Researchers discovered a ribonucleic acid molecule that produces black coloring on butterflies' wings in a study published last month.
GW biology professor Luca Livraghi used the genome-editing technique of clustered regularly interspaced short palindromic repeats, or CRISPR, to show that a RNA molecule called lncRNA generates black coloring on many butterflies' wings. Livraghi said his study is the first to describe the role of lncRNA molecule in the evolution of butterflies, as researchers had previously thought a protein was responsible.
"There aren't many examples of this described in the literature yet and even less so of these RNA molecules being implicated in evolution of these species," Livraghi said.
Livraghi said the CRISPR technology used in the study acts like "molecular scissors" that allow researchers to remove certain RNA sequences from an organism. He said researchers found that when they removed the specific RNA molecule, the black pigmentation did not appear on the butterflies' wings.
"If you have a gene that you hypothesize, for example, is involved in producing a certain color or a certain pigment on the wings, you can basically go in and use these little molecular scissors to mutate that gene to render it nonfunctional by cutting it with these scissors," Livraghi said. "Once you do that, you let the organism develop, and you can then look at the resulting effect it will have on the developing wings."
RNA molecules and DNA molecules both contain genetic material, but RNA is made of different sugars than DNA and their main function is to direct the body on what proteins to produce. Livraghi said researchers previously assumed that the RNA molecules' protein production caused the black pigments to appear on butterfly wings, but his study reveals that the protein generation step is not necessary.
"We were really surprised to find that actually you don't need that step of the generation of the protein, but that the RNA molecule itself is able to somehow translate its expression into specific pigments in the wings," Livraghi said.
Researchers used in situ hybridization, a laboratory technique where researchers place tissue on a glass slide and expose it to a dyed DNA sequence, which then binds to the matching sequence in the tissue, exposing the matching molecule to determine the stage the RNA molecule activates the trait. Livraghi said the process causes genes to "light up" and allows researchers to locate the active molecule when the black pigment appears.
"If you actually dissect the butterfly wing before its final development, the first few days of metamorphosis, there won't be any pigment whatsoever in that wing," Livraghi said. "It'll look basically blank, but however, genes are being turned on and off in specific sequences and specific places."
Experts in insect biology and genetics said the methods of the study show how researchers studying organisms can use gene editing technology to identify molecules that spur certain traits in organisms.
Marc Halfon, a professor of biochemistry at the University at Buffalo, said the technology used in this study is "powerful," and the results that researchers find about one species can indicate similar genetic processes in other species that result in similar traits.
"The applications are very broad even though you're looking just at a single species, at a single process here, the information we glean from that could really have a lot of implications for many things that are going on," Halfon said.
Michael Perry, an assistant professor of biology at the University of California, San Diego, said the researchers' use of in situ hybridization to locate the noncoding RNA molecule -- a molecule that does not produce proteins -- is helpful because researchers often locate RNA molecules by creating an antibody for the protein it creates, and in situ hybridization allows researchers to see the molecule without the protein.
"Through a combination of slightly more clean genetic experiments, knocking out this sequence specifically and then finding out where this RNA was expressed and showing that it's expressed in patterns complementary to the wing patterns, it was nice," Perry said.
Perry said the study was successful due to the use of in situ hybridization. He said researchers often accidentally remove more genes than they mean to through CRISPR, which makes them wrongly attribute the cause of the trait changes.
"So when you inject sometimes you do damage just to the whole local region," Perry said. "Sometimes you knock out a bigger piece of a chromosome, and so what the new studies have done has been to identify the actual mechanism."