Final Flashcards
The three cytoskeleton components ordered on size
1)Micro tubules
2)Intermediate filaments
3)Microfillaments
Main roles of the cytroskeleton
1)Scaffolding
2)Movement of material
3)generation of force in contraction
Types of microtubules
1)Cytoplasmic:Found in the cytosol of axons and miototic spindles, shape cells and move vesicles
2)Axonemal:Organized into special structures like flagella, cilia, and basal bodies
Tubulin
Compose micro tubules, they form straight hollow cylinders which form the 13 piece structure called a protofillament
Characteristics of microfillament alpha and beta subunits
Contain a N-terminal GTP binding domain, a central domain, and a C terminal domain capable of interacting with MAPS. Alpha forms the minus terminal and B forms the plus terminal
Tubule arrangements and their functions
1)Single:Standard usage
2)Doublet:Cilia and Flagella
3)Triplet: Basal bodies and centrioles
Three steps in tubule formation
1)Lag phase:Nucleation occurs via the aggregation of dimers into oligmers
2)Elongation:Sub units are slowly added onto the nucleus formed
3)Plateau:Tubulin concentration limits the addition and removal of subunits
Critical concentration
At this concentration the rate of assembly and disassembly is balanced on both ends
What happens when
Cc<C+ and C-
Cc>C+ and C-
C->Cc>C+
Cc<C+ and C- , Loss on both ends
Cc>C+ and C-, Gain on both ends
C->Cc>C+, Tredmilling
Conformational change occurring as new sub units are added to the microtubule
On the B end, whenever a new sub unit is added, GTP is hydrolyzed into GDP
Stability of the MT when tubulin is high
1)GTP bound B tubulin stabilizes the tip for growth
2)GDP bound tubulin destabilizes the MT
This happens because GTP cannot be turned into GDP fast enough
MTOC
Attaches to the minus end of the MT acting as an anchor and helping to nucleate the microtubule for rapid growth using its y tubulin
Centriole
Centriole walls are formed by 9 sets of triplet microtubules and they’re involved in basal body formation for cilia/flagella. Cells without centrioles have poorly organized mitotic
spindles but can still divide
Regulation of MT assembly
1)ATP to drive their transport of vesicles of organelles
2)Stabalizing/bundle proteins
3)Plus-End Tubulin Interacting Proteins
4)Microtubule-Destabilizing/Severing
Proteins
MT bundling proteins and their roles
Allow for the interaction with other cellular structures and help space them
1)Tau:Form bundles in axons
2)MAP2:Form loose bundles in dendrites
TIPS
Stabilize microtubules by capturing the growing end and protecting it from catastrophe
Severing proteins
Destabilize the micro tubule and prevent growth
Op18-Binds to tubulin dimers to prevent polymerization
Catatrophins-Promotes peeling of subunits apart on the ends
Katanins- Severe the ends of micro tubules
Microfillaments
Smallest fillament unit involved in cell migration and muscle contraction, alpha actin is used in muscles and b and y actin are involved in all other roles
G and F actin
G actin:smallest actin subunit polymerizing with a lag, elongation, and plateau
F actin:Polymer of G actin wound into a helix using ATP hydrolyses
two actin structures involved in full cell movement
Lamellipoda and filopodia
Proteins responsible for regulating polymerization of microfillaments
1)Profillin
2)Cofillin
3)Thymosin
Profillin
Binds to ADP g-actin and catalyzes the exchange of ADP for ATP, promoting polymerization
Cofillin
Binds to ADP actin, severing it and promoting depolymerization
Thymosin
Binds to ATP actin to prevent them from joining the microfillament chain
Proteins involved in capping of microfillaments
1)CapZ:Binds to the + end to prevent loss/gain
2)Tropomodulin: binds to the - end to prevent loss/gain
Severing proteins of microfillaments
1)Cofillin
2)Gelosin:breaks actin MF and caps newly exposed + end to prevent future polymerization
Formins
Controls the assembly of microfillament polymerization via nucleation to speed up growth and prevent capping proteins from attaching
Cross link protiens of MF
1)Filamin-Act to join two microfillaments together where they intersect
2)Fimbrin-Bundle microfillaments into tightly linked and ordered structures
ARP2/Complex
Nucleates the branching on MF leading to the formation of Lamellipodia
Intermediate filaments
Most stable, least soluble, non polarizable cytoskeleton component that act as scaffolding and a bridge between cell components
Cytoskelton components role in movement
MT-Bending resitence
MF-Generate tension
IF-Withstand pressure
Steps in assembling an intermediate fillament
1)2 of the the polypeptides coil together to form the 45nm rope
2)Two dimers assemble into an anti parallel tetramer leading the loss of polarity
3)Eight tetramers form one unit
4)Units associate with each together to elongated into intermediate filaments
5)Pieces can then be modified from the middle with no energy cost
Kinesin
Move down MT anterograde
Dynein
Move down MT retrograde
Cell motility
Movement of an organism through the environment, past of through the cell, or within the cell
Two methods of motility in cells
1)Kinesins(+) and Dynin(-) movement along MT
2)Actin microfillament and myosin motor proteins
Anatomy of kinesins
1)Globular head region that attaches to the MT
2)Neck holding the stalk
3)Coiled stalk to allow flexibility of movement
4)Light chain which attaches to cargo
Movement of Kinesin
1)Leading head chain binds ATP
2)ATP causes the swinging forward of the other head
3)Trailing chain finds new MT binding site
4)new leading heavy chain releases ADP, and old leading chain turns ATP into ADP and Pi and Pi is released
Dynein classes
1)Cytoplasmic dynenin which position the centrosome and golgi complex and mitotic spindles
2)Axonemal Dyneins:move flagella and cillia
Cillia vs flagella movement
Flagella move with a propagated bending motion due to inhibition and activation of dynenins
Cillia move with oar like perpendicular force due to the nexin linkage
Axoneme
Nine micro tubule doubles surrounding a central singlet pair linked using nexin
Intraflagelleur transport
Movement of structural components between the doublets and the membrane, kinesin 2 moves towards the tip and cytoplasmic dyneins move material back
Similarities between kinesins vs Myosin motors
1)Use ATP hydrolosis
2)One form is associated and the other dissociated
3)Similar shape
4)Move to plus end
Diffrences between kinesin and myosin motors
1)Larger steps along the track
2)Uses MF instead of MT
Bipolar filaments
2 heavy chains and 4 light chains forming Filaments with opposite polarity at the two ends
Thick filament
Staggered arrays of myosin 2
Thin Filament
Actin microfillaments with other bound proteins
Tropomyosin
actin filaments that play a critical role in regulating the function of actin filaments in both muscle and nonmuscle cells by blocking binding sites of troponin
Troponin
binding site of myosin head in the power stroke
Tropomodulin
Caps the (-) end of the microfilaments for stability
CapZ
Caps the (+) end of the microfilament forming the Z line
Nebulin
Stabilizes and binds the thin filaments to the Z-line
Alpha Actin
Cross links the Z-line to the microfillament keeping the thing filament in a parallel array
Myomesin
Bundles the myosin molecules together
Titin
Attaches the thick filament to the z line and keeps thick filaments in the correct positions relative to the thin filaments in contractions
Sliding filament model
Muscle contraction is due to the thin filaments sliding past the thick filaments with no change in length of the fibre
Steps in the movement of actin
1)ATP binds to myosin dissociating the myosin/actin cross bridge
2)ATP is hydrolyzed to ADP+Pi putting myosin into cocked position
3)Energized myosin binds to the actin filament
4)Pi is released pulling the actin forward
5)ADP is released but myosin remains bound to actin
6)ATP binds to myosin, dissociating the myosin/actin bridge
Ca2+ regulation of contraction
Electrical signals open RYANDINE receptors that release Ca2+ from the Sarcoplasmic reticulum, contraction can only occur when Ca2+ is present as it binds to tropomyosin and allows myosin to bind to troponin
Cell crawling
1)Signal is received to crawl
2)Extension of lamellapoda push the membrane forward using ARP2/3, WASP,WAVE
3)Protrusion attaches to the substrate generating tension
4)Tail is released snapping the cell forward
Cell cycle phases and role
1)G1+G2, Doubling mass and proteins
2)Sphase, Duplication of DNA
3)Mphase, actual duplication occurs splitting the cell
4)G0, holding phase
Prophase
Nuclear envelope dissolves, chromosomes condense, cytoskeleton dissembles
Prometa phase
Spindles interact with the chromosomes, phosphorylation of the nuclear pores causes them to dissociate, and inner nuclear membrane dissociates with lamins and chromsomes move to equator, spindle forms
Lamin A
Depolymerization of intermediate filaments and nuclear lamina
Kinetochore
- Kinetochores serve as a site of:
1. Microtubule/chromosome attachment
2. Motor proteins, necessary for chromosome mobility
3. Signaling for important mitotic checkpoints
Metaphase
Chromosomes are aligned and attached to microtubules
Astral Microtubules
- project toward the cell cortex
- help position the spindle apparatus in the cell
Kinetochore microtubules
-connect to chromosomes to the spindle poles
-align the chromosomes on the metaphase plate and pull
them to the poles during anaphase
Polar microtubules
-extend past the chromosomes and interact with polar microtubules from the other spindle pole
-maintain integrity of the mitotic spindle and push the
spindle poles apart during anaphase
Anaphase
Centromere split and chromosomes separate in two steps due to enzyme Seperase
In anaphase A, the chromosomes are slowly pulled centromere first toward spindle poles
as kinetochore microtubules get shorter
* In anaphase B the spindle poles themselves move away from each other as polar
microtubules lengthen
Proteins involved in separation of chromosomes during anaphase
1)Specialized kinesins:binds to the end and promotes depolymerization of MTs at both the Kinetochore and centrosome
2)Bipolar kinesins-5:motor binds to overlapping polar microtubules causing them to move apart
3)Cytoplasmic dyneins:Moves towards the (-) ends of the microtubles towards the cell membrane
Telophase
- At the beginning of telophase the daughter chromosomes arrive at the poles of the spindle
- Mitotic spindle disassembles
- Chromosomes uncoil into interphase chromatin
- Nucleoli reappear and nuclear envelopes reform
- During this period, cytokinesis may also start
Cytokenesis
Causes parental cell to split into two at the spindle midzone using actin filaments
Cleavage of cell in cytokenisis
Cleavage depends on a belt-like bundle
of actin microfilaments that form just below
the plasma membrane in early anaphase, contraction is regulated via bipolar myosin 2 and actin filments
3 checkpoints in the cell cycle
1)Start, Cell cycle will only trigger inf conditions are favourable
2)G1/M Checkpoint, determine if all DNA is duplicated before division
3)Metaphase to anaphase check- are chromosomes attached to the spindles?
Steps involved in cell signalling
1)Synthesis of ligand
2)Signal is released via exocytosis
3)Transit of signal molecules to target cells
4)Signal molecule binds to a protein receptor on the target cell
5)Ligand- receptor interaction results in confirmation change in receptor
Endocrine signalling
Horomone is produced from the tissue and reaches target cell via circulation of blood stream
Paracine signalling
Diffusion of signalling ligand diffuse and act local over a short range
Autocrine signalling
Act on the cell that produces it
The different signalling molecule types
1)Eicosoids/ fatty acids
2)Amino acids
3)Steroids
4)Polypetides and proteins
3 classes of cell receptors
1)G protein
2)Enzyme linked
3)Ion channel linked receptors
KD
Disassociation constant or the state in which half the receptors of a cell are occupied
agonists
drugs that activate the receptors that they are bound to
Antagonists
Drugs that bind to receptors without activating it preventing the regular messenger from binding and inhibiting signal production
Process of ligand to intracellular signal
1)First messenger arrives and binds to transmembrane receptor
2)receptor changes shape and the cytoplasmic domain can create a second messenger or directly signal
3)Deactivation or activation of some molecule leads to cellular process
Path ways of intracellular g protein signal
1)Convergence:Signals from multiple signaling pathways converge
2)Divergence:Signals from one ligand activate multiple pathways
3)Cross talk:Signals from different receptors affect components of multiple pathways
Desensitization
When receptors are not used for some time cells no longer respond to the signals, this is done via
1)Less receptors on the surface
2)Adapt to signal by lowering affinity for ligand
GPCR
G-protein coupled receptors, they regulate a number of cellular processes via ligand binding
Anatomy of GPCR
3 Extracellular loops- involved in ligand binding
3 intracellular loops-signalling proteins binding site, in the inactive state the protein binding site is buried in the active state the ligand bind disrupts the trans membrane interactions via alpha helix rotation allowing g protein binding
Heteromeric G-protein
Held into the plasma membrane by the covalent attatchments of lipid chian, Galpha is a fatty acid and binds to GTP/GDP and Gy/Gb are isoprene proteins that can serve in signalling
Off and On for Galpha
GTP-ON
GDP-OFF
GPCR signaling pathway
1)Ligan binds to GPCR causing confirmation changes unmasking the g protein binding site
2)Galpha releases GDP and binds to GTP
3)Galpha dissociates from Gb and Gy via lowered affinity with Galpha now acting as an effector protein. Gb/Gy can also associate with K+ channels and open gated channels
4)Effector can lead to production of a second messenger eliciting a downstream response
5)G protiens are on until Galpha turns GTP->GDP +Pi
6)Galpha now reduce affinity for effector and now re-associating with Gb/Gy
How to prevent over stimulation of a GPCR
1)GPCR cytoplasmic domain can be phosphorlated
2)The phosphorlated GPCRs are now more attracted to Arrestin which compete for g protein binding site
Arrestin
Target receptors to clatherin coated pits, removing the receptor from the cell surface to which it can then
1)Move into endosome and continue signalling
2)Lysosome degradation
3)Returned to surface for alter resensitization
cAMP
Formed by Adenylcylcase, inactive until it is bound by Galpha, when it binds to other proteins it can modify their activity, its main target is Protein Kinase A
Target of cAMP
Protein kinase A
How can cAMP have differing effects in differing cells?
1)Diffrent cells have diffrent PKA
2)AKAP help organize where PKA is moved within the cell
AKAP
Coordinates protein-protein interactions by moving PKA to different cellular locations
ID3 and DAG
Formed when PIP2 is cleaved by phospholipase C
How are ID3 and DAG activated
1)Gaq is activated on the G protein
2)Gaq activates PLC which cleaves PIP2 into IP3 and DAG
3)IP3 diffuses through the cytosol and binds to ligand-gated calcium channels
How to increase Ca2+ in the cell
1)Voltage gated channels opening
2)IP3
3)Ryodine receptors
RTK
Receptor tyrosine kinases, regulate differentiation, cell division. cell survival, and are activated by growth factors that lead to the dimerization of the RTK
Mechanisims of RTK dimerization
1)Ligand binding links the two individual receptors together
2)Ligand binding leads to conformational changes and dimerization of receptors occurs
Regions of RTK
1)Extracellular-Binds ligand
2)Single transmembrane domain-Participates in dimerization
3)Dimerization/aggregation- Activates RTK
4)Large cystolic domain-Location of the kinase and phosphorylation target and SH2 binding site
adapters RTK
Act as linkers of two or more proteins to form a signalling complex with SH2 or PTB domains to link RTK
Activation of RTK pathways
1)Recruitment of the enzymes to the membrane, placing them in close proximity to the target
2)Binding of the pTYR onto RTK resulting in conformational changes in catalytic domain
3)Phosphorylation of the enzyme to increase or decrease the catalytic activity.
Termination of RTK signaling
Same as GPCR but with a motif signal instead of arrestin
1)Endosome signalling
2)Lysosome destruction
3)Return to surface
Regulators of GTPase
1)GAP:activate GTPase by hydrolyzing GTP
2)GDI:Inhibit GDP dissociation
3)GEF:replace GDP for GTP
RAS-MAP signal pathway
- The pathway is activated when a growth
factor (EGF) binds the extracellular domain
of an RTK - Ligand binding leads to:
a) dimerization of two receptors
b) trans-autophosphorylation of the cytosolic
portion of the RTK protein
c) The SH2-domain containing protein Grb2
has high affinity for the pTyr on the RTK
d) Grb2 is constitutively associated with a
RasGEF called Sos - Recruitment of Sos to the membrane
brings the RasGEF into close proximity
with Ras, allowing GDP to be replaced with
GTP (Ras Activation!) - Ras-GTP (not Ras-GDP) has a high
affinity for the protein kinase Raf
(MAPKKK)
a) RAF is recruited to the membrane and is
activated by Ras-GTP (via phosphorylation)
b) Raf phosphorylates Serine and
Threonine residues in a protein kinase
called MEK (MAPKK) - MEK in turn phosphorylates proteins
called ERK or mitogen-activated
protein kinase (MAPK) - Activated ERK/MAPK can phosphorylate
over 160 different proteins, including the
transcription factors Ets and Jun - These transcription factors activate
genes involved in cell proliferation
RASGAP
Facilitates GTP to GDP hydolyses to deactivate RAS which helps prevent cancer
MAP kinase and diffrent cell response
1)Diffrent MAPKKK->MAPKK->MAPK combos
2)Scaffolding protein can tether diffren MAP kinase pathways together and restrict them to specific areas
Nuclear receptors
Receptors that are found in the cytoplasm and are acitvated by horomones.
1)Ligand binding domain
2)DNA binding domain which bind to DNA elements with a two finger zinc motif that enhances or down regulates gene expression
Transcription control of nuclear receptors
1)Hormone enters target cell and binds to cytoplasm receptor
2)Nuclear localization signal is exposed
3)Complex enters nucleus and activates/inhibits genes
