PlantMolecular Biology 31:713-719, 1996. @ 1996 Kluwer Academic"Publishers. Printed in Belgium. 713 Stringently and developmentally regulated levels of a cytoplasmic double-stranded RNA and its high-efficiency transmission via egg and pollen in rice H. Moriyama, K. Kanaya 1, J.Z. Wang 2, T. Nitta* and T. Fukuhara Laborato~ of Molecular Cell Biology, Faculty of Agriculture, To~'o University of Agriculture and Technology; Fuchu, Tokyo 183, Japan (*author for correspondence); J Present address: Laborato©' of Glico Molecular Biology, Frontier Research Program, The Institute of Physical and Chemical Research, Wako, Saitama 351-01, Japan; 20n leave from Institute of Genetics, Academia Sinica, Bei/ing, PCR Received24 October 1995;acceptedin revisedform8 April 1996 Key words: biparental transmission, copy number, cultured cell, cytoplasmic inheritance, double-stranded RNA, Orvza sativa Abstract A very restricted amount of high-molecular-weight double-stranded RNA (dsRNA) has been tbund in healthy japonica rice plants. We discriminated dsRNA-carrying rice plants from noncarriers. The endogenous dsRNA was localized in the cytoplasm (about 100 copies per cell) and was transmissible to progeny plants by mating. In crosses between carriers and noncarriers, the RNA was transmitted efficiently to F1 plants via both egg and pollen. The rice dsRNA was maintained at an almost constant level by host plant cells from generation to generation. The high-efficiency transmission of the endogenous dsRNA to progeny plants appears to depend on the autonomously controlled replication of the dsRNA localized in cytoplasmic vesicles. However, an increase in copy number (about 10-fold) of the dsRNA was observed during the suspension culture of host cells. The number of copies of dsRNA returned to the original low value in regenerated plants, suggesting that the copy number is stringently and developmentally regulated in rice cells. Introduction Endogenous double-stranded RNAs (dsRNAs) are dis- tributed extensively in the plant kingdom, from algae to higher plants [2, 5, 6, 10, 15]. Host plants with these plasmid-like dsRNAs are apparently asymptomatic. Although horizontal transmission of the dsRNAs from plant to plant has not been proven, the dsRNAs are known to be transmitted to progeny cells during cell division [2]. Thus, the known endogenous dsRNAs are transmit- ted vertically from generation to generation via seeds [1]. The uniparental, namely, maternal, inheritance of plant dsRNAs via seeds could be simply anticip- ated when the dsRNAs are localized in the cytoplasm [2]. However, the transmittance efficiency of the cyto- plasmic dsRNAs during fertilization is distinct from the efficiency of the cell organelles. Some of low- molecular-weight dsRNAs, encapsidated by proteins to form isometric particles, known as cryptoviruses, are transmitted biparentally to progeny plants; the trans- mission is generally found to be about 50% efficient when noncarrier plants are pollinated by carriers [1]. Double-stranded RNAs that do not appear to be associ- ated with distinct virus-like particles in the cytoplasm are also transmitted biparentally in Phaseolus [14], alfalfa [7], Brassica [16] and barley [24]. Thus, details of the inheritance of the cytoplasmic dsRNAs should be examined extensively in a variety of plants with special reference of copy number and the subcellular localization of the dsRNAs in host cells. A high-molecular-weight dsRNA occurs in every tissue and at every developmental stage in japonica rice [8, 22, 23]. The rice of dsRNA of 13 952 nucleotides (nt) in length has one, extremely long, open reading frame (ORF) of 13 716 nt that appears to encode a large