New findings suggest a surprising level of accuracy in regulation of protein controlling body plan development
By Melissa Lee Phillips | July 12, 2007
The mechanism that sets up the basic body plan of the developing fly is surprisingly precise, according to two papers in this week's Cell. The finding contradicts earlier studies that suggested that expression levels of the key protein were only loosely controlled.
"The way they've done this analysis is really spectacular," said Nipam Patel of the University of California, Berkeley, who was not involved in the work. "They're getting down to the physics level of what's going on."
Researchers twenty years ago identified the transcription factor Bicoid, which controls the temporal and spatial expression of a cascade of genes that determine the anterior/posterior axis of the fly body plan. The transcription factor forms a concentration gradient across the embryo - high Bicoid levels at the fly's anterior end activate anterior-specific genes while low Bicoid levels at the posterior end activate posterior-specific genes. Scientists have considered Drosophila a model system for studying body patterning in animals.
"Attempts like this to delve into the details will certainly provide paradigms for other systems," said Steve Hanes of the Wadsworth Center at the New York State Department of Health in Albany, who was not involved in the study.
Previouswork suggested that Bicoid's expression levels were only loosely controlled but that its downstream patterning genes had very precise expression levels. It wasn't clear how noisy Bicoid expression could be converted into precise target expression, said lead author Thomas Gregor and his colleagues at Princeton University in New Jersey. Since previous studies were done only in fixed tissue, Gregor and his colleagues decided to examine Bicoid expression in live flies. They fused green fluorescent protein to the bicoid gene in Drosophilamelanogaster and used two-photon laser-scanning microscopy to quantify the Bicoid gradient as it formed during the first three hours of embryonic development.
In the first paper, the authors show that the Bicoid gradient forms within the embryo within about one hour of fertilization. Over the course of a single nuclear division, the researchers found that Bicoid concentration varies four-fold in each nucleus. At the beginning of every cycle, however, the nuclear concentration of Bicoid returns to the same level.
The four-fold variation in nuclear Bicoid concentration throughout a single nuclear cycle is "really quite stunning," Hanes told The Scientist. It's not yet clear how that variation will influence gene expression, but "new models for how Bicoid stimulates expression of its target genes will have to take this into account," he said.
They also examined how Bicoid diffusions throughout the cell, to try to determine how it establishes its gradient. Two independent methods showed that Bicoid diffuses very slowly through the cell -- too slowly, in fact, to explain how it can set up its gradient in only an hour. The authors speculate that the diffusion rate may be different in different parts of the embryo or at different timepoints of embryogenesis.
"The way that the system works is probably more complicated than we think," Patel said. "Simple diffusion is not sufficient to explain it."
In the second paper, Gregor and his co-workers found that the embryo can distinguish between very small differences in Bicoid expression. Just a 10% difference in Bicoid concentration in a nucleus can determine whether or not downstream genes are activated. Also, Bicoid expression was almost exactly the same in corresponding regions of different flies.
The precision with which Bicoid is expressed within and between flies "makes things quite a lot simpler to understand," said Claude Desplan of New York University in New York City. "Before, it was some kind of mysterious phenomenon where a non-robust gradient... leads to a very robust expression pattern of target genes."
However, it remains possible that Bicoid's expression still contains some variability that is filtered before target gene activation, Hanes said. "I am not fully convinced of their arguments that the noise level for Bicoid is minimal."
Jun Ma of Cincinnati Children's Hospital Medical Center in Ohio, also not a co-author, agreed that Bicoid expression may still contain some noise. "I would not rule out that there is a multi-layer correction mechanism to basically ensure that the system works well," he said. If Bicoid expression is as controlled as the authors say, that leads to a different question, Ma said: "What makes the Bicoid gradient so precise?"
It's also a mystery how the embryo can distinguish between such small differences in Bicoid concentration when deciding which genes to activate in different parts of the embryo, Gregor said. "It's very difficult to imagine how this would work," he told The Scientist.
Overall, the findings "raise as many questions as they answer," Hanes said.
Melissa Lee Phillips
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