When Rudyard Kipling told how the leopard got his spots, he missed the mark. Leopards have “rosettes”; spots are for cheetahs, says Gregory Barsh, a geneticist at the HudsonAlpha Institute for Biotechnology. But whatever you call the markings, how wild cats and their domestic counterparts acquire them has long been a mystery. Now, Barsh and his colleagues have found an answer. In so doing, they have shown that a 70-year-old theory explaining patterns in nature holds true for fur color in cats, and likely other mammals as well.
“This is an important paper unveiling part of the genetic basis [of ] coat color markings so prominent in many mammals,” says Denis Headon, a developmental biologist at the Roslin Institute. It also offers a glimpse of how those genes operate during development, forming what he calls a “highly adaptable mechanism” that responds to genetic tweaks to produce diverse coat patterns, from stripes to spots.
Biologists have identified hair follicle cells as the source of the black, brown, yellow, and red pigments that color hair or fur. “But we didn’t know when and where the process of the establishment of the color pattern took place,” Barsh says.
In 1952, computing pioneer Alan Turing suggested molecules that inhibit and activate each other could create periodic patterns in nature if they diffused through tissue at different rates. Thirty years later, other scientists applied his theory to develop a hypothesis about how spots, stripes, and other color patterns form during development. In this scheme, activator molecules color a cell but also trigger the production of inhibitors, which diffuse faster than the activators and can shut off pigment production. Last year, that idea was proved correct in plants called monkeyflowers: Researchers showed that dark, activated speckles on the petals become ringed with unpigmented tissue as inhibitors spread. And researchers had shown molecules following the Turing pattern help trigger the development of hair follicles in mice. But how coat color develops in mammals remained largely mysterious because mice and other easy-to-study lab animals lack spots or stripes.
So Barsh’s team turned to domestic cats to track the identity of molecular activators and inhibitors of coat color. A decade ago, they tracked down a gene, Tabby, that, when mutated, gives tabby cats black blotches instead of their usual dark stripes. Hudson-Alpha geneticist Christopher Kaelin found that same mutation in king cheetahs whose spots were unusually big and blotchy, suggesting the same genes color both wild and domestic cats.
To see what other genes and their mutations operate during development, Kaelin and HudsonAlpha colleague Kelly McGowan spent several years collecting discarded tissue from clinics that spay feral cats, which are often pregnant. They first noticed temporary thickenings of the skin of 28- to 30-day-old embryos, where black stripes would later appear in the fur. “There’s a change [in the skin] that precedes and mimics what you observe in adult [fur],” McGowan explains.
The researchers then isolated and sequenced the active genes in individual skin cells of early embryos. At about 20 days old, embryos showed a sharp increase in the activity of several genes involved in a key developmental pathway, known as Wnt signaling, in skin areas destined to briefly thicken before the area becomes permanently dark. One of the most active genes was Dkk4, as they reported on 16 November in a preprint on bioRxiv. The team also found that mutations that inactivated Dkk4 accounted for the loss of distinct markings in Abyssinian and Singapura breeds, making their spots too small to distinguish. Tabby and Dkk4 “are in the same pathway,” and likely work in both domestic and wild cats, Barsh explains, though he doesn’t yet know how they are connected.
Dkk4 is a known inhibitor of Wnt signaling, which helps determine cell fates and spurs cell growth during development in many animals. The team found that in domestic cats, Wnt and Dkk4, respectively, are the activator and inhibitor. In dark skin, they exist in about equal amounts. But in paler areas, the faster moving Dkk4 protein most likely turns off Wnt, shutting down pigment production and thereby generating stripes, just as Turing’s theory had predicted. “It is remarkable, although not altogether surprising, that we see Wnt-Dkk4 signaling again playing a critical early role,” says Larissa Patterson, a developmental biologist at Rhode Island College.
“This paper provides thought-provoking insights into potential mechanisms of pattern diversity in wild cats,” Patterson adds. It “greatly adds to the evidence” that this process is at work in cats and, most likely, other mammals, agrees Roland Baddeley, a computational neuroscientist at the University of Bristol.
Researchers had already shown the Turing mechanism involving Wnt and Dkk4 sets up the formation of hair follicles—but not coat color—later in mouse development. Barsh’s team, however, found that the color pattern in cats and possibly other mammals is established well before hair follicles appear, suggesting early color patterns may guide hair follicle pigmentation.
That simple interactions among well-known molecules can explain the variety of coat color patterns in mammals is an example of nature’s thriftiness, Headon says. “It suggests that the same molecules and pathways are likely to be reused for patterning of very different structures and at very different scales to form the intricate elements of the vertebrate anatomy.”
sciencemag.org, 2 December 2020