Lecture 19 & 20 - Actin Flashcards
Actin filaments (also known as ________) are ______ ______ of the protein actin
microfilament; helical polymers
Actin subunits
also called globular or G-actin
- polypeptides carrying a molecule of ATP or ADP
Is actin and ATPase?
yes; actin subunits have ATP, actin filaments hydrolyze this ATP into ADP
What provides polarity to the filament?
the way the actin subunits assemble
- at minus end (pointed end), the ATP-binding pocket is exposed
- at plus end (barbed end), the ATP-binding pocket is buried
Where does most growth happen on a filament?
from the (+) end
What is the rate-limiting step in the formation of actin filaments?
nucleation => very slow
- doesn’t happen in a cell
How many actin molecules are needed to make the growing complex more stable?
3 actin monomers; 2 actin molecules bind relatively weakly to each other
What gives the appearance of the filaments moving in the (+) end direction?
actin monomers are joining the (+) end while other are falling off the (-) end
What process pushes membranes forward?
actin treadmilling
What drug stabilizes F-actin?
phalloidin => cannot grow or shrink actin, we die
What are some examples of structures assembled from actin filaments?
- microvilli
- cell cortex
- adherens belt
- filopodia
- lamellipodium/ leading edge
- stress fibers
- contractile ring
What are the 2 structures that migrating cells make?
protrusive structures termed filopodia and lemellipodia
What do actin-binding proteins do?
they regulate polymerization and length of filaments
- monomer-sequestering proteins bind to actin monomer and prevent it binding to filament
- actin-polymerizing proteins
- filament-severing proteins cut filaments
- filament-capping proteins prevent dissociation
CapZ
a capping protein; prevent G-actin addition and loss
- binds to (+) end
- looks almost identical to G-actin but lacks actin binding site for other G-actin subunits
Cofilin
cuts actin into small pieces
What does actin filament crosslinking do?
makes complex structures
Alpha-actinin
a crosslinking protein that makes F-actin bundles
- a dimer
- crosslinkers must have 2+ actin binding sites
- rigid
Filamin
a flexible actin crosslinker protein
- crosslinkers must have 2+ actin binding sites
- a dimer
- flexibility allows different angles
Arp2/3 complex
promotes actin branching
- composed of 7 subunits structurally similar to G-actin
- initiates a new branched filament by binding to the side of the filament and recruiting actin monomers
What links the actin cytoskeleton the membrane?
ERM proteins
E= Erzin
R = Radixin
M = Moesin
- binding sites for actin and membrane proteins are hidden but revealed when Erzin is phosphorylated
How do cells assemble complex actin structures?
by dynamically changing the activity and localization of actin-binding proteins
How do actin binding proteins become activated?
with a signal
- Rho GTPases are the signal (Cdc42, Rho, Rac)
- on state is GTP bound
- off state is GDP bound
What do Rho GTPases do?
they coordinate the activation of all actin-binding proteins required to build a specific cytoskeletal arrangement
- each Rho GTPase protein signals the formation of a different actin structure
- they locally activate actin-binding proteins
Profilin and Formin
profilin: a small protein, key regulator of actin polymerization
formins: a group of proteins involved in the polymerization of actin and associate with the fast-growing end of actin filaments
- grabs monomers and puts them near (+) end
Where are filopodia and lamellipodia present?
at the leading edge of migrating cells
What pushes the cell forward?
actin polymerization and branching
- researchers removed the nucleus to see if transcription was involved, cell still moved
What are the motor proteins that walk on actin filaments?
myosins; transport a wide variety of cargo; moves towards (+) end
Myosin power stroke
- Without ATP myosin is attached to actin filament in rigor => myosin head is stuck to actin
- ATP binding to myosin head domain causes the release of actin filament
- ATP is hydrolyzed causing a large conformational shift in the ‘lever arm’
- Release of phosphate & myosin head binds to actin
- power stroke is the force generating step
- ADP is released, myosin head remains tightly bound (rigor state)
What part of myosin makes actin move?
the motor domain; without it, the filament does not move
What are the 2 types of myosin?
conventional and unconventional
- conventional form bipolar filaments
- unconventional do not form bipolar filaments
Myosin V
tethers and transports organelles
- globular tail binds cargo
- dimerization
- neck = Ca2+ regulatory region
- head = actin and ATP binding
Myosin 1
has only one head, binds membranes, and powers membrane deformation
- doesn’t move along actin filaments
- does power stroke, but doesn’t really release actin
