Virus-Resistant Transgenic Horticultural Crops: Safety Issues and Lessons from Risk Assessment Studies - Transgenic Horticultural Crops: Challenges and Opportunities - page 280

own.potential. risks?.Do.they.alleviate.a.previous.concern.that.was.no.longer.seen.
as.a.problem.only.to.raise.new.issues.that.are.potentially.more.dificult.to.assess?.In.
the.following.section,.we.will.describe.some.of.the.latest.approaches.toward.virus.
resistance.in. transgenic.plants. and.discuss.if. and.how.these.technologies.alleviate.
some.of.the.existing.concerns.
f uture t rendS
Given. the. discovery. and. elucidation. of. the. antiviral. pathways. of. RNA. silencing,.
many.new.approaches.have.been.used.to.develop.transgenes.more.likely.to.stimu-
late.RNA.silencing.via.the.design.of.sophisticated.transgenes..Since.RNA.silencing.
theoretically.is.stimulated.in.a.plant.cell.by.the.presence.of.dsRNA,.many.of.these.
transgene. constructs. attempt. to. transcribe. RNA. molecules. that. are. more. likely. to.
form.dsRNA.structures..One.strategy.for.accomplishing.this.is.the.use.of.inverted.
repeats,. which. involves. the. creation. of. a. transgene. containing. two. copies. of. the.
viral.target.complementary.DNA.(cDNA).sequence.separated.by.a.spacer.DNA.of.
some.length. 25,125 .After.transcription,.it.is.thought.that.the.resulting.RNA.will.form.
a. hairpin. with. the. inverted. RNA. forming. a. double-stranded. structure.. The. use. of.
introns.is.a.similar.approach.in.that.two.inverted.regions.of.viral-derived.cDNA.are.
separated. by. an. intron. 126 . Once. the. intron. is. spliced. by. host. machinery. following.
transcription,.a.dsRNA.structure.is.formed..Another.approach.consists.of.producing.
two.complimentary.pieces.of.RNA.which.can.then.form.a.dsRNA.from.bidirectional.
promoters. 127 .These.new.strategies.have.been.shown.to.produce.a.signiicantly.higher.
proportion.of.virus-resistant.transgenic.plants.than.the.use.of.full-length.coat.protein.
transgenes. 25,125,126 .In.addition,.they.hold.a.seeming.advantage.over.a.full-length.coat.
protein.gene.in.the.sense.orientation.as.they.are.generally.unable.to.produce.a.func-
tional.protein,.alleviating.concerns.arising.from.the.presence.of.the.coat.protein.in.
plant. material.. Coat. protein. expression. can. also. be. prevented. by. using. transgenic.
approaches.involving.a.transgene.that.produces.an.RNA.product.which.is.untrans-
latable,. either. because. it. lacks. the. necessary. translation. start. codon. for. ribosomal.
processing.or.because.it.is.oriented.in.antisense.directions..It.should.be.noted.that.
the.use.of.introns,.often.derived.from.plants.themselves,.potentially.poses.the.risk.
of. silencing. host. genes. from. where. the. intron. was. derived,. if. the. RNA. silencing.
machinery.incorrectly.processes.the.transgene.RNA..A.similar.concern.applies.to.
the.siRNA.technology.overall. 128
Another.approach.utilizing.the.knowledge.of.viral.silencing.is.to.produce.resis-
tance.by.using.modiied.plant.microRNA.(miRNA).cistrons.to.produce.a.range.of.
artiicial. antiviral. miRNAs. 129,130 . The. durability. of. this. approach. compared. to. the.
use.of.longer.dsRNA.approaches.has.not.been.demonstrated. 131
Some. nonviral. sources. of. virus. resistance. have. also. been. investigated.. These.
would.theoretically.alleviate.concerns.about.synergism,.recombination,.and.transen-
capsidation..These.include.the.transfer.of.host.resistance.genes.against.viruses.into.
other.hosts.via.genetic.engineering,.or.the.silencing.of.host.genes.that.are.necessary.
for.viral.replication. 25 .The.use.of.plant-generated.antibodies.against.viruses,.which.
failed.to.progress.for.many.years.though.initially.perceived.as.promising,.has.recently.
been.shown.to.be.effective.in.controlling.viruses.in.plants.expressing.the.transgenes.