William R. Hiatt

The Application of Antisense RNA Technology to Plants

Antisense RNA consists of sequences complementary to a target RNA and can act as a regulatory molecule by binding to the target species via base pairing. Gene expression can be inhibited in this manner; such regulation occurs naturally in bacteria and has been demonstrated experimentally in eukaryotes (for reviews see 1–6). Until recently, work in eukaryotes has utilized organisms such as Drosophila (7), Dictyostelium (8), Xenopus (9), and mammals, including cell lines of various species (10 and transgenic mice (11). Despite this diversity in experimental organisms and approaches, few generalizations can be made regarding the mechanisms of inhibition. However, in studies which involve the inhibition of an endogenous gene by the introduction of a stably integrated antisense construct, a substantial reduction in the amount of target mRNA has been frequently observed. Examples include hsp26 mRNA in Drosophila cells (12), discoidin 1 (13) and myosin mRNA (8) in Dictyostelium, MYC mRNA in mammalian cells (10), and myelin basic protein mRNA in mice (11). Although the mechanism(s) by which antisense RNA reduces mRNA is unclear, multiple steps in mRNA biogenesis have been implicated including transcription (10) and transport from the nucleus (13, 14). RNA:RNA duplex formation in the nucleus has been proposed to reduce the stability of mRNA (13). A substantial excess of antisense RNA has often been required for an effective reduction of target mRNA levels (12–15).