HOUSTON -- (Nov. 5, 2010) -- A new study by Rice University
bioengineers finds that the workhorse proteins that move cargo
inside living cells behave like prima donnas. The protein, called
kinesin, is a two-legged molecular machine. Rice's scientists
invented tools that could measure the pulling power of kinesin both
singly and in pairs, and they report this week in Biophysical
Journal that kinesins don't work well together -- in part
because they are so effective on their own.
"Researchers have been investigating the mechanical properties
of individual motor proteins for some time now, but this is the
first time anyone's been able to tie a defined number of molecular
motors to a cargo and watch them work together," said lead
researcher Michael Diehl, assistant professor in bioengineering at
Rice. "We know that more than one of these motors is attached to
most cargoes, so understanding how they work together -- or fail to
-- is a key to better understanding the intracellular transport
system."
Cargoes inside cells are hitched to teams of motor proteins and
hauled from place to place like horse-drawn wagons. Like
stagecoaches or wagons, many cargoes are pulled by several horses.
But unlike a wagon, cellular cargoes often also have multiple teams
pulling in opposite directions.
"Motor proteins move directionally," Diehl said. "They either
move toward the cell's nucleus or they move away from the nucleus
toward the periphery. Grouping different types of motors together
allows cells to regulate cargo movement. But when there are
multiple motors pulling antagonistically in opposite directions,
what determines which group wins? What influences the balance? How
do they cooperate or compete to get the right packages to the right
place? Those are the kinds questions we're trying to answer."
Diehl said intracellular transport has become an increasingly
hot topic over the past decade, in part because researchers have
found that breakdowns in the transport system are linked to
neurodegenerative diseases like amyotrophic lateral sclerosis (ALS)
and Huntington's disease.
One question Diehl and lead co-authors Kenneth Jamison and
Jonathan Driver helped answer in the new study is how much pulling
power a pair of kinesins could apply to a cargo compared with the
amount applied by a single kinesin.
The apparatus they created to study the problem was years in the
making. Driver and Jamison, both graduate students, used strands of
DNA to make a scaffold, a sort of molecular yoke that they could
use to hitch a pair of kinesins to an experimental cargo. The cargo
in their tests was a microscopic plastic bead. Using laser beams in
an instrument called an optical trap, they attached teams of
bead-pulling proteins to microtubule roadways.
As the motors walked down the road, they pulled the bead away
from the center of the optical trap. At the same time, the lasers
in the trap exerted counterpressure in an effort to move the bead
back to the center of the trap. Eventually, the light won out,
forcing the motors to let go and the cargo to snap back to the
middle of the beam. By measuring the precise movements of the bead
during this reaction, Diehl's team was able to determine exactly
how much force a team of motors exerted on a bead.
"Compared with other motors, kinesin is actually a pretty strong
performer," he said. "Single kinesin motor molecules can produce
relatively large forces, and they rarely step in the wrong
direction when walking along microtubules. This is remarkable
behavior, considering kinesin is a molecular-scale machine that
experiences significant thermal and chemical fluctuations."
Given how well they perform alone, it would be easy to assume
that a group of kinesins would pull harder than a single kinesin.
But Diehl points out that a team of kinesins can only harness the
combined potential of both motors under certain circumstances.
"Our analyses show that the two kinesins must stay in close
proximity to one another to cooperate effectively," he said.
"Otherwise, one of the motors will tend to assume all of the
applied force imposed on the cargo. Kinesin is relatively fast and
efficient on its own, but they have trouble keeping up with one
another when they are connected together."
Diehl said the group suspects that other classes of motor
molecules, which are somewhat weaker than kinesin, may function
better in groups. The team is carrying out follow-up experiments to
see if that's the case, and they are examining how such
distinctions may play a role in regulating cargo movement in
cells.
Diehl's research group, which is located in Rice's new
BioScience Research Collaborative, has spent years refining the
tools used in the new study, and the work is paying off in numerous
ways. Within the past four months, the group won an R01 grant from
the National Institutes of Health worth more than $1.4 million, and
Diehl also published a theoretical study of motor proteins with
Rice chemist Anatoly Kolomeisky.
SOURCE