The Silence of Genes in Transgenic Plants
M A I K E S T A M, J O S E PH N. M. M O L and J A N M. K O O T E R* Department of Molecular Genetics, Institute for Molecular Biological Sciences, BioCentrum Amsterdam, Vrije Uni ersiteit, De Boelelaan 1087, 1081 HV Amsterdam, The Netherlands
Received : 31 May 1996 Accepted : 16 September 1996
Ingenetically modiﬁed plants, the introduced transgenes are sometimes not expressed. They can be silenced. Transgenes can also cause the silencing of endogenous plant genes if they are suﬃciently homologous, a phenomenon known as co-suppression. Silencing occurs transcriptionally and post-transcriptionally but silencing of endogenous genes seems predominantly post-transcriptional. If viraltransgenes are introduced and silenced, the posttranscriptional process also prevents homologous RNA viruses from accumulating ; this is a means of generating virus-resistant plants. A major goal of current research is to dissect the mechanism(s) of these sequence-homologydependent gene silencing phenomena. Various factors seem to play a role, including DNA methylation, transgene copy number and therepetitiveness of the transgene insert, transgene expression level, possible production of aberrant RNAs, and ectopic DNA–DNA interactions. The causal relationship between these factors and the link between transcriptional and post-transcriptional silencing is not always clear. In this review we discuss various observations associated with gene silencing and attempt to relate them. # 1997 Annals ofBotany Company
INTRODUCTION Analysis of the large collection of genetically modiﬁed plants generated in recent years has expanded our knowledge of physiological processes and gene regulation mechanisms tremendously. However, transgenes do not always behave as expected. This has revealed the existence of hitherto unknown cellular processes. There is considerable variation in the expression oftransgenes in individual transformants which is not due to diﬀerences in copy number. Thus, gene activity is not exclusively determined by the strength of the promoter which controls transcription ; epigenetic eﬀects also inﬂuence expression levels. This sometimes leads to gene inactivation either by blocking transcription or by inhibiting mRNA accumulation. Until now, gene silencing has been aconfusing ﬁeld of research. Most of us stumbled upon this phenomenon fortuitously by analysing transgenic plants. The mechanisms by which silencing is achieved are still poorly understood. Despite the diﬀerent silencing systems being examined, several cases of gene silencing have features in common which gives us insight into the factors involved. Gene silencing also occurs in untransformed plants whereit reduces expression of endogenous genes. A number of mutations in various plant species appear to result from epigenetic gene silencing. For example, paramutation in maize (Brink, 1973) and tomato (Hageman, 1993) probably involve gene–gene interactions. Although paramutation was known long before the discovery of transgene-mediated silencing, it is only recently that the underlying molecularmechanisms have become apparent. Several features re* For correspondence.
semble those associated with transgene-mediated silencing. Because of space limitations, we will only discuss the silencing of transgenes and endogenous genes in transgenic plants. A detailed description of paramutation and related phenomena in untransformed plants can be found in other reviews (Matzke and Matzke, 1993 ;Patterson and Chandler, 1995) and some recent articles (Das and Messing, 1994 ; Hollick et al., 1995 ; Patterson et al., 1995 ; Ronchi, Petroni and Tonelli, 1995). Various aspects of transgene-mediated silencing discussed here can also be found in other reviews (Finnegan and McElroy, 1994 ; Flavell, 1994 ; Dougherty and Parks, 1995 ; Matzke and Matzke, 1995 ; Baulcombe and English, 1996 ; Meyer,...