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Unmasking
fungi
Mention fungal cells, and Whitehead postdoc Robert
Wheeler thinks of M&Ms.
“Imagine a fungal cell as a sugar-coated pathogen,”
he says. “The organism is encased in a hard, white
‘candy’ shell, which is covered with a thin
layer of sweet ‘paint.’”
 |
Robert Wheeler |
The layers, which consist of complex sugars called betaglucan
and mannan, hint at an evolutionary arms race that plays
out over millions of years. First, immune systems detect
beta-glucan. The pathogens can’t get rid of the
protein because it serves a critical structural purpose,
so they hide it with a coat of mannan.
“This may very well be one more tactic in the
ongoing hide-and-seek game between our immune systems
and pathogenic fungi,” says Whitehead Member Gerald
Fink, who is Wheeler’s advisor.
| “I wouldn’t have tackled this project
without automated imaging,” says Whitehead
postdoc Robert Wheeler. “The Cellomics equipment
allowed me to conduct 96 experiments quickly at
the same time.” |
The pair worked with baker’s yeast (a fungus)
to identify the genes associated with the cloaking device.
They began with a library of yeast strains. Each one
contained a mutation on a specific gene. Wheeler placed
the strains in 96-well plates and stained them with
substances that bind to beta-glucan and mannan. He prepared
52 plates, as he wanted to screen all 4,080 yeast genes,
and loaded them into a Cellomics ArrayScan. He instructed
the microscope to take pictures, find cells and quantify
beta-glucan exposure for each strain.
A computer program processed the photos, locating cells
by searching for the mannan stain. It drew a circle
around each cell and tried to detect beta-glucan inside.
When the circles glowed bright, Wheeler knew he’d
poked holes in the cells’ outer coat of paint.
When the circles remained dark, he concluded the paint
was intact. Thus he pinpointed the genes responsible
for maintaining the yeast’s mask by disabling
them, one by one, and then monitoring the result.
“I wouldn’t have tackled this project without
automated imaging,” says Wheeler. “The Cellomics
equipment allowed me to conduct 96 experiments quickly
at the same time.”
His findings, which appeared in the April issue of
the journal PLoS Pathogens, have implications
for drug development. Pharmaceutical companies could
target the genes that control the mannan layer. Wheeler
hopes they will create more drugs like caspofungin,
which he showed can boost immune response by unmasking
fungi.
| Written by Alyssa Kneller with
contributions by David Cameron |
|
