(Press-News.org) Contact information: Diana Lutz
dlutz@wustl.edu
314-935-5272
Washington University in St. Louis
Samurai sword protein makes strategic cuts in cell skeletons
Carefully placed and timed cuts give cell skeletons the best pattern for a particular function
Just as our bodies have skeletons, so do our cells. They're equally indispensible in both cases. Without our bony skeletons we'd go limp and fall down. And without our cytoskeletons, our cells, which come in roughly 200 different shapes and sizes, would all become tiny spheres and stop working.
Using cells from the stem of a seedling as a model system, Ram Dixit's lab at Washington University in St. Louis seeks to understand the molecular mechanisms that organize and pattern the hundreds or thousands of microtubular "bones" of the plant cytoskeleton. In their model system, the microtubules form parallel bands like barrel hoops around the cell's girth.
Dixit's lab shows in the Oct. 24 online issue of Current Biology that misaligned microtubules that grow over existing microtubules are cut at the crossovers by the enzyme katanin, named for the katana, or samurai sword. Once a microtubule is cut, the part downstream of the cut falls apart, disintegrating into individual tubulin units.
Because katanin shows up at crossovers just before a microtubule is cut and because there is no cutting in a mutant plant line lacking katanin, the WUSTL scientists are sure that katanin and katanin alone is responsible for this activity. In the mutant the microtubules form disorganized cobwebs.
The scientists also showed, by chilling cells to destroy their cytoskeletons, that katanin organizes the cytoskeleton in the first place as well as maintains its organization once it has formed.
Stars, hoops, fans and cobwebs
The cytoskeleton — whether it's in an animal or a plant cell — is the framework that organizes the interior of the cell, Dixit says. It has two basic functions. It helps shape and support the cell, and it serves as a highway along which molecules and organelles move from one part of the cell to another.
To perform its functions, the cytoskeleton has to be organized in a specific pattern, however.
Animal cells have something called a centrosome, also called a microtubule organizing center, or MTOC. The surface of the MTOC is studded with microtubule nucleation complexes from which microtubules arise and to which they remain tethered. Given these constraints, its not hard to see why microtubules form starburst arrays around centrosomes.
But there are many cell types that have ordered microtubule arrays that aren't created by centrosomes. Some nerve cells, for example, have very long projections (axons) that are chock full of microtubules.
The microtubules are aligned with the axis of the axon and they're not connected to the cell's centrosome in the cell body, which can be some distance away. "How do you order microtubules and generate a specific pattern when you don't have centralized control?" asks Dixt, an assistant professor of biology in Arts & Sciences.
VIDEO:
A short movie clip shows a single severing event. The microtubule array is fluorescent green and the microtubules' growing tips are marked red. The white arrow follows the growing tip...
Click here for more information.
The same question arises with muscle cells, which also have linear microtubule arrays and with the cells that line the gut, which have flat microtubule arrays in their flanks. In both cases, the arrays are distant from the cell's microtubule organizing center.
The cytoskeletons in the cells of land plants are also patterned according to function without the help of a centrosome.
The guard cells that open and close the stomata on the under surface of plant leaves, for instance, have fan-shaped arrays that follow their curves. Pavement cells on the leaf surface that are shaped like interlocking puzzle pieces have net-like arrays. And rapidly elongating cells in plant stems have transverse arrays that then reorient toward the longitudinal direction as growth slows.
Lit up like a Christmas tree
It's hard to study these arrays in animal cells, says Dixit. "The microtubules go deep into the cytosol and are hard to manipulate or image at high resolution. Besides animal cells, all have a centrosome, so that's always a confounding factor.
"So we use plant cells as a model system instead," Dixit says. They don't have centrosomes; instead the microtubules nucleate at dispersed sites in the cell cortex, a layer of cytoplasm on the inner side of the plasma membrane. And when the cells are not dividing, the microtubules are plastered to the inside of the plasma membrane, where they're easily accessible and easy to image.
The cells Dixit's lab use are from a lineage of Arabidopsis plants created by Erica Fishel, PhD, then a WUSTL graduate student in biology, that express two fluorescent tags, or marker proteins. One colors the entire microtubule fluorescent green and the other marks its growing tip cherry red.
Quan Zhang, PhD, who was a postdoctoral research associate in the Dixit lab, crossed this marker line with an Arabidopsis mutant that does not produce the katanin enzyme.
While a WUSTL undergraduate, Tyler Bertroche (AB '11), generated a plant lineage where katanin is tagged with green fluorescent protein. Thanks to their efforts, the Dixit lab now has several different lineages of color-coded wild-type and katanin-mutant Arabidopsis.
Time is of the essence
Dixit Lab
A short movie clip shows a single severing event. The microtubule array is fluorescent green and the microtubules' growing tips are marked red. The white arrow follows the growing tip of one microtubule as it crosses over existing microtubules. A pink arrow appears when a crossover is cut. The microtubule disintegrates behind the cut and disappears.
"So how do cells pattern a microtubule array and how do different cells do it differently?" Dixit asks.
Microtubules originate in the cell cortex but once a cytoskeleton is established, they also sprout from existing microtubules in what is called branching nucleation.
Dixit and other scientists had shown through simulation that these branches would disorder the array unless some kind of pruning or culling mechanism was also in play.
Meanwhile, scientists at the University of Manchester in England had observed that misaligned microtubules were severed at junctions where a growing microtubule crosses an existing microtubule.
Zhang in the Dixit lab began with these observations and devised experiments to clarify the molecular mechanisms underlying them. He showed that they are insensitive to the geometry of the crossover or the stoutness of the underlying microtubule bundle. Instead, all that mattered was time.
How rapidly crossovers were cut determined what sort of array formed. In the transverse arrays, microtubules were cut, on average, within 41 seconds of a crossover event. In net-like arrays, on the other hand, cutting was three times slower and not as tightly controlled.
Plants have several severing proteins that might cut microtubules, but of these, katanin was the likeliest suspect. And in the katanin mutant, none of the crossovers were severed, demonstrating that this enzyme is solely responsible for cytoskeleton patterning.
To make sure, the scientists tagged katanin green and the microtubules red. When the cell was color coded in this way, they were able to see that katanin almost always localized to crossover sites before they were severed.
Coming in from the cold
These experiments showed that katanin was responsible for maintaining array patterning, but does it also create the pattern in the first place? To find out, Zhang used a trick he found in the literature, Dixit says.
Microtubules are cold sensitive and fall apart when they are chilled. "So Quan would put cells in the freezer for four or five minutes, take a slide out, run to the microscope, and watch to see what happened as the cells warmed up," Dixit says.
In the wild-type cells, the microtubules quickly re-appeared and became well ordered. In the katanin mutant, however, the microtubules re- appeared but never became organized.
The punchline, Dixit says, is that microtubules get cut at crossovers; the timing of the severing differs from one pattern to the next; katanin does the cutting; and if katanin is not present, the cells can neither generate or maintain ordered arrays.
INFORMATION:
Samurai sword protein makes strategic cuts in cell skeletons
Carefully placed and timed cuts give cell skeletons the best pattern for a particular function
2013-10-24
ELSE PRESS RELEASES FROM THIS DATE:
Curing HIV/AIDS gets tougher: Study shows far more 'hidden' and potentially active virus than once thought
2013-10-24
Curing HIV/AIDS gets tougher: Study shows far more 'hidden' and potentially active virus than once thought
Discovery of a larger than expected latent reservoir of HIV confounds 'shock and kill' cure strategy
Just when some scientists were becoming more hopeful about ...
Researchers design global HIV vaccine that shows promise in monkeys
2013-10-24
Researchers design global HIV vaccine that shows promise in monkeys
Preclinical study provides strong rationale for clinical trials
BOSTON -- The considerable diversity of HIV worldwide represents a critical challenge for designing an effective ...
Genetic mutation provides clues to battling childhood obesity
2013-10-24
Genetic mutation provides clues to battling childhood obesity
As the number of children battling obesity continues to grow, researchers are racing to identify causes and possible interventions. Now, a new paper publishing October 24 in the journal Cell identifies a possible ...
Bee sting allergy could be a defense response gone haywire, Stanford scientists say
2013-10-24
Bee sting allergy could be a defense response gone haywire, Stanford scientists say
STANFORD, Calif. — For most people, a bee sting causes temporary pain and discomfort, but for those with a bee venom allergy, the consequences can be devastating: ...
Researchers identify gene variant that raises risk for colorectal cancer from eating processed meat
2013-10-24
Researchers identify gene variant that raises risk for colorectal cancer from eating processed meat
Discovery sets first step towards identification of genetic variants linked to carcinogenic risk from diet and nutrition
A common genetic variant that affects ...
Reservoir of hidden HIV larger than previously thought
2013-10-24
Reservoir of hidden HIV larger than previously thought
New findings put spotlight on need for new drugs to target HIV proviruses
In the fight to cure human immunodeficiency virus (HIV), researchers have been dealt a blow. A new study by Howard Hughes Medical Institute ...
Researchers apply brainpower to understanding neural stem cell differentiation
2013-10-24
Researchers apply brainpower to understanding neural stem cell differentiation
How do humans and other mammals get so brainy? USC researcher Wange Lu, PhD, and his colleagues shed new light on this question in a paper that will be published in Cell ...
Novel genetic mutations cause low metabolic rate and obesity
2013-10-24
Novel genetic mutations cause low metabolic rate and obesity
Researchers believe the gene could be a useful therapeutic target for treating obesity and type 2 diabetes
Researchers from the University of Cambridge have discovered a novel genetic cause of severe ...
For fish and rice to thrive in Yolo Bypass, 'just add water'
2013-10-24
For fish and rice to thrive in Yolo Bypass, 'just add water'
From a fish-eye view, rice fields in California's Yolo Bypass provide
an all-you-can-eat bug buffet for juvenile salmon seeking nourishment
on their journey to the sea. That's according ...
Physicists decode decision circuit of cancer metastasis
2013-10-24
Physicists decode decision circuit of cancer metastasis
Rice U. research reveals 3-way genetic switch for cancer metastasis
Cancer researchers from Rice University have deciphered the operating principles of a genetic switch that cancer cells use to decide when to metastasize and ...
LAST 30 PRESS RELEASES:
Less intensive works best for agricultural soil
Arctic rivers project receives “national champion” designation from frontiers foundation
Computational biology paves the way for new ALS tests
Study offers new hope for babies born with opioid withdrawal syndrome
UT, Volkswagen Group of America celebrate research partnership
New Medicare program could dramatically improve affordability for cancer drugs – if patients enroll
Are ‘zombie’ skin cells harmful or helpful? The answer may be in their shapes
University of Cincinnati Cancer Center presents research at AACR 2025
Head and neck, breast, lung and survivorship studies headline Dana-Farber research at AACR Annual Meeting 2025
AACR: Researchers share promising results from MD Anderson clinical trials
New research explains why our waistlines expand in middle age
Advancements in muon detection: Taishan Antineutrino Observatory's innovative top veto tracker
Chips off the old block
Microvascular decompression combined with nerve combing for atypical trigeminal neuralgia
Cutting the complexity from digital carpentry
Lung immune cell type “quietly” controls inflammation in COVID-19
Fiscal impact of expanded Medicare coverage for GLP-1 receptor agonists to treat obesity
State and sociodemographic trends in US cigarette smoking with future projections
Young adults drive historic decline in smoking
NFCR congratulates Dr. Robert C. Bast, Jr. on receiving the AACR-Daniel D. Von Hoff Award for Outstanding Contributions to Education and Training in Cancer Research
Chimpanzee stem cells offer new insights into early embryonic development
This injected protein-like polymer helps tissues heal after a heart attack
FlexTech inaugural issue launches, pioneering interdisciplinary innovation in flexible technology
In Down syndrome mice, 40Hz light and sound improve cognition, neurogenesis, connectivity
Methyl eugenol: potential to inhibit oxidative stress, address related diseases, and its toxicological effects
A vascularized multilayer chip reveals shear stress-induced angiogenesis in diverse fluid conditions
AI helps unravel a cause of Alzheimer's disease and identify a therapeutic candidate
Coalition of Autism Scientists critiques US Department of Health and Human Services Autism Research Initiative
Structure dictates effectiveness, safety in nanomedicine
Mission accomplished for the “T2T” Hong Kong Bauhinia Genome Project
[Press-News.org] Samurai sword protein makes strategic cuts in cell skeletonsCarefully placed and timed cuts give cell skeletons the best pattern for a particular function