Taming the sindbis virus

For both cell culture and gene therapy, viruseshave long been considered an efficient tool forintroducing foreign genetic material intocells. Unfortunately, the viruses most suitedto these jobs tend to kill the cells they infect,making them useful only for transient geneexpression. To get around this problem, ateam of researchers led by microbiologistCharles Rice at the Washington UniversitySchool of Medicine (St. Louis, MO) hasdeveloped a noncytopathic strain of Sindbis,a lytic virus that is already widely used intransient expression systems. The scientists,whose work is described in the October issueof PNAS (95:12989–12994, 1998), transfectedcells with recombinant viral RNA containinga puromycin resistance gene, and then grewthe cells in the presence of puromycin. Onlycells carrying a noncytopathic mutant viralRNA could survive this double selection, andthe resulting mutations were mapped andincluded in a new gene expression vector. TheWashington University team is now focusingon using the system to express genes at physi-ological levels for experiments in basic cellbiology and virology.

Blocking an MS message

Using a mouse system that closely mimics theprogressive demyelination found in multiplesclerosis (MS), a condition which afflicts anestimated 300,000 people in the US alone,researchers have developed a promising newform of immunotherapy that may help pre-vent patient relapses. The scientists, led byAbdolmohamad Rostami and CrisConstantinescu at the University ofPennsylvania (Philadelphia, PA), discoveredthat the concentration of the cytokine inter-leukin-12 (IL-12) increased in the mice duringexperimentally induced relapses. The teamreasoned that “if the level of IL-12 was up dur-ing the relapses and acute attacks. . .wethought that if we could decrease the level, wemight be able to prevent the relapse,” explainsRostami. As predicted, mice injected with amonoclonal anti-IL-12 antibody were protect-ed from relapses, and injecting purified IL-12could induce a relapse. The antibodies did notappear to have any side effects on the mice,and a single injection appeared to providelong-term protection. The researchers, whosework appears in the November 1 issue of theJournal of Immunology (161:5097–5104,1998), are now developing similar anti-IL-12antibodies for testing in humans.

NATURE BIOTECHNOLOGY VOLUME 16 DECEMBER 1998 1299

IN BRIEF

RESEARCH NEWS BRIEFS

Lilliputian lab

In an effort to make DNA analysis morestreamlined and cost effective, a team of engi-neers and geneticists from the University ofMichigan, Ann Arbor have developed a devicethat can perform enzymatic reactions on DNAand analyze the results in an integrated systemthe size of a single silicon chip. As reported inScience (Burns et al. 282:484–487, 1998), thedevice was used to digest and amplify a regionof Mycobacterium tuberculosis DNA. The vari-ous systems required were built onto the chipusing standard photolithographic techniques,in a process conceptually similar to stenciling.“The device is unique due to the fact that ituses complex integrated systems where they allwork together simultaneously,” says Dr. DavidBurke, a coauthor of the study. Included on thechip are a nanoliter injector, a sample mixingand positioning system, a temperature-con-trolled chamber, an electrophoretic separation

The tet system just gotbetter

The tetracycline (tet) regulatable system forgene induction/repression, while conceptual-ly brilliant, has been fraught with technicaldifficulties, including high basal levels ofexpression in the absence of induction. NowHelen Blau and colleagues at the StanfordUniversity School of Medicine have devel-oped a reversible tet-inducible retroviral sys-tem (RetroTet-ART) in which activators andrepressors are expressed together in cells.Guided by the crystal structure of tetR, a tran-srepressor was engineered with a dimeriza-tion domain distinct from the transactivator.When the transactivator and the transrepres-sor were expressed in vitro, no complexescontaining both proteins were observed in gelretardation and immunoprecipitation stud-ies. This allowed the transrepressor to bindand repress gene expression in the absence ofdox, whereas the transactivator binds andinduces gene expression only in the presenceof dox. The RetroTet-ART system was thentested in cells by reversibly expressing p16, a

Research News Briefs written by AlexanderCastellino, Alan Dove, and Margret Einarson.

protein known to arrest growth. In the pres-ence of increasing dox, cell growth decreased.The growth arrest was reversed upon doxwithdrawal. Helen Blau says, “The system isremarkably easy to use, as all the componentscan be introduced into cells in less than twoweeks. It will prove a boon when it comes toexpressing products toxic to cells, proteinsthat arrest cell growth, and regulating proteinexpression in gene therapy and transgenicanimals.” These findings are reported inNature Genetics (20:389–393, 1998)0

system, and a fluorescence detector, at a finalproduction cost of approximately six dollarseach. Further development of this technologywill allow DNA analysis to be performed “onsite,” whether that means a doctor’s office, therain forest, or the farm.

In work that has far-reaching implica-tions for animal husbandry and thepreservation of endangeredspecies, researchers report in theOctober issue of Animal Repro-duction Science (53:265–275,1998) a technique for trans-planting cryopreserved ovariantissue from elephants into mice.The tissue, collected in SouthAfrica and frozen, was able toundergo normal develop-ment in immune-deficientmice, producing matureovarian follicles and, in atleast one case, an oocyte. Inthe past, female germplasmhas been notoriously diffi-cult to preserve, compli-cating many species con-servation efforts. “As far as

we can tell, this general procedureshould work for any mammalian

species except the egg-layingMonotremes. Certainly, wewould hope that this procedurewould be used for other endan-

gered species,” says John Critser,the scientific director of theCryobiology Research Institute atIndiana University and seniorauthor on the study. The scientistshope to fine-tune the system to

reliably produce mature oocytes,which could then be used for invitro fertilization in breeding

programs. While emphasizingthat several technical hurdles

remain, Critser argues that “geneticmaterial from rare and endangered

species that would otherwise be lost can bepreserved at this point [by freezing].”

Endangered egg surrogates

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