Silencing of waxy genes in rice containing Wx transgenes

In order to study gene silencing in a monocot system we introduced a waxy (Wx) gene into rice. In the pollen grain of the transgenic wild-type plants, two types of Wx gene silencing were observed: Type I in which all the pollen grains showed the mutant (wx) phenotype, and Type II in which 50% of the pollen grains showed the wx phenotype. Analysis of Wx gene expression in the progeny of selfing and outcrossing indicated that Wx gene silencing was meiotically transmitted to the offspring. The number of transgene copies and transgene loci was determined by Southern blot analysis and suggested that the Wx transgene may have a paramutagenic effect on the endogenous Wx genes. In contrast to the pollen grain, the wx phenotype was not observed in the endosperm. However, the level of WAXY (WX) protein in the endosperm of Type I lines was similar to that in non-transgenic seed, while in Type II lines two classes of seeds, showing high and low levels of the protein segregated. When the same transgene was introduced into wx mutants in which no Wx transcript was detectable, the transgene behaved as a dominant Mendelian factor and no silencing was found, suggesting that the activity of the endogenous Wx gene influences the silencing phenomenon. Our study of Wx gene silencing in rice extends the well-known phenomenon of gene silencing, so far observed mainly in dicots, to a cereal.