is shape change on cytosolic side, cell stuck on surface by integrins cell behavior changes motile or non motile etc |
how does a cell move |
it is like a rock climber it anchors itself then feels around with the front then if it finds somewhere it likes it will unanchor itself and move forward and repeat the process ( integrins bind to surface and reinforce it then feel around
|
cell-cell interactions |
wilson sponge sorting experiments; steinberg embryo tissue sorting hierarchy, urchin mesenchyme cell adhesivity; four fams of integral mem proteins [ selections, certain immunoglobulins, certain integrins, cadherins |
wilson sponge sorting experiment |
work with sponge on cross sections & pores where water comes in b/c filter feeds, sponge flagellated feeding cell inside sponge (choanocytes), epithelial cells on outside, amoebocytes (middle make body of sponge), wilson took sponge, sift & mashed sponge thru it made lil pieces, sit overnight in seawater & in morning saw lil sponges, cells knew where to be relative to each other, had two species of sponge (red & yellow) mixed them & got lil red and yellow sponges (knew who they are & where to be) cell-cell interact. |
hulterfreter and steinberg embryo tissue sorting hierarchy |
chick embryos, used enzymes and techniques, mixed dif cells from chick embryo and found they found their spots, cells interact w/each other ,cells know who to interact with and who not to & know where to be |
urchin mesenchyme cell adhesivity |
blastula have micromeres (lines on blastula) and gastrula have mesenchyme; micromeres turn into mesenchymes when the cell starts gastrulation; embryo start to become more animal like; micromeres jump out of association and crawl around hollow part and lay down ECM (skeleton, mesenchyme do it), (micromeres stick tightly to neighbors) mesenchyme detached to everyone,when cell changes cell to cell recognition, its behavior changes, it depends on proteins (integral membrane proteins) |
selectins |
integral membrane glycoproteins; bind to specifc sugar oligosaccharide sequence on other cells surface, they are a type of lectin general class of protein that bind to specific oligosach., large extracell. domain span membrane and small cytoplasmic tail , three types E(endothelial cells) ,L,(leukocytes),P (platelets); binding of selecting to specific oligosach. require Ca ions |
how to selectins help in removal of bacteria |
white blood cell roll across epithelial cells and can go thru it with a signal if there is bacteria, selecting bind to sugar on white blood cell and changes it behavior and sticks tightly to epithelial cell and slip thru into tissue and kills bacteria; called leukocyte attraction and diapedesis |
immunoglobulins |
on lymphocytes, integral proteins, involved in immune function/recognition, but some involved in Ca-independent cell to cell adhesion(they stick to other immunoglobulin on another cell)( need Ca ions) ; non immune cells can use this
V-CAM (vascular cell adhesion molecules),N-CAM(neural cell adhesion molecules),L1 (non-immune cell adhesion)
[loops on them can bind to other immunoglobulin proteins, cell to cell] |
integrins |
most included in adhesion of cells to ECM, few in cell-cell adhesion, cooperate with immunoglobulins in leukocytes and neuron binding, Ca dependent for binding |
Cadherins |
atleast 30 related glycoproteins, calcium dependent cell adhesions, cell express some cadherin bind lightly to each other; three types E (epithelial), P-(placental),N-(neural), tied to cytoskeleton on cytoplasmic face by cattiness proteins ( less than .05mm Ca ions than not stiff enough, then add a lil dimerize, more than 1 mm of Ca ions will bind to each other (same kind of cadherin)) |
what do each integral membrane bind to |
... |
plant cell walls |
arrangement of cellulose fibrils parallel to arrays of mt under plasma mem.; cellulose synthesizing enzyme complexes (rosettes) oriented by mt, reorganization of mt arrays lead to reorientation of cellulose fibrils, alternating layer of cell fibrils, cell fibril parallel, cytoskeleton in cell is same orientation (parallel) which is mt, mt anchored in plasma mem. , rosette put to cellulose constrained by mt; cell grows by stretching parallel bundles of cellulose , can lay cellulose in between [mt- microtubules] |
cytoskeleton |
filaments that form elaborate 3D structures; microtubules (mt, hollow rigid cylindrical tubes, walls made of tublin), microfilaments (mf,solid thinner structures made of actin), intermediate filaments(if, tough ropelike fibers made of various protein types), cortex ( structured viscous region of cytoplasm under plasma mem., reflects degree of organization of cytoskele. assoc. w/surface of cell esp. mt & mf,; highly struct. cytoplasm reinforcer so cytoplasm doesn't tear; plant and fungi don't have it |
cytoskeleton function |
structure support, intercellular transport, contractility and motility, spatial organization, cytoskeleton built on edge of advancing side, scaffold ( support maintains shape including cortex, internal framework( puts organelles in specific patterns), mRNA moved to sites for translation, endomembrane vesicle transport, NT vesicle movement to synapses, move cell (flagella and cilia), chromosome movement in mitosis and meiosis |
study of cytoskeleton |
microscope needed, fluorescence microscopy (labeled antibodies, covalently linked dyes on molecules, hybrid protein genes w/ flourescent tags(GFP)), video microscopy ( electronic enhancement of image in vitro motility assays, genetic engineering techniques, electron microscope technique |
microtubules (MT) |
in almost all eukaryotic cells, wall made of cylindrical array of 13 linear filaments of globular proteins (protofilaments) [hollow tube stiff can be long, made of tubular protein dimers (alpha, beta bond) together by end to end to build filament], tubulin are GTP binding proteins , tubulin in parallel orientation so has polarity (+ & - ends,beta (+)on 1 end and alpha(-) on other); usually have one end attached to centrosome (mt organizing center) |
where are new tubulin added on mt |
on alpha(-) end tubulin is added slowly (slow growing or are being pulled off here); on beta (+) end tubulin is added fast (fast growing)
GTP in beta end regulates activity |
tubulin dimer |
occupy 8 nm length together (one beta to other), two places tubulin dumpers can be ( tubulin assemble mt, can be in cytosol (floating not assembled) but when beta has GTO it binds with alpha and beta and makes mt (assemble, adds to existing ends)); GDP sticks to other tubulin dumers but not so strong so it falls apart, so in cytosol it gets rid of GDP and gets GTP instead |
mt associated protein (MAPs) |
found on mts isolated in many cells types, globular heads attached to tubulin, filamentous tail extends out, may interconnect mts (cross bridge) stabilize mts or influence assembly rates, bind to mt and say if assembly, disassemble or if can attach other stuff, tails of MAP proteins stick to each other, can extend half life of mt, (ex MAP2 protein globular head binds to tubulin and has long tail) MAP proteins have different effects on mt |
function of mt |
internal scaffold for structural support, resists compression or bending, support cells; cell polarity (est different structures end to end),determine cell shape ( elongated cell processes (ex. axon) supported by mt arrays), plants( array of mt under plasma mem influence organization of cellulose fibrils in cell wall, cel internal organization, motility (flagella and cilia movement) spindle apparatus ( chromosome movement), centrosome (where all mts all go) |
mt motors and motility |
mt dont generate force but molecules associated with them do (railroad, mt= track), for mts motor proteins are kinesins and dyneins ( attach them to one mt and it works on other, or attach to one and drags it along) [lazer tauzer pulls cells to laser use wavelength thats not absorbed b cell can grab it and move cell where you want) |
kinesins |
move vesicles/organelles away from center of cell to peripheral (outside), [made of two identical heavy chains, 2 identical light chains] (+) directed motor proteins move to (+) end of mt [(-) directed move to center of cell], consume ATP and alternate attach/detach from mt surface, each step progresses 8 nm (one tubulin dimer)(1 ATP) , tail attach cargo (light chains), move from (-) to (+) end of mt; movement is progressive (stay on chain for long time without falling) heavy chain generates force and moves |
cytoplasmic dyneins |
move stuff from edge to center (-) directed, progressive like kinesins, force generated proteins in cilia / flagella, huge protein (1.5 E6 D) [2 identical chains and several light and intermediate chains], consume ATP and moves along mt, are on different protofilaments w/ kinesins but on same mt, heavy chain ( catalytic function and moves), light chain (has cargo)[ vesicles have both kinesis and dyneins attached but one works while other is inactive, (both 8nm a step) |
mt organizing centers(MTOC) |
cell controls arrangement of mts by regulating when/where they are assembled , mt assembly in vitro (in glass purified form) has 2 phases; nucleation( form ring of 13 tubulin dimers, slow growth, 1st step form mt wall), elongation (addition of tubulin to end , fast, usually (+) side, tubulin dimers added faster to (+) side than (-) side but disassembly faster on (-) end then (+) end; MTOC est. sites where nucleation can occur/be regulated, MTOCs have certain special y tubulin (found only here, anchor for (-) end, growth on (+) end |
in cells why is nucleation faster? |
nucleation is faster because the ring is pre-formed and have y tubulin attached to it; something assembles them put y tubulin (gamma tubulin) on it then alpha then beta tubulin above it
y-tubulin = gamma tubulin |
centrosomes |
major MTOC in cells usually near nucleus , animal cells centrosome has 2 cylindrical mt structures at right angles to one another called centrioles, centrioles have characteristic "9x3" arrangement of mt (9 triplets of 3 mt that fuse), centrosomes surrounded by fuzzy cloud PCM (pericentriolar material), mts originate in PCM and grow out from cloud, centrosomes is also og site of spindle apparatus assembly (cell has reserved dimers not in mt, if cell crawling centrosome in front side, |
in plant cells what acts as centrosome? |
in plants nuclear envelope acts as centrosome where mt radiate out and fungi too; centrosome grows mt and pushes it away from edges so its in the middle |
nucleation of mt in MTOC |
study of PCM reveals clues, PCM thought to serve as attachement/organization sites for ring shaped structures that serve to organize 13 initial y-tubulins, only alpha tubulins can attach to y-tubulins so polarity of mt is est.
|
mt dynamics |
y-tubulin orients dimers so alpha part of dimer attaches to y-tubulin, stability of mt arrays depends on structure ; cilia & flagella are extremely stable (MAPs), cytoplasmic mt are labile and changing, dramatic rearrangements are achieved by two mechanisms: rearrangement of existing mt, or disassembly/reassembly of arrays;rearrangements allow cells to respond to changing conditions, assembly associated w/ hydrolysis of ATP by beta tubulin |
cilia and flagella |
hairlike motile organelles project from surface work like oars, no homology to prokaryotic flagella, no fundamental differences between cilia and flagella, some differences n cytoskeleton anchoring structure of cilia and flagella but not organelles themselves, major motility structure ( power stroke and recovery stroke) |
ciliary structure |
entire organelle is covered with membrane , extension of plasma membrane, core (axoneme) contains distinct arrays of mt that run longitudinally thru entire length, "9+2" array of mt (9 outer pairs of mt & 2 mt in middle) 9x3 centriole (also called basal body) connected to 9+2 array of flagella (cilia) ( cilia has to have basal body) |
basal body |
root of cilium, if shave cilium regrow from basal body, 9x3 array of mt , two of the three continue into the axoneme, no central mt, homologous structure and content to centriole |
motility of cilia |
depend on dynein arms attached to outer parts of mt for it to move, sliding filament model (dynein on one pair interacts w/ mt of adjacent pair to generate force), bottom of axoneme is fixed (basal body) so mt pairs slide past one another, regulate size and frequency of movements by regulating dynein activity (central pair, Ca ion and cAMP) , central two mt apparently determine which pairs of outer pairs interacts to move ( only move with dyneins) |
intermediate filaments (IF) |
solid unbranched filaments, intermediate in diameter between mt and mf, radiate through cytoplasm of wide variety of animal cells;cross linked to mts by plectin, molecular diversity (many different types of IF proteins depends on cell type)
rope like cable like |
Microfilaments(MF) |
solid fibers made of one protein type (actin), actin subunits called G-actin (globular), when assembled into fibers called F-actin, units have polarity & all aligned in same order so fiber has polarity, assembly into MF requires ATP binding to actin, hydrolysis of ATP occurs sometime after each actin monomer binds to MF and can promote assembly, found in most eukaryotic cells, depending on type and context can be highly organized arrays, loose networks or tightly anchored bundles; have (+) &(-) directed motor proteins |
IF Types |
keratins -epithelia
vimentin-mesenchyme
desmin-muscle
lamins-all cells (nuclear envelope)
neurofilament proteins-neurons
etc |
common features of IFs |
central rod shaped,alpha helical domain w/globular domains on each end; two proteins interact by wrapping alpha helical domains around each other to form a ropelike dimer with N-termini and C-termini next to each other;two dimers lined head to tail to form functional tetramer (no polarity); line up tetramers into form filament: these self assemble into IF array |
IF characteristics |
highly resistant to pulling forces, provide tensile / stretching resistance to tissues, resistant to chemical disruption harder to extract and study than other cytoskeletal components; ex. keratin mats in epithelial cells make dense IF mat in mature cells, resistant to chemicals, water and bacterial invasion |
critical need for IF |
knockout mice deleted for various IF genes show specific health problems (tissue specific functions), ex mice deleted for type one keratin(epidermal cell expression)sensitive to mechanical pressure trauma (skin bruises or blisters easily),for us is epidermolysis bullosa simplex (mutation in human k14); overexpression of some IF genes lead to problems :too much neurofilament expression kills neurons, not all IF genes have such essential functions: mice missing vimentin gene show no obvious abnormalities |
ALS: lou Gehrigs disease |
amytrophic lateral sclerosis, neurodegenerative disease, killed popular baseball player lou gehrig, associated with excessive accumulation of NFs in affected neurons, mechanism obscure but too much NF kills neurons ( might plug up axon and prevent proper transport of materials up/down axon |
What does ATP and ADP do in MFs |
ATP assembles and ADP disassembles
similar to tubulin and GTP |
how does cell crawl around (actin part) |
cells crawl and cell has actin cortex under plasma membrane develops pressure on cytoplasm so cell sticking and pushes stuff out,cell will find something to adhere to and it will get reinforced (squeezing cell), on rear end proteins level cell off (pulling) feeling around important for cell motility
Cortex ( layer of cytoplasm under plasma membrane where actin are highly concentrated) |
actin types |
major contractile protein on muscle cells, cells possess a family of actin genes, each expressed for a specialized use, depending on cell type, actin genes are highly conserved across evolutionary time, yeast and mammalian actin can copolymerize and make fibers, |
MF assembly |
G actin binds ATP, when G actin bind to make F actin ATP hydrolyzed, (+) on filaments grows quickly and add G-actin (-)slowly site of depolymerization, changes in cytoplasmic conditions can shift equilibrium towards or away from polymerization, much actin polymerization initiated at plasma membrane, nucleating protein complexes: ARP complex (analogous to MTOC) |
Myosin MF motor protein |
sole function is motor of actin, all motors for actin are myosins, bind to MF: almost all (+) directed , all have large globular heads which bind MF and hydrolyze ATP, two types: unconventional (cytoplasmic myosins atlas 14 dif types, let cell crawl), and conventional ( muscular myosin only type in muscle cell type 2),cell may express several different types at once |
muscle contractions |
skeletal muscle cell(muscle fiber)structure unusual, a single cell is huge long and formed by fusion of many 100s procursor cells, highly ordered internal arrays of contractile units (myofibrils) , each myofibril is repeating linear array of contractile units (sarcomeres), each sarcomere has characteristic banded pattern |
sarcomere |
banding pattern arises from partial overlap of thick filaments (myosin II) and thin filaments (actin), H zone (thick filaments only,myosin), I bands(thin filaments only,actin), A band outside H zone overlap of both types, Z disk ( actin then myosin then actin), A-(mix of actin and myosin),M-(proteins keep all filaments lined up), myosins on left side all point to one direction and on other side point to other direction (double ended), (+) end is embedded in the Z line (Actin filament) (myosin is (+) directed) |
sliding filament model |
contraction of myofibril achieved by sliding interactions between myosin and actin in sarcomeres, each myosin head alternatively hydrolyzes ATP, binds actin, and changes shape to pull actin in one direction, myosin in contact with actin for a short period unlike kinesin with MT (myosin pulls actin on each side and contracts muscles)( needs fresh ATP every time) [ Ca ions release causes protein to protein interactions turn on myosins, pull Ca ions out and myosins relax] |
nonmuscle motility |
more difficult to study b/c components less ordered, more labile and typically restricted to limited regions of the cell; depends on cytoplasmic actin and sometimes myosin like motor proteins |
cytoplasmic actin array |
formation and stabilization of actin arrays depends on actin binding proteins ( ABPs about 100 dif types),ABPs determine organization and behavior of actin arrays, some assemble and stabilize some disassemble |
non muscle motility |
F-actins and myosin motors produce variety of cellular motility processes; cytokinesis, phagocytosis (endocytosis), cytoplasmic streaming, vesicle trafficking, integral membrane protein lateral movements, cell substrate interactions (movements), changes in cell shape |
specific examples of adherent cells |
cell migration on surface, cell not uniformly attached to surface; instead is attached at specific points (focal adhesions), each focal adhesion has MFs in tightly packed bundles extending into cytoplasm (stress fibers), stress fibers contain actin, myosin, and other muscle like contractile proteins, stress fibers not present in actively crawling cells, only in tightly adhering cells |
specific examples of crawling cells |
actively crawling mammalian cells spreads out like a ruffled veil (lamellipodium, LMLP), LMLP extended by actin arrays linked to/ organized by ABPs (ex. ARP 2/3 complex), LMLP adheres to substrate depends on membrane integrins contacting/ binding substances on surface (ex. fibronectin), bound integrins send signal into cytoplasm to mobilize more motility components to adhesion site, actin polymerization drives forward extension of LMLP; myosin driven contractions pull rest of cell forward to adhesion site |
developmental significance of motility |
examples; neural crest cells(arise on dorsal side of embryo, just over developing spinal cord,must crawl laterally or ventrally over most of embryo to destinations many mm away, developing neurons(axon end emerges from spinal cord and grows out to destination (muscle cells, gland cells) in body, growing end defined by MF arrays); both follow specific fibronectin and other ECM pathways to destinations |
cytosol |
contains many metabolic pathways, protein synthesis, cytoskeleton |
nucleus |
contains main genome, DNA and RNA synthesis |
ER, endoplasmic reticulum |
synthesis of most lipids, synthesis of most proteins for distribution to many organelles and to plasma membrane
|
golgi apparatus |
modification, sorting,and packaging of proteins and lipids for secretion or delivery to another organelle |
lysosomes |
intracellular degradation |
endosomes |
sorting of endocytosed material |
peroxisomes |
oxidation of toxic material |
general flow of material |
ER-golgi apparatus- lysosomes or vesicles to go out |
approaches to study endomembranes |
light microscopy, electron microscopy, cell fractionation, autoradiography (pulse-chase methods),genetic analysis, Green fluorescent protein markers |
cell fractionation |
isolate components in test tube and can control environment, break up cell in blender (homogenization), put in centrifuge and pull out heavy stuff and repeat and next heaviest stuff comes out, the solution becomes rich in substance but isn't pure, (example ER and membrane turn into circles called microsomes when they are broken up) [rough ER more dense than smooth ER so they end up at the bottom or below them] |
the two types of autoradiography |
pulse labeling( follow movement of molecules by labeling, example to find direction of flow, will add lil color to a part and follow the color), pulse chase ( is the following part, remove color or radiation) |
are proteins directly synthesized in organelle? is sort and transport of proteins into each organelle? |
most protein encoded in nuclear genome and most organelle don't have ribosomes, sorting is highly selective and energy dependent |
signal sequence |
it tells the protein where to go and if it is on the wrong protein that protein will still go to the destination |
steps in synthesis 1 |
mRNA of secretory protein bind to free ribosome in cytosol (translation begins), Near N-terminus of new protein, signal sequence of amino acids emerge (6-15 amino acids, non polar), signal sequence recognized by signal recognition particle (SRP) and binds to it [SRP particle: 6 polypeptides and 7S RNA in complex] |
steps in synthesis 2 |
binding of SRP to ribosomes/ mRNA stops translation, complex routed to surface of rough ER and binds specifically to SRP receptor (docking complex) on ER cystolic surface (SRP bind to SRP receptor, ribosome associates with membrane channel to lumen (translocon)) |
steps in synthesis 3 |
when ribosome bound to channel SRP interacts to widen channel, SRP is then released back into cytosol, translation resumes and polypeptide is inserted into lumen of ER as translation continues |
signal receptor transporter |
signal to send protein to place, receptor senses the signal, and transporter brings protein to the right place |
integral protein membrane synthesis |
initally the same process for ER localization and translation of polypeptides into lumen (signal sequence), but sequence of amino acids contain stop transfer sequence, it is about 15 hydrophobic uncharged amino acids, blocks further insertion of protein into lumen of ER, so ribosome lifts off surface of ER to complete translation, resulting protein spans membrane |
into nucleus (protein) |
has large pores, protein and RNA get through
small molecules diffuse freely but large molecules and complexes are actively transferred, bi-directional transfer |
nucelocytoplasmic transfer |
KAP(nuclear import receptor) recognizes the signal for nuclear protein and lets it thru the pore, KAP can also recognize nuclear export signals and take proteins out of nuclear thru pore |
Mitochondria import of protein |
most protein made in nuclear and have to be imported into the mitochondria, synthesized in cytosol, signal transfer sequence on N-terminus,receptor on mitochondria (on nothing else), sequence docks onto receptor and moves thru membrane and inner membrane thru a channel and it unfolds, then folds back in the matrix with help of chaperone then signal sequence is cleaved off |
proteins all start in ER |
synthesis happens on surface of ER (cytosol), proteins end up in lumen of ER (signal sequence) as being completed, signal sequence binds to receptor on ER surface and gets bound to ER as it is being made and then gets into the ER |
how proteins get into ER |
rough ER is a large endomembrane system that is in much of the cytosol(has ribosomes),mRNA binds to ribosome, protein grows 5' to 3' , signal sequence is on growing side and SRP (signal recognition particle) attaches to the signal (stalls translation), binds to SRP receptor on ER membrane, gets protein into the translocation channel and SRP displaced and is recycled (translation continues) [protein thread goes thru channel into ER and signal sequence is cleaved off], inside ER are chaperones that help it fold |
what happens when the signal sequence is cleaved off on the N-terminus |
there is a new N-terminus under the old one |
what are the locations of N-terminus and C-terminus depending on the stop transfer sequence |
can have N terminus inside and C outside, N terminus outside and C inside and both can be outside of ER channel
|
stop transfer sequence |
next one in line that stalls translations, it stops transfer across the membrane but not translation, rest of protein is made in cytosol |
cases where N terminus and C terminus could be according to the stop transfer sequence |
when N terminus normal and internal signal sequence (first part of protein made in cytosol , SRP binds to it and start transfer sequence tell it to go into channel and SRP falls off and translation continues, start transfer sequence threaded thru until stop transfer sequence and C terminus in cytosol; N terminus has start sequence then it will be inside; Start sequence in middle then C will be inside[start and stop sequences are hydrophobic] |
proteins in the ER when they don't fold properly |
when mistakes (don't fold properly), chaperones try to refold them if not then unfold protein response if don't get enough chaperones, receptors on ER surface signal molecules and tell nucleus to make more chaperones by activation chaperone gene expression then refold proteins with the extra chaperones |
glycosylation in ER |
most proteins made on rough ER are glycosylated (specific sugar groups are added), sugar sequence assembled by membrane bound enzyme (glycosyltransferases),in mammalian cells specific 14 sugar core oligosaccharide assembled, core then transferred as a block to specific R group sequence(asn-x-ser/thr) on new protein as it is translated, can be modified later in golgi processing, presence of sugar groups critical to proper folding/assembly of protein |
transport from ER |
new proteins not exported until folded (chaperons help), new material to be exported from ER are to collected at smooth surfaced regions facing the center of the cell, regions are called transitional elements (exit site for transport vesicles), small vesicles are pinched off ER and are transported towards Golgi apparatus[motor proteins on cytoskeleton (mt) take them places and fuse w/ target][ vesicles covered in protein coat called COP or catherin] |
how are vesicles formed and how do vesicles go into cytosol |
cargo receptors pick up cargo and adaptor proteins allow them to recognize the coat protein and make vesicle formation then make coat protein and a structure helps pinch off vesicle then coat taken off and naked vesicle go to its destination ; adaptor helps cargo receptor link with coat protein, it helps recognize each other |
Path from ER to Golgi |
[lots of coat proteins, different locations have different coat proteins] once vesicle is formed and budded off RAB and V-snare (on vesicle) are caught by t-snare and tethering protein, RAB is docked by tethering protein and V-snare is docked with T-snare then the vesicle is fused into the membrane and the cargo goes inside |
golgi apparatus and who was it discovered by and how |
by camillo golgi in 1898, unusual staining technique(some scientists didn't think golgis were real, confirmed existence by EM observations in 1950s
cis (accepts vesicles from ER and moves to Trans golgi) and trans golgi( vesicles exit from here) |
polarity of Golgi apparatus |
side facing ER is cis and side facing plasma membrane is trans, middle of stack is medial region;cis is usually convex and trans is concave, new material enters cis, go thru medial and leave thru trans; trans is sorting station b/c proteins are placed in different vesicle types for different destinations in cell |
golgi processing |
newly made proteins leave ER and enter Golgi on cis side and pass to trans; modification in golgi as protein passes thru[ parts of protein might be removed, amino acids might be modified, carbohydrate content might be modified by a series of rxns (removal and addition of specific sugar components) and sorted and targeted for delivery to ultimate location] |
movement thru golgi(3 different models) |
cisternal maturation model(new vesicles enter of cis and leave thru trans, each cisterna moves as a unit thru stack and matches as it moves, so new cis every time and old cis tunes to trans, different rxns happen at different layers so vesicles goes to all); Vesicular transport model( cisternae are stable compartments help in place, materials are shuttled thru the stack by small vesicles that transfer from unit to unit); Both both processes happen at same time |
types of secretions |
constitutive(for membrane proteins constantly secreted material, goes on all the time, unregulated) and regulated(material released only when signal received or under special conditions,example is synapse transmission)
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example synapse transmission for regulated secretion |
vesicles go thru mt track, cytosolic side stays cytosolic side, contact of vesicles dumped outside (excreted), hormones from blood stream and get signal to release then to outside of cell thru vesicles like synapse transmission, cargo is nt in vesicles acetylcholine for synapse transmission (nerve terminus), highly regulated, vesicles ready to fuse just waiting for signal (at plasma membrane) of nt |
endocytosis |
material into cell,destruction of old cytoplasmic membrane, material transported to lysosome (contains digestive hydrolytic enzymes for macromolecules, acid pH to activate these enzymes), hydrolysis of contents to monomers for absorption into cytosol (to be used elsewher) |
cholesterol and its way into cell to bring in cargo |
hydrophobic, packed into LDL (conglomerates associated with proteins) now soluble, cell gets thru by endocytosis, need receptor for it on membrane then docks and receptor activated then bring them in thru vesicles (same as exocytosis but inward), unchaste vesicles and repeats process, fuse with endosome and bring cargo in, cargo fall off in (pH 5) and receptor sent back to plasma membrane, endosome move cargo to lysosome (w/ its hydrolytic enzymes) and makes amino acids and choldesterol for cells to use |
cell signaling |
basis of cellular detection of environmental info and cell responses to that info, signal transduction process; from outside to inside |
signal transduction : components |
signal molecules (ligand), receptor molecules on cell surface, signal transduction molecules (cytoplasm and bind to stuff), effector molecules (cytoplasm and bring about change) |
signal transduction events |
recognition of signal (ligand binds to receptor molecule), transfer signal across membrane, signal transduction in cytoplasm, activates specific effector molecules in cytoplasm producing response and it could include (alteration of enzyme activity, rearrangement of cytoskeleton, change in gene expression, change in membrane permeability, activation of DNA synthesis, cell death (apoptosis)), end of response leads to destruction or inactivation of signal [cell sense environment then responds to it] |
what kind of responses are fast? what kind are slow? |
responses that alter protein synthesis in nucleus are slow and responses that alter protein function in cytosol are fast |
why does a cell get a signal as in what can the cell be told to do |
survive, divide, differentiate, and die (apoptosis), get the signal from other cells, |
can the same signals be used on different receptors and cause different responses? |
yes, signal molecules can be reused; example is acetylcholine can cause muscle contractions, salivary secretion and decrease heart rate |
what are the different kind of signaling? (4 different types) |
endocrine(hormones, long distance, affects other cells in other parts of body, have hormone receptors), Paracrine( short range, for neighbors, local general area have receptor), contact dependent( signal molecules starts on membrane so other has receptor and needs to be next to it), neuronal ( short range cells contact each other synaptic cleft, tight collection, pre and post synaptic cells) |
chemo attractant |
cell sense bacteria and follows it to ingulf it |
lymphocyte migration |
cells migrate to heal wound are attracted to chemoattractants because of receptors to respond |
cell surface receptors |
most signaling membrane receptors are on the surface |
intracellular receptors |
signal cross membrane and receptor in cytosol |
hydrophobic receptors (hormones) |
for steriods, cross membrane to get receptor in cytosol, when steroid hormone bind to receptor it moves to nucleus and regulated target genes and makes responses (regulated transcription affecteds gene expression)[ ex. cortisol (signal) binds to intracell. receptor and it gets conformational change then goes to nucleus and activates target gene then does transcription and makes mRNA] ( chaperone in cytosol helps in conformational change)
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ion channel linked receptors |
ligand binds to channel to open it |
G protein coupled receptors (GPCR) |
work thru g protein to activate target |
enzyme linked receptors (tyrosine kinase |
signal dimerize receptos and activate it |
example signal transduction system |
G-protein linked signaling system, Signal molecule (ligand) usually small hydrophilic molecules but odors hydrophobic, receptor molecules (large transmembrane protein, 7 membrane spanning segments), transduction systems (membrane associated heterotrimeric G proteins, membrane and cytosolic series of other proteins |
heterotrimeric G protein |
has alpha, beta, and gamma parts, uses GTP and GDP( inactive when bound), alpha and gamma have lipid tail attached to membrane, and beta is not attached to membrane but is attached to gamma, |
G protein process |
signal bind to receptor, activates G protein by removing GDP and replacing with GTP (alpha disassociates from beta and gamma and becomes active with GTP, and beta and gamma active without GTP) and can now transduce signal thru the cell |
target protein and G protein |
g protein active when GTP is on alpha and when beta and gamma are dissociated from alpha, alpha can bind to target protein and activate it; if GTP is hydrolyzed then alpha inactive and beta and gamma associate back and become inactive, |
example of g protein path |
acetlycholine is signal and binds to receptor which activates G protein then beta-gamma complex opens K+ channel, if GTP hydrolyzed then become inactive again and so does K+ channel |
sequence of events relay system |
G protein consists of 3 subunits (alpha, beta, and gamma), when activated by receptor/ligand G alpha binds GTP and detaches from G beta/gamma complex and moves laterally thru membrane, G alpha and GTP can bind to and activate effector molecule |
sequence of events : ending response |
signal ends when GTP on G alpha is hydrolyzed to GDP, G alpha slowly hydrolyzes its own GTP and thus inactivates itself, G alpha and GDP reassociates with G beta/gamma complex and resumes inactive form |
effector molecules |
target down stream of G protein to biological output, common effector (adenylyl cyclase), catalyzes rxn ATP (cAMP), cAMP ( is 2nd messenger activates many different cytoplasmic processes), diffuses thru cytosol binds other enzymes and activates them, Protein kinase A (PKA) is activated by cAMP, starts a complicated chain rxn |
g protein process with 2nd messengers |
signal (Ligand) binds to receptor (GCPR) then activates G protein and alpha binds to target protein then ATP makes cAMP and cAMP activates PKA and PKA activates phosphorylase kinase (proceeds down cascade) |
PKA |
protein kinase A, tetramer, 2 regulatory subunits (bind to cAMP) and 2 catalytic (phosphorylates), catalytic is separated from complex and phosphorylates after cAMP binds to regulatory subunits |
example 2 of G protein process with cAMP |
ligand binds to GCPR and activates G protein then G alpha binds to target protein then makes cAMP using ATP then cAMP activates PKA then proceed down cascade |
intrinsic GTPase |
random process where heterodimer is only on for a lil then GTP hydrolyzes to GDP, protein active for a lil then turns off, proteins can influence GTPase and slow it down or prolong it or speed it up |
GCPR |
C-terminus inside and N-terminus outside so it can bind to ligand (signal) |
2nd messengers |
small molecules, speed signaling, in signal amplification (acetylcholine), makes from one signal to lots of signals each step can amplify |
Other Second Messenger Systems |
Lipid Based: Phosphatidylinositol (PI) derived system
When activated, PI 4,5-bisphosphate is hydrolyzed into diacylglycerol (2 fatty acids and glycerol) and Inositol 1,4,5- trisphosphate (IP3)
Both products are second messengers and affect different parts of cell
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G protein process with IP and diaglycerol |
signal molecule binds to GCPR and activates G protein then G alpha activates Phospholipase C, which activates inosital phospholipid, which does two things, it activates diacylglcerol which attaches to PKC, inosital phospholipid also activates IP3 which opens the Ca 2+ channel that attaches to the PKC to activate it (PKC needs the diacyclglycerol and Ca ions to be active), PKC then creates the cell response |
other 2nd messenger system 2 |
calcium ions (in resting 10^-8 and in stimulated cells its 10^-4), membranes impermeable to Ca ions and PM and ER actively transport Ca out of cytosol; opening of Ca ion channels allows Ca into cytosol; cytosolic response mediated by calmodulin proteins ( modulates Ca ion activity) go from low to high concentration |
another major signaling system (RTKs) |
receptor tyrosine kinase (RTKs), example insulin receptors, many RTKs are also enzymes when activated they phosphorylate specific tyrosine R groups on themselves and on cytosolic target proteins, they use a different G protein pathway |
RTKs |
single monomers until signal dimerizes them, alpha helix is in the membrane, tyrosine kinase receptor proteins are inactive until phosphorylates when ATP is hydrolyzed then relay proteins can bind to tyrosine receptors when tyrosine gets phosphorylated |
Ras |
a monomeric G protein that is in use in RTKs pathway, uses GTP when active and (GDP inactive), tethered to membrane by lipid tail |
RTK path with Ras |
signal binds to receptor and dimerizes RTK then tyrosine receptor proteins get phosphorylated and adaptor protein gets attached to site and Ras activating protein attached to adaptor protein and activates Ras by switching GDP to GTP on Ras, then Ras carries on signal transmission |
MAP kinase and Ras |
Ras activates MAP kkk by conformational change then MAP KKK activates MAP KK by phosphorylation then MAP KK activates MAP K by phosphorylation then targets downstream proteins [ if Ras is mutated then GTP always on (either hydrolization is slow or it lost ability to hydrolyze GTP) so it is always sending signal ] |
RTK signaling without G proteins |
don't need G proteins, RTK dimerize and Jak attaches then phosphorylated tyrosine then get adaptor protein and stat proteins (transcriptional regulators cause genes to be expressed or repressed), stats usually inactive if not phosphorylated so in cytosol Jak activates stat then dimerizes with phosphorylation then sends signal |
why is it hard to figure out what happens downstream? |
messy, and complex |
signal pathways |
ancient paths, can swap parts with different species, few GCPR in plants don't have RTKs steroids or cAMP, animals an plants diverged before multicellularity because cell signaling is cell to cell interactions, communication
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