The mystery of calico cats is morethan skin deep. The broad blackand yellow patches in their fur are

the outward manifestations of a moresubtle genetic quirk. True calicoes arefemales, and like all female mammals,they carry two X chromosomes in theircells. Early in development, however,each embryonic cell randomly selectsone X for future use and signals theother to condense permanently into aninert mass called a Barr body. (In thisway, females achieve parity with males,which have only one X chromosome anda largely inactive Y.) In calico cats theresulting mosaicism is visible becauseeach of their X chromosomes carries adiÝerent pigment gene.

After three decades of work, re-searchers are beginning to understand

how mammalian cells manage to turnoÝ an entire chromosome. The key ap-pears to be a gene on the inactive X thatproduces an RNA molecule of unknownfunction. There are several explanationsfor how the gene accomplishes its feat.ÒMy personal bias,Ó remarks Carolyn J.Brown, one of the discoverers of thegene, Òis that the RNA molecule is im-portant in forming some kind of cageor structure that segregates the X andallows inactivation.Ó

Brown and other members of the lab-oratory of Huntington F. Willard at CaseWestern Reserve University made theirdiscovery while looking at gene expres-sion on the Barr body. A few genesÑabout a dozen are now known in hu-mansÑevade the general ÒoÝÓ signaland therefore remain active on both Xchromosomes. Yet in 1990 WillardÕsgroup found one gene that had a uniquedistinction: it was active only on theBarr body. Moreover, the gene was lo-cated in the small region of the X chro-mosome that previous research had de-

termined was essential to X inactivation.Those characteristics hinted that the

gene, which WillardÕs group dubbed theX inactive-speciÞc transcript gene (Xist),might play a pivotal part in turning oÝthe X chromosome. Willard and Brownand their colleagues released word ofXist in January 1991. Several months lat-er Sohaila Rastan and Neil BrockdorÝand their colleagues at the Medical Re-search Council in Harrow, England, re-ported discovering a corresponding Xist

gene in mice.Last October in Cell, both the Willard

and Rastan teams published their anal-yses of the human and mouse forms ofXist. The genes produce exceptionallylarge RNA molecules, and the humanand mouse RNAs are generally similarto each other. Yet unlike most RNA,which leaves the cell nucleus and istranslated into protein, the Xist RNAdoes not carry information for makingproteins at all. Indeed, as WillardÕs ex-periments using ßuorescent molecularprobes showed, the Xist RNA never

Kitty, We Shrunk Your Brainhelsea Clinton and other cat lovers, don’t take thisthe wrong way, but the brains of your pets aren’t allthat they used to be. The tabby curled on the sofa

has lost almost a third of the neurons of its more robustPleistocene ancestor. Such is the conclusion of Robert W.Williams of the University of Tennessee and Carmen Cava-da and Fernando Reinoso-Suárez of the Independent Univer-sity of Madrid. Their finding does not mean that cats havebecome more stupid—mercy, no. Rather it reveals a mech-anism that may facilitate certain types of rapid evolutionarychange.

The brains of domestic cats are not unusually tiny. If thebrain sizes of lions, ocelots and all other feline species areplotted against their body weights, the domestic cat’sbrain falls neatly on the curve. “Its brain is exactly the sizeyou’d expect based on its body size,” Williams says. But,he observes, “even though people had studied those curvesad nauseam, nobody ever really knew what they meant interms of cell number and cell size. What does it mean tosay that the brain got smaller? Did it lose parts, or did theparts get smaller?”

In search of an answer, Williams, Cavada and Reinoso-Suárez compared the visual systems of modern house catswith those of Spanish wildcats (Felis sylvestris tartessia).Fossil evidence indicates that the Spanish animals are vir-tually indistinguishable from the wildcats that roamednorthern Africa and Europe 20,000 years ago. The Span-ish wildcats are taller and usually about twice the weight ofthe more familiar F. catus. Unlike feline homebodies, whichare primarily nocturnal hunters, the wildcats hunt by day.

The clear-cut results of the comparison showed that “thereduction in brain weight involved the loss of brain cells,”Williams says. Domestic cats had only about half as manyneurons in the ganglia (nerve clusters) that connect theirbrain to their retinas. The wildcats had about 50 percentmore neurons in their lateral geniculate nuclei, the brain

structures that first receive signals from the optic nerves.In the retinas of the wildcats, the density of the cone pho-toreceptors—which make color vision and vision in brightlight possible—was also more than twice as great. The re-searchers are confident that similar losses have occurredthroughout the cat brain.

Twenty thousand years is relatively little time for somuch change to have evolved. Williams thinks he and theothers have found “a scintilla of evidence” about the mecha-nism. When they examined a wildcat embryo, they foundthat its brain contained approximately the same numberof neurons as that of a domestic cat embryo. “So it looksplausible to us that the way the domestic cat got a smallerbrain was by losing more cells rather than by producingfewer cells,” Williams concludes.

Programmed cell death is a common feature of embry-onic development for most animal species. In domesticcats, about 80 percent of the cells in the visual ganglia diebefore or shortly after birth—far more than in other verte-brates. Conceivably, then, the smaller modern cat speciesmight have arisen fairly rapidly through a change in thedevelopmental program that generally raised the amountof cell death. Williams cautions, however, that the idea“still really needs to be nailed down.”

To Williams’s knowledge, the study is the first attemptto compare species within an evolutionary lineage. Theshrinkage in cats is not entirely human doing: most of itoccurred long before people began domesticating catsless than 5,000 years ago. Indeed, because many mam-mals have become smaller since the last ice age, furtherwork on other animals may find similar massacres of graymatter. Williams believes dogs are likely to be another ex-ample of “absurdly rapid evolution,” much of it at the handsof human breeders. Cat fanciers may find some consola-tion in that thought: Who knows how much was deleted enroute from Great Danes to Chihuahuas? —John Rennie

C

Spot Marks the XIn females, one chromosomemay lock itself inside an RNA

SCIENTIFIC AMERICAN April 1993 29Copyright 1993 Scientific American, Inc.

seems to leave the nucleus. Instead itclusters tightly around the inactivatedX chromosome that makes it.

Those results suggest several modelsfor how inactivation might occur. Oneis that as the Xist RNA is produced, itbinds to the chromosome, perhaps inassociation with other molecules. Theresulting cage of RNA may directly in-capacitate most genes. Alternatively, thepresence of the RNA might enable thechromosome to interact with other fac-tors on the nuclear membrane or else-where that deactivate it. Yet anotherpossibility is that the RNA itself doesnot serve a function but that the act oftranscription in that region induces con-formational changes in the chromosomethat lead to its inactivation.

In recent months the association be-tween Xist and X inactivation has beenfurther strengthened by Larry J. Shapiroof the University of California School ofMedicine at San Francisco, Jacob Wahr-man of the Hebrew University of Jerusa-lem, John R. McCarrey and Donald D.Dilworth of the Southwest Foundationfor Biomedical Research in San Antonioand others. In independent studies,those investigators have found that thetranscription of Xist precisely mirrorsthe inactivation of X chromosomes invarious tissues.

In January, Graham F. Kay, anothermember of RastanÕs group, announcedthat the transcription of Xist in early em-bryonic cells seems to precede X inacti-vation by a day or so. ÒThat implies tous that Xist expression is not simply aconsequence of X inactivation and sup-ports the case that it could be causal,ÓBrockdorÝ comments. Brown agrees thatXist is Òa smoking pistolÓ but empha-sizes that its importance during inacti-vation remains to be proved.

New experiments should settle thatissue. ÒThe idea weÕre working on is toknock out the Xist genes in an embry-onic stem cell,Ó BrockdorÝ explains. ÒIfXist is required, we should abolish theability of those cells to undergo X inac-tivation.Ó Investigators can also insertactive copies of Xist into cells to seewhether neighboring genes are shut oÝ.

Other questions also remain. ÒIf Xist

is involved in X inactivation, then thereis something that is turning it on orturning it oÝ,Ó Brown says. Researchersare keenly interested in determining howthe Xist RNA interacts with the chro-mosome. At this point, they can onlyspeculate about how the informationconcerning which X chromosome shouldbe inactivated is passed from one cellto its progeny. Until those answers arefound, researchersÕ understanding of Xinactivation is likely to stay as patchy asthe calico cat herself. ÑJohn Rennie

30 SCIENTIFIC AMERICAN April 1993 Copyright 1993 Scientific American, Inc.