Science cover story: Gene-edited butterfly mutants reveal secrets of ancient "junk" DNA

A new study explains how DNA located between genes — known as "junk" DNA or noncoding regulatory DNA — fits into a basic program that has been preserved for tens to hundreds of millions of years, while allowing wing patterns to evolve
extremely quickly.
In the October 21 issue of Science magazine, "Deep Cis Modulation Homology of Butterfly Wing Patterns" was published
as a cover story.

The study supports the idea that an ancient color pattern has been encoded in the butterfly genome, and that noncoding regulatory DNA works like a switch, turning some modes on and off others
.

"We were curious to see how the same genes make these very different-looking butterflies," said
Anyi Mazo-Vargas, Ph.
D.
, lead author of the study and a former graduate student in the lab of Robert Reed, a professor of ecology and evolutionary biology in the College of Agriculture and Life Sciences.
Mazo-Vargas is currently a postdoctoral fellow
at George Washington University.

"We found that there is a very conserved set of switches (non-coding DNA) that work in different locations, are activated and drive genes," Mazo-Vargas said
.

Previous work in Reed's lab has identified key color-patterning genes: one (WntA) controls the stripes, and the other (Optix) controls the color and iridescent colors
of the butterfly's wings.
When the researchers disabled the Optix gene, the wings took on a black color, and when the WntA gene was deleted, the stripe pattern disappeared
.

Pattern detail of the wings of the Agraulis vanilla butterfly, altered by modifications to noncoding DNA sequences by the gene-editing tool CRISPR/cas9
.

This study focused on the effect of
non-coding DNA on the WntA gene.
Specifically, the researchers conducted experiments on 46 noncoding elements in five species of butterflies
, the largest family of butterflies.

To allow these noncoding regulatory elements to control genes, the tightly wound DNA coils become loose, which is a signal that regulatory elements are interacting with genes, activating them, or in some cases turning them off
.

In this study, the researchers used a technique called ATAC-seq to identify regions
in the genome where this unraveling occurs.
Mazo-Vargas compared the ATAC-seq profiles of the wings of five butterfly species to identify genetic regions
associated with wing pattern development.
They were surprised to find that a large number of regulatory regions were shared
among very different butterfly species.

Mazo-Vargas and his colleagues then used CRISPR-Cas gene-editing technology to disable 46 regulatory elements at once to see the effect
on wing patterns when these noncoding DNA sequences were disrupted.
After deletion, each non-coded element changed one aspect of
the butterfly wing pattern.

The researchers found that in four species — Junonia coenia (eye-shaped spot-winged butterfly), Vanessa cardui (ramie butterfly), Heliconius himera (black sleeve butterfly) and Agraulis vanillae (bay fritillary) — each noncoding element has a similar function for the WntA gene, proving that they are ancient and conserved, likely originating from a distant
。

They also found that D.
plexippus used four other different regulatory elements to control its WntA gene, possibly because it had historically lost some genetic information and had to reinvent its regulatory system to form unique color patterns
.

Reed said: "We have come to realize that most evolution occurs in mutations in
these non-coding regions.
I hope this paper will become a case study showing how people can use a combination of ATAC-seq and CRISPR to start asking about these interesting regions in their own research systems, whether they work
on birds, fruit flies, or nematodes.
”

Theodore Morgan, project director at the National Science Foundation, said: "This study is a breakthrough in our understanding of genetic control of complex traits, not only in
butterflies.
This study not only shows how instructions for color patterns in butterflies have been highly conserved throughout evolutionary history, but also reveals new evidence
governing how DNA fragments positively and negatively affect traits such as color and shape.
”

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