Cytoskeleton II Flashcards by Joel Glotfelty

Cellular movement/migration, intracellular transport, and ciliar and flagellar beating are all dependent on the

Cytoskeleton

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Involved in spindle assembly, chromosome alignment, and segregation in mitosis

Kinesins and dynein

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Cleave the cell in the contractile ring during mitosis

Actin and myosin II

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The driver of morphogenesis

Cytoskeleton

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The folding of the epithelial sheet is driven by myosin II-dependent contraction of the

Adhesion belt

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The transport of vesicles between cellular compartment and to the cell periphery requires

Cytoskeleton

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Molecular motors carry cargo on

Microtubules and actin filaments

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Intracellular transport involves which cytoskeletal elements?

Actin filaments and microtubules

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In the intracellular transport on actin filaments and microtubule, the traffic goes

Both ways

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What are the three classes of cytoskeletal motors?

) Myosins
) Dyneins
) Kinesins

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Move along actin filaments

Myosins

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Move along microtubules

Dyneins and kinesins

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ATPases that have multiple isoforms encoded by multiple genes

Myosins, dyneins, and kinesins

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Have related structures and mechanisms, but different functions

Kinesins and Myosins

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A particular isoform (type) of myosin, dynein, and kinesin on an actin filament or microtubule moves in

One direction

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Vesicles/organelles can move on microtubules and actin filaments and have more than one kind of

Motor

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The downstream targets of cellular signaling cascades

Molecular motors

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Myosins, Dyneins, and Kinesins are all classified as

Mechanochemical enzymes

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Work by generating tension or movement of an object along a filament or microtubule

-ex: muscle contraction, chromosome segregation, vesicle movement

Myosins, Dyneins, and Kinesins

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Myosin has a large gene family. The enzyme is made up of

Two heavy chains and two light chains

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Most myosins move towards the

(+) end of actin (“plus-end directed)

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The globular head, or motor domain, of myosin contains the

ATPase

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Variable domain of myosin. Contains coiled coil for dimerization, and/or binds to membrane or target vesicle

Tail domain

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The myosin superfamily shows homologous motor domains, but variable

Tail domains

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The classification of a myosin protein is based on its

Motor domain

Each member of the myosin super family has specific cellular

Localizations and functions

Mutations in myosin family cause inherited human diseases such as

Myopathies, deafness, blindness, etc

Myosin has many roles including

1. ) cytokinesis 2. ) intracellular transport 3. ) endocytosis

First isolated as the motor responsible for anterograde axonal transport - Smaller than myosin - mutations cause human inherited diseases

Kinesin

Like the myosins, most kinesis are what type of movers?

(+)-end directed -some are (-)-end

The head domain of kinesins,which contains the ATPase, is structurally related to

Myosin and G proteins

Located in the heavy chain and contains the ATPase and microtubule binding site -conserved across kinesin family

Motor domain ("head")

Which type of kinesin motor is (+)-end directed?

N-terminal motor

Which type of kinesin motor is (-)-end directed?

C-terminal motor

Located next to the motor region and determines the polarity of the kinesin

Kinesin Neck region

Allows for dimerization when it is a coiled coil

Kinesin Stalk region

Binds to the target, variable across kinesin family, and binds light chains

Kinesin Tail region

A minus-end directed motor -an ATPase

Dynein

Dynein is a very large protein. It weighs approximately

2 Million daltons

Dynein is an AAA protein, meaning it is

ATPase associated w/ diverse cellular activity

Dynein typically contains 6 domains. The energy of hydrolysis leads to a conformational change which leads to

Work

Dynein is found in -necessary for beating

Ciliary/flagellar arms

The cytoplasmic forms of dynenin function in

retrograde axonal transport

Functions during mitosis and in the transport of mRNA, vesicles, organelles, and viruses

Dynein

The tail of dynein binds cargo or to another

Microtubule

What is an example of a dynein associated disease?

Primary Ciliary Dyskinesia (PCD) -i.e. kartakener syndrome

Characterized by respiratory tract infections and male infertility

Primary Ciliary Dyskinesia (PCD)

In PCD, there is a mutation in the outer arm ciliary dynein heavy chain. This results in

Immobile cilia

PCD is associated with

Situs inversus

All function by converting the energy of hydrolysis of ATP into mechanical work

Cytoskeletal motors

Causes a conformational change in the cytoskeletal motor

Binding and/or hydrolysis of ATP

The rate-limiting step for cytoskeletal motors is

Product release following hydrolysis

Accelerates the rate limiting step

Binding actin or microtubule

You can think of the actin filament/microtubule as a

Nucleotide exchange factor (NEF)

Differ in processivity and the fraction of the kinetic cycle they spend attached to microtubules or actin

Kinesin and Myosin

Filament forming myosin

Myosin II

Results in attachment of myosin II to actin and causes rigor mortis in mucle

Depletion of ATP

The binding of ATP thus causes the

Release of Myosin II from actin

In the myosin II reaction, the rate limiting step is -accelerated by actin binding

Pi release

Operates by an analogous mechanism to myosin

Kinesin

The binding of ATP weakens the affinity of kinesin for

Microtubules

After ATP hydrolysis, Kinesin binds microtubules with a

Higher affinity

Binding to microtubules following ATP hydrolysis accelerates product release and conversion from

Weak to strong binding (power stroke)

For myosin and kinesin, when ATP is bound we see which stroke?

Recovery stroke

For myosin and Kinesin, when ATP is hydrolyzed, we see tighter binding to actin/microtubules and which stroke?

Power stroke

In dynein, the microtubule binding site is remote from the

ATPase domain

When nexin is present between two microtubules, they can not slide. The force generated by dynein movement then causes the cilium to

Bend

A given motor moves in only

One direction

Motor-cargo relationships are highly

Specific

Single headed myosins: the tail can bind to the plasma membrane, and in microvilli, the actin filaments oriented so the (+)-end is pointed to the tip of the microvillus

Myosin I

Form bipolar filaments as in: muscle, contractile ring of cytokinesis, stress fibers, and adhesion belts

Myosin II

Allows for contraction, as in sliding filaments

Myosin II

Carries cargos such as ER, vesicles, proteins, and nucleic acids, on its tail and walks towards the (+) end of the actin filament

Myosin V

Designed for long range transport: processive, longer steps than other myosins

Myosin V

Bipolar myosin filaments are involved in

Contractile functions

For vesicles to reach the end of the growth cone or cell periphery they must travel from the cell body to the periphery, i.e. they must

Transfer from microtubules to actin filaments

Which three motors are involved in vesicle transport?

Dynein, kinesin, and myosin V

Defines the cell's polarity and organization

Transport of vesicles

Are transported from the ER to the cell periphery and back -Can travel in both directions on a microtubule (motor-dependent)

Vesicles/organelles -also proteins and nucleic acids

Can occur on + end of a microtubule via a + tip protein, as a microtubule grows

Vesicle/organelle transport

Can have more than one molecular motor on its surface -can travel on both microtubules and actin filaments

Vesicle/organelle

The model system for transport is the transport of

Pigment granules in skin

Pigment is in vesicles called

Melanosomes

Develop from neural crest cells, as do many neurons

Melanocytes

They are transported on microtubules and actin filaments to the periphery of the cell where they are taken up by keratinocytes. -Myosin V is required for transport to the periphery

Melanosomes

Normal melanosome distribution in the skin requires

Myosin V

Binding of cargo causes a transition into it's active conformation -without cargo it exists in an inactive folded conformation

Myosin V

Transports melanosomes and synaptic vesicles to the cell's periphery

Myosin V

Target myosins to specific intracellular membranes

Rab GTPases

The motor domain of myosin Va moves along actin carrying the melanosomes to the periphery of the

Melanocyte

The myosin V tail binds to its cargo by way of

Melanophilin (Mlph) and Rab27aGTP

Mice null for Rab27a, melanophilin (Mlph) or myosin Va results in clustering of melanosomes around the

Nucleus

In humans, mutations in myosin Va, melanophilin, or Rab27a cause - results in melanosomes remaining in the basal level of the epithelium - causes silvery hair and light skin in children of dark-skinned parents

Griscelli type 1 and Elejalde synromes

Recent work on melanocytes shows that melanophilin not only links myosin Va to melanosomes, but that it also regulates

Dynein

Involved in recruitment of vesicles carrying receptors into dendritic spines is associated with long term potentiation, synaptic changes that are believed to be associated with learning and memory.

Myosin Vb