How Does Intracellular Molecular-

Motor-Driven Transport Work?

Joseph Snider2, Frank Lin2, Neda Zahedi3,

Vladimir Rodionov3 , Clare Yu2 and Steve
Gross1,2
1
Department of Developmental and Cell Biology
University of California, Irvine;
2
Department of Physics
University of California, Irvine;
and
3
King’s College, London
A Cell Is Like a City
• Workers • Proteins
• Power Plant • Mitochondria
• Roads • Actin fibers, microtubules
• Trucks • Kinesin, dynein, myosin
• Factories • Ribosomes
• Library • Genome
• Recycling center • Lysosome
• Police • Chaperones
• Post office • Golgi apparatus
• Communications • Signaling networks
 Intracellular Traffic
 How is intracellular
transport regulated?

© Scientific American
Microtubules (MT)
are like freeways
and actin filaments
are like local
surface streets.
 Filaments
• 10 nm diameter
Actin filament • 2.77 nm rise
• 26 subunits/74 nm repeat

- end + end

+ end

 25 nm diameter
Microtubule  13 protofilaments

- end
Motor proteins

Myosin

Kinesin
 Kinesin Myosin-V Dynein
Cargo Cargo

KLC
Pi Dynactin
binding

KR2
KR1
MR2
Head
Ca2+ (ATPase) Stalk
Pi

2 1 c
KAPP
3 6
4 5
KR3
MR1

KHC Lever (?)

Head (ATPase) MT binding

Motor
proteins
move cargo
along
filaments Molecular Biology of the Cell, 3rd Ed, 1994

Biochemistry, 4ht Ed, 1995
 Herpes Virus Transport in
Neurons Along Microtubules

• Virus Movie: VirusMov.mov

Microtubules (MT)
are like freeways and
actin filaments are
like local surface
streets.

How does the cell regulate

the transport of vesicles?
Model System:
Melanophores (Pigment granule cells)
• Cells
change color by
Dispersing or Aggregating
pigment granules
• granules move bi-directionally along
microtubules
(Kinesin-II and Dynein)
•Granules also move along actin filaments
(Myosin-V)
Dispersing Pigment Granules

• Dispersion: Granules move out from nucleus:

Dispersion1.mov
Aggregating Pigment Granules

• Aggregation: Granules move in toward nucleus:

Aggregation1.mov
Dispersion: Aggregation:
MTActin

Spread ActinMT
cargos Bring cargos
throughout back to
the cell nucleus
 Experiment
• Pigment cells (melanophores) only have
actin filaments (no microtubules)
• Pigment granules (melanosomes) are
tracked
• Record position r vs. time t
• Plot average <r2> vs. time t
 Experiment: Cargos traveling solely on actin filaments
go farther during dispersion than during aggregation.

Do cargos go faster during dispersion? No, the velocity of

the cargos is the same for aggregation and dispersion.
 Why do cargos go
farther during
dispersion than
during aggregation?
• During dispersion cargos go “straight” to the end of the
actin filament and do not turn at intersections with other
actin filaments. This is good for spreading out pigment
granules uniformly.
• During aggregation cargos have a 50-50 chance of
switching to another actin filament at each intersection.
So they don’t go as far. Frequent switching is a good
way to find a nearby microtubule.
 Two Types of Theoretical
Modeling Confirm This Scenario
• Langevin solution interpolates between
short time ballistic (straight-line) motion
and long time diffusive motion.
• Computer simulations of cargos moving
along actin filaments also confirms this
picture.
 Solution to Langevin Equation
r (t )  D  t    1  e  
2
  t /

• Langevin solution interpolates between

short time straight line motion and long
time diffusive motion.
• Fitting displacement data yields D and τ
which can be used to obtain the mean free
path ℓ (distance traveled before turning).
• The mean free path is given by
l  D / 2
 Fitting Parameters

• Fit to Langevin Solution

r (t )  D  t    1  e  
2
 t /

• Fit to power law


r (t )  At
2
Langevin Fits Yield Mean Free Path

• Dispersion: <ℓ> = 539 ± 9 nm

l  D / 2
• Aggregation: <ℓ> = 237 ± 12 nm
 Compare Langevin ℓ with Electron
Micrographs of Actin Filaments

Dispersion: Langevin ℓ L/2 where L≈1300 nm is a typical

filament length implying cargos go to end of filament
Aggregation: Langevin ℓ ≈ 1.5 d where d≈160 nm is the
typical distance between filament intersections consistent
with cargos switching with 50% probability at intersections
Electron Micrographs of Actin Filaments

The actin filaments appear denser during aggregation

which would encourage frequent switching from one
filament to another. (Not enough EMs to confirm this.)
Filament Density and Touching Number
• To quantify the
density of filaments,
randomly place
circles on the EM.
• Circle diameter =
568 nm = cargo
diameter
• Touching number =
number of filaments
in contact with circle
Touching Number nt

Aggregation Dispersion
< nt > = 7.8 < nt > = 4.2
 Simulations of Cargos Moving on
Actin Filaments
• Distribution of filament lengths taken from
EMs (electron micrographs)
• Vary density of filaments (touching
number)
• Vary switching probability at filament
intersections
 Simulations of Cargos Moving
Along Actin Filament Networks

Trajectories more localized Trajectories more spread out

 Switching Probability

Aggregation: nt = 7.8, <ℓ> = 237 nm, switch = 50%

Dispersion: nt =4.2, <ℓ> =539 nm, switch = 0% - 6 %
 Result of Simulations of Cargos
Moving on Actin Filaments
Using the density of filaments taken from EMs and
the Langevin mean free path, we find:
• For aggregration, switching probability is 50% at
filament intersections
• For dispersion, cargos go to end of filament and
then attach to a new filament (switching
probability is very small ~ 0%)
• Result: Average mean free paths agree with EMs
and Langevin, confirming scenario
 Simulations of Cargos Moving
Along Actin Filament Networks

Trajectories more localized Trajectories more spread out

Cargo displacement after 30 sec from
simulations

•Cargos are more

localized during
aggregation
•Cargos are more
evenly spread out
during dispersion
 SUMMARY:
We have explained how and why
cargos go farther during dispersion
than during aggregation
• During dispersion cargos go “straight” to the end of the
filament and do not turn at intersections with other
filaments. This is good for spreading out pigment
granules uniformly.
• During aggregation cargos have a 50-50 chance of
switching to another filament at each intersection. So
they don’t go as far. Frequent switching is a good way
to find a microtubule that leads to the nucleus.
 Possible Way that the Switching
Probability Is Regulated
• During aggregation, there are about 60 motors per
cargo, but only one active motor pulls a cargo along a
filament. Another motor can attach to a nearby filament
and cause a switch to the new filament. Switch
probability is 50%.
• During dispersion, there are about 90 motors per cargo,
but only 2 active motors pull a cargo. Another motor
may try to attach to another filament but it is not strong
enough to cause a switch to a new filament. Switch
probability is 0.
 Actin Collaborators
• Steven Gross (Cell and Dev. Biology, and Physics and
Astron., U.C. Irvine)
• Joseph Snider (Physics and Astron., U.C. Irvine)
(Langevin and simulations)
• Francis Lin (Physics and Astron., UC Irvine) (data
analysis)
• Neda Zahedi (King’s College London and U. Conn.
Health Sci. Ctr.) (experiments)
• Vladimir Rodionov (U. Conn. Health Sci. Ctr.)
(experiments)
• Snider et al., PNAS 101, 13204 (2004).
• Website: http://bioweb.bio.uci.edu/sgross/
THE END
Fraction of cargos that have touched a
MT 15 times by time t (from
simulations)
 Collective Motion vs. Single Motor
Properties
•Cargos go further in dispersion due to collective
motion rather than the properties of individual
molecular motors.
•Analogy: To understand traffic flow patterns in
southern California, you don’t need to know how a
car works. Learning about tires and internal
combustion won’t tell you why there’s a traffic
jam.
Actin Filament Length Distribution
 Quantification of motion
• Particle tracking: 8nm resolution, 30 Hz
• Analysis: Displacement vs. time R (t)
random motion