Notebook

WHY X Y? Cloning is not just for females anymore. The same University of Hawaii researchers that cloned 50 healthy female mice a year ago (T. Wakayama et al., "Full-term development of mice from enucleated oocytes injected with cumulus cell nuclei," Nature, 394:369-74, July 23, 1998) recently reported the first-ever cloning of an adult male mouse (T.W. Wakayama, R. Yanagimachi, "Cloning of male mice from adult tail-tip cells," Nature Genetics, 22:127-8, June 1999). Named "Fibro" after the fibroblast cells from which he was cloned, the mouse was the first documented case of a male mammal cloned from adult cells, cells taken from the tip of a mouse's tail. Past successful clonings have used cells from the female reproductive system. Fibro was one of only three mice that reached full term out of 274 attempted clonings. The other two died soon after birth. Senior author of the study, Ryuzo Yanagimachi, a professor of anatomy and reproductive biology at the University of Hawaii Medical School, suggests that the most important potential application of his findings right now may be the ability to preserve endangered male species through cloning. But scientists must first increase the rather low success rate. "We need another breakthrough to find out if we can improve this," says Yanagimachi. An important clue to why these cloning efforts were not more successful may be that, as noticed in previous studies, cloned placentas are twice as large as normal placentas. "I don't know what this means," comments Yanagimachi, "but I suspect that some communication between placenta and fetus is not going well." The problem, therefore, may be with the placenta and not with the fetus, he says. --Eugene Russo PRIONS IN FUNGI? Now that the idea of conformational changes in prions causing transmissible spongiform encephalopathies is gaining acceptance, some researchers are reevaluating past examples of possible protein infectivity in species other than mammals. Philippe Silar and Marie-Josee Daboussi, lecturers at the Institute of Genetics and Microbiology at the University of Paris South, reviewed evidence of prionlike molecules in filamentous fungi that might finally make sense as proteins that assume more than one conformation, one of which transmits a change in phenotype (P. Silar and M.-J. Daboussi, "Non-conventional infectious elements in filamentous fungi," Trends in Genetics,15:141-4, April 1999). The earliest references predate description of the double helix. "In model systems like Paramecia and yeast, elements with similar patterns were known for a long time. But at that time, explaining their nature was out of the reach of technology," explains Silar. Such changes are easy to see in fungi, which consist of cells strung into many long threads, or hyphae. Silar and Daboussi tracked down two Ph.D. theses, from 1965 and 1979, that describe changes in color and the growth pattern of hyphae that are transmitted without involvement of a nucleic acid. Like the mammalian prions, similar proteins in fungi "are caused by altered conformation of a normal cellular protein. But unlike the mammalian protein, prions in fungi are not infectious like ... viruses are," Silar adds. --Ricki Lewis HPV LINKS Human papilloma virus (HPV) is present in most cases of cancer, but scientists have remained unsure what role the virus plays in the cancer's development. Luis Villarreal, professor of biological sciences at the University of California, Irvine, School of Biological Sciences, and colleagues found that by grafting human cervical tissue into mice, they could trace the development of cervical cancer and experiment with factors that increase its risk (J.A. Taylor et al., "Immunohistochemical analysis, human papillomavirus DNA detection, hormonal manipulation, and exogenous gene expression of normal and dysplastic human cervical epithelium in severe combined immuno- deficiency mice," Journal of Virology, 73:5144-8, June 1999). "This human-mouse experimental model allows us to create cancer cells and study how they grow, which wasn't possible before. Cervical cancer only occurs in humans, so animals and cells could never give us the exact cause," Villarreal explains. The researchers found preliminary evidence of the HPV in nearly all the abnormal cervical growths in the model. In addition, they found that high doses of estrogen appeared to result in noncancerous growths in the cervical tissue. As they continue their studies, they expect to learn more about how the virus behaves. Viruses typically cause disease by interweaving their DNA with the DNA of host cells. By mapping out how the virus integrates itself into cervical cells, the scientists hope to gain greater understanding of how the virus contributes to cancer development. --Ilene Schneider Three-dimensional molecule model of human MMP-2 X-ray crystal structure CRYSTALLIZING MMP-2 The recently elucidated structure of an enzyme known to have a critical role in tumor growth could prove to be a useful tool for the design of novel drugs that can stop cancer metastases. After seven painstaking years of X-ray crystallography research, a team from the Karolinska Institut in Stockholm and Biostratum Inc. of Research Triangle Park, N.C., recently successfully crystallized a molecular target that may be integral to the design of a number of antiangiogenesis and other anticancer therapies (E. Morgunova et al., "Crystal structure of human MMP-2 reveals the exposed propeptide loops ready for proteolytic activation," Science, 284:1667-70, June 4, 1999). Called matrix metalloproteinase 2 (MMP-2), the enzyme is one of a family of proteinases that catalyze the degradation of the extracellular matrix to allow for cell movement--a function that makes MMPs integral to the process of tissue growth and wound repair. According to senior author Karl Tryggvason, a professor of medical chemistry at Karolinska and cofounder of Biostratum, crystallizing MMP-2 was a difficult task because of the protein's considerable size and multiple domains. "I don't know exactly why we succeeded," says Tryggvason, who adds that several major pharmaceutical companies were in the running. "We were, I guess, lucky." Although researchers had previously crystallized parts of other MMPs, this was the first crystallization of an entire MMP structure. Although MMP inhibitor clinical trials are already in the works, the molecules tested generally act too broadly and have unwanted side effects. These recent findings should enable the characterization of new, more specific anticancer compounds. The newly elucidated structure should also serve as a reference in future attempts to solve the structures of other MMPs. --Eugene Russo TICK GLUTTONY Ixodid ticks attach to their vertebrate hosts for several days, even weeks, consuming huge blood meals, which can increase their body weight 100-fold. Since the ticks are exposed to host inflammatory and immune responses, they need mechanisms to fight host defenses while feeding. Inflammation leads to a sensation of itch or pain, prompting the host to remove the ticks by grooming. Three histamine-binding proteins (HBPs) from the saliva of Rhipicephalus appendiculatus ticks were characterized and reveal a strategy by which ticks prevent inflammation (G.C. Paesen et al., "Tick histamine-binding proteins: isolation, cloning, and three-dimensional structure," Molecular Cell, 3:661-71, May 1999). Histamine, a principal mediator of inflammatory reactions, is released in response to tissue damage. It is mainly secreted by mast cells and basophils and binds to H1 and/or H2 receptors on the surface of target cells involved in inflammation. The crystallographic structure and biological activity of the tick HBPs indicate that they sequester histamine at the wound site, outcompeting the host's histamine receptors for the ligand and feeding successfully. The crystal structure of a histamine-bound HBP revealed a lipocalin fold novel because it contains two binding sites with different affinities for histamine. Whereas lipocalins mostly bind hydrophobic compounds, the binding pockets of the tick HBP were designed to accommodate hydrophilic (histamine) ligands. Guido Paesen from the Natural Environment Research Council in the United Kingdom comments, "By mopping up free histamine, soluble HBPs offer a new approach to the control of histamine-based diseases independent of membrane-bound histamine receptors." Traditional drugs used to control the pathological effects of histamine such as allergies and peptic ulcers, act by blocking the histamine-binding sites of either H1 or H2 receptors. --Nadia S. Halim MICROORGANISMS CLEAN UP Microorganisms living at the bottom of lakes and streams may offer help in protecting surface drinking water from contamination by the gasoline additive MTBE (methyl tertiary-butyl ether). Scientists with the U.S. Geological Survey in Columbia, S.C., have found the first instance of naturally occurring microorganisms able to degrade MTBE to nontoxic byproducts under aerobic conditions (P. Bradley et al., "Aerobic mineralizations of MTBE and tert-butyl alcohol by stream-bed sediment microorganisms," Environment and Science Technology, 33:1877-9, June 1, 1999.) Automobiles guzzle more than 4 billion gallons of MTBE, a potential carcinogen, each year (S.P. Hoffert, "Haze of uncertainty surrounds gas additive," The Scientist, 12[13]:1, June 22, 1998). MTBE evaporates into the atmosphere and contaminates rain, which ends up in streams and lakes. Also, leakage from underground gasoline storage tanks leads to contaminated groundwater discharged into streams. "This finding offers the possibility that under the right conditions there is a way for MTBE to degrade naturally in the environment," comments Charles Kulpa, professor at the University of Notre Dame, Ind., and director for the center for bioengineering and pollution control. Sediment samples from stream beds were used to degrade 14C radiolabeled MTBE. Within 105 days 73 percent of the MTBE was converted to radiolabeled carbon dioxide. Scientists also observed that tertiary-butyl alcohol, another fuel additive, degraded in a similar manner. Man-made sediments containing the microrganisms may potentially be used to clean contaminated groundwater. --Nadia S. Halim Conus snail feeding on a worm TOXIC EVOLUTION The phrase "snail's pace" now has new meaning. The venomous gastropod Conus has the world's fastest known mutating genes, according to a recent study (T.F. Duda Jr., S.R. Palumbi, "Molecular genetics of ecological diversification: duplication and rapid evolution of toxic genes of the venomous gastropod Conus," Proceedings of the National Academy of Sciences, 96:6820-3, June 8, 1999). The speedy evolution occurs in genes that produce small toxic proteins the cone snail secretes to stun its supper. Harvard University researchers examined the genetic sequences of toxins from two distantly related cone snails collected from different sites around Oahu, Hawaii. Thomas F. Duda Jr., a Harvard University biology graduate student, estimates that the cone-snail toxin genes mutate five times faster than the highest rate for a mammalian gene and three times quicker than the top rate for a fruit fly gene. Those mutations have resulted in a variety of toxins that vary drastically in terms of both chemical composition and size. "You've got about 500 species and at least a dozen toxins per species." Duda speculates that such a large variety of toxins has allowed Conus to hunt organisms ranging from worms to mollusks to fish. Conus may also deploy rapid mutations as a preemptive strike against a potential meal's ability to develop resistance, he hypothesizes: "There may be some kind of arms race going on between predator and prey." --Paul Smaglik

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