Cytoskeleton: Intermediate filaments, cell motility Flashcards
Name key cytoskeletal functions
Structure and support
Intraclelualr transport
Contarctility and motility
Spatial organization
Are intermediate filaments simailr to actin and mts
Nawwwwww
Very diff
Not as highly conserved
Not same polarity, dynamics, proteins or formation either
What is purpose if
Mechanical stability
How if formed
Formed from a number of diff elongated subunits - independent of actin and mt
Flexible rope like fibres
Are if polar
Apolar
Not motors
Diameter if
100 angstroms
Where are if in cell
Extend across cell. Mechanical strength
Form nuclear lamina - mesh work on inner surface of nucleus
Where are ifs found in cell types
Nuclear lamins = evolutionary precursors - multiple duplications have given rise to cytoplasmic ifs
Found in animal cells
Need to enhance support for squishier organizsms
Describe nuclear lamina
Forms a mesh under nuclear envelope
Provides structural integrity to nucleus
Dissamebels during mitosis when nucleus disassembles
Assembly and dissamebly controlled by phosphorylation
Describe intermediate filament assembly - dimers
Alpha helical region in monomer
- fibrous proteins, differences in amino (n) and carboxy terminal domains = give ifs unique properties
Forms coiled coil dimer= coiled coil region relatively constant
Describe intermediate filament assembly - tetramer
Diners associate laterally via coiled coil domains
Antiparalele Arrnagment
Staggered tetramer of 2 coiled coil domains
Repeating subunit = equivalent to actin monomer or tubulin dimer
APOLAr = bc of antiparallel arrangement
Describe 2 tetramers - next step arrgangemt
Head to tail interactions generate elongated 2 stranded filaments
2 tetramers packed together
Describe intermediate filament - final assembly product
Then bundle the 2 tetramers with another = 2 stranded filaments associate laterally into 10nm fibres
All same diameter
= 16 units
Highly twisted rope like structure flexible and strong
Head to rail association so no polar ends
What do intermediate filaments impart
Mechanical strength
Ifs span cell and anchor at attachment sites
Desmosomes = packed with If - hair pin structure (Link if network between cells, Cadherin fam adhesion proteins), hemidesmosome - if stop working = detach from bm
Strengthens individual cells and whole cell layers
What happens when defect if
Loss of integrity of epidermis
All epithelial tissu rests on bm
Hemidesmosomes attaches num to cells of stratum germinativum
Weakened ifs allow epidermis to detach from basal lamina = bad
Describe neurofilaments
Impart strength to axons
Nf-l + nf-h heterodimers
Nlf-l + nf-m heterodimers
Nf-h c terminal tail creates cross bridges for tensile strength
Neurofilamets can be transported as cargo by mt motors to arrange them within neuron = grabs and brings them throughout axon tp where you need more support - cargo for motors - doesn’t have motors tho
If vs actin/mt - conserved
If proteins not as conserved as actin and tubulin - more cell type specific
If vs actin/mt - shape
If elongated fibrous
Actin and tubulin = globular, stronger, rope like filaments
If vs actin/mt - ntp
If polymeriztaion does not involve ntp binding or hydrolysis = generally les dynamic = more stable
If vs actin/mt - where
Actin and tubulin = expressed in all eukaryotic cells at some phase of development
Ifs = tissue specific
If vs actin/mt - Movemnt
Ifs not involves in movement - either of the whole cell or within cell = no motors
If vs actin/mt - polarity
If not polar
Describe cell motility
Plays a central role in many biological processes
Why cell motility important - 3 functions
Embryology = cellular migrations for gastrulation, nervous system development
Wound healing = essential migration of firbrobalsts and vascular endothelial cells
Metastasis = cancer, tumour cells migrate from initial tumour mass into circulation = migration
Describe how bundles of actin filaments and myosin 2 filaments formed in non muscle cells
Formed transiently to perform specific functions and then disassemble = dynamic system
Describe where myosin 2 is in non muscle cells
Relatively abundant in cortex of non muscle cells
Stress fibres are prominent in cultured fibroblasts = Represent a temporary contractile bundle of actin filaments and myosin 2
How is Assembly of myosin 2 filemants regualted in non muscle cells - step 1
Transient increases in calcium - binds calmodulin = activates myosin light chain kinase = mlck
How is Assembly of myosin 2 filemants regualted in non muscle cells - step 2
Mlck phsophorylates 2 light chains associated with Myosin head = has 2 effects
How is Assembly of myosin 2 filemants regualted in non muscle cells - step 2 = results = effects
Release of myosin 2 from a binding site on the head, allows myosin 2 to form bipolar filaments = heads open up
Change in conformation of myosin head exposing acting binding site =allows interactions between actin filament and myosin 2 filaments = leads to contractile response
Describe additional regulation of mlck
Involved rho gtpase
Active rho (rhogtp) = activates rho kinase = promotes activation of mlck
Rho kinase all phosphorylates and inactivates mlck phosphatases (remove phsophate) = prevents dephosphorylation and inactivation of mlck
= exposes head and allows tails to do coiled coil conformation
Describe how interactions between stable actin filaments and myosin 2 thick filaments is regulated in muscle cells
In skeletal muscle = actin myosin interacted regulated by actin based regulatory system
Calcium binds troponin = moves tropomyosin, myosin ready but tropomyosin in way
Tropomyosin binds in groove of actin filament helix and blocks binding site of myosin head
= myosin head can interact when tropomyosin gone
Name the 4 steps of cell Motilty
Protrusion
Adhesion
Traction
Deadhesion/tail retraction
Describe protrusion - Gen
Actin polymerization
Involves forward of the membrane at the front of the cell - at Leading edge of cell
Describe adhesion - Gen
Integrins
Required for protrusion to be translated to movement along a surface
Describe traction - Gen
Myosin 2
Process leading to forward movement of nucleus and cell body
Also sometimes myosin 1
Describe deadhesion/tail retraction - Gen
Myosin 2
Mehcanistically distinct processes involved in last step of cell locomotion
Some cells do leave parts behind tho
What is basic engine for cell motility
Actin cytoskeleton
rapidly moving cells can move without microtubuels - like keratinocytes, neutrophils but crawling motility always needs actin
(Myosin related process = retraction, external signals = bind cortex and mt gets signal = knows where to go, gradient based, retrograde g actin comes back to feed cell, coil in breaks up actin behind polymerizing actin )
Analogy of cell motility
Microtubules = steering wheel
Actin polymerization = engine = generate force
What must cells acquire in order to migrate
Spatial asymmetry to migrate = need clear distinction between front and back of cell
Marked by increase in concentration of actin filaments at a particular region of cell
Describe protrusion - step 1
2 types of protrusion structures can be discerned at front of migrating cell = filapodia and lamellapodia
Describe filapodia - step 1
Cdc42
Thin, cylindrical projections, contain actin filaments organized into tight bundles which exclude myosin’s
Describe lamellapodia - step 1
Rac
Broad, flat, sheet lie in which actin filaments are more loosely organized in orthogonal criss crossed pattern due to actin filament cross linking protein and arp2/3 complex
Senses environment and leads to leading edge
Describe polarity fo actin filaments in protrusion structures - step 1
Have barbed + end of actin pointing at leading edge
Accepted model = generated by actin polymeriztaion alone
Af cross linkers import bc increase rigidity of actin bundles or networks to facilitate pushing forward on membrane
Each monomer on to filament pushes membrane further
*70degrees -arp2/3
Describe lamellipodia formation- step 1
Cell signal - like chemoattractant binding to receptor - at membrane = activates wasp
Wasp = activates arp2/3= initiates f actin polymerization
Arp2/3 binds to walls of f actin = initiates branching f actin - at 70 degrees
F actin grows at barbed end and depolymerizes at pointed end = causes pm to push out - leading edge
Describe ex of actin polymerization model for membrane protrusion
Pathogenic bacteria = listeria monocyogenes, gram + , can cause life threatening illness = encephalitis
Listeria can invade mammalian cells and once internalized = bacterium escapes membrane bound endosomes and replicates and resides happily within cytoplasm of host cell
Force pushing bacteria
Within cell = bacterua hijacks host machinery required for actin polymerization and generates actin comet tail = propels bacterium within host cell
Bacterial motility directly coupled to actin polymeriztaion - independent of myosin function -actin formed by arp2/3 bacterial version
Describe step 2 = adhesion = specifics
Required to stabilize and consolidate forwards protrusive movements
Experiments have shown that front of cell = preferential locus where adhesions from
Newly synthesized and cycled integrins transported to leading edge of new cell bc leading edge constantly losing adhesion receptors - as cell moves forwards
Inetgrins - binds surface then actin and myosin pull in that direction
Integrins recyeld = endocytosed often bc only limited number of them
Describe step 3 - tractions
Also need contractile force to move cell forward
Contractile force must overcome resistance in order to effective translocate the cell body - resistance due to adhesive interactions of Integrins and breaking these interactions -requires application of physical stress via contraction of myosin 2 and af
MYOSIN 2 BASED
Pulling of cell body forwards by contractile forces acting between back edges of leading lamella and nucleus could also involve myosin 1
Describe step 4 - deadhesion/tail retraction
Rapid cell migration requires efficient mechanism to release adhesion at rear of cell
Mechanism for deadhesion seen in fibroblasts = involves membrane ripping = Integrins left behind on substratum as cell moves forward, Integrins remain on cell surface and are endocytosed into vesicles and reused at leading edge - ripping which leads to tail retraction = myosin 2 mediated process
