32 Students + 15,000 Insects Equals One Biological Clue

One of biology's most challenging puzzles — how organisms change in response to the pressures of natural selection — might just have become a little easier to tease apart, thanks to research that took 32 undergraduate students and more than 15,000 caterpillars (of children's book fame) to complete. 
Using tobacco hornworm moths (Manduca sexta) as a model system, a team of researchers at the University of Arizona developed a method that provides a conceptual framework for simplifying very complex biological processes into a small number of parameters that can be used to make predictions about an organism's response to selection. 
Published in the journal The American Naturalist, the method dramatically reduces an organism's physiological, developmental, endocrine, genetic and molecular complexity into manageable chunks amenable to experimental study. The approach can be applied not only to insects, but across many taxa, including amphibians, mammals, insects, green algae and plants.
By greatly reducing the complexity of the underlying regulatory networks that govern how genetic information is translated into phenotypic traits, the new approach promises to facilitate a process called genotype-phenotype mapping, a long-standing goal of evolutionary biology. 
Mapping the relationship between genes and traits is a necessary step in figuring out how evolution acts on an organism through natural selection, but elucidating those relationships has proved frustratingly difficult, according to the lead researcher, Goggy Davidowitz. Davidowitz is an associate professor in the UA's Department of Entomology in the College of Agriculture and Life Sciences and a joint faculty member in the UA's Department of Ecology and Evolutionary Biology in the College of Science.
For example, biologists recently announced that in sequencing the giraffe's genome, they had identified 70 different genes that had undergone changes in their sequence compared with those of the giraffe's closest relative, the okapi. But how exactly are those genetic differences translated into the traits that make a giraffe different from an okapi?
A Hurdle to Overcome 
Figuring out which gene affects which trait — and in what contribution — is the hurdle that evolutionary biologists must overcome if they want to understand how organisms change over the course of their evolutionary history. 
"We know that natural selection can favor the evolution of certain traits — for example, a caterpillar's body size and the time it needs to develop into an adult — but we are less clear on how this happens," Davidowitz says. "The phenotype of an organism — the sum of its traits — is at least partly defined by its genes, but there is no one-gene, one-phenotype relationship. Instead, many traits are determined by more than one gene, just as a single gene may affect multiple traits."
To make things even more complicated, environmental conditions can influence how a genotype is translated into a phenotype because of genes being expressed differently, a phenomenon known as phenotypic plasticity. 
"With bioinformatics generating a huge body of gene sequences and multiple genes involved in traits, we are facing a big information overload," Davidowitz says. "So how do you map a trait to the genotype? We may know what genes are involved, but how do you get from those genes to the phenotype?"
To address that question, Davidowitz and his team imposed on Manduca sexta caterpillars 10 generations of simultaneous selection for two traits: body size and development time from larva to adult. They divided the animals into groups representing four combinations of those traits: large size/short time; large size/long time; small size/short time; and small size/long time. 
The researchers chose M. sexta caterpillars for their studies because their physiology has been well-studied. 
"Body size is highly correlated with fitness, as is the time it takes the larva to develop," Davidowitz explains. "Both traits are highly correlated with survivorship, and we know how they are regulated, so we are in a good position to impose selection on them in the lab."
According to previous work by other researchers, eight different molecular pathways are involved in caterpillar growth. 
"Combining those eight pathways in every possible way gives you 40,000 permutations to analyze — not a manageable number," Davidowitz says.
By distilling these combinations into key events that happen during the development of the animal, the new approach brought that number down to three — a very manageable number. 
"If you want to understand how an organism responds to selection, you need a model, and you can't do that with 40,000," Davidowitz explains. "But you can do it with three. And once you have that, you can start seeing how organisms respond to selection and make predictions of how they respond."
Events That Determine Body Size 
In their experiments mimicking evolution in the lab, the group focused on three key events that determine the final body size for an insect: the decision to stop growing; the actual cessation of growth; and how much growth occurs until growing actually stops. 
Using those parameters, the researchers were able to predict the caterpillars' body size in response to simultaneous selection to 100 percent and their development time to 93 percent.
Davidowitz says the study is one of few that addresses the evolutionary response of simultaneous selection on more than one trait.
"Because the three key events we focused on in the caterpillar's development encapsulate all of the underlying complexity, you can ignore that complexity up to a point," he says. "This opens the door to make a very complex problem simple, regardless of what species you are looking at." 
Co-authors on the study are Derek Roff at the University of California, Riverside, and Frederik Nijhout at Duke University.
The research report is published in the online edition of The American Naturalist.

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