Analysis of external factors that affect formation and accumulation of D-RNAs in Bromovirus infections

Abstract

Defective RNA molecules (D-RNAs) are being studied in several plant RNA virus groups. In the genus Bromovirus, D-RNAs have been described for Broad bean mottle virus (BBMV), they are formed by a single internal deletion in the RNA2 and have been generated de novo by serial passages. In contrast, in Brome mosaic virus (BMV) D-RNAs are generated by a single or double internal deletion in the RNA3 without serial passages in their hosts. In this work the external effects of the host and the growth temperature in the generation and accumulation of D-RNAs in BBMV, BMV and Cowpea chlorotic mottle virus (CCMV) are studied. The BBMV and BMV D-RNAs were generated and accumulated with or without serial passages of the viruses in different hosts and cultivars. Plants grown at 12, 16, 20 and 24°C in a growth chamber and in a greenhouse (22 ± 5°C) generated D-RNAs after being inoculated with virusfree D-RNAs, with and without passages. The D-RNAs observed in BBMV and BMV presented some common characteristics: both were formed de novo after serial passages or without passages, their deletion borders had short repeated and palindrome sequences that favour recombination. In addition, growing inoculated plants at lower temperatures greatly facilitated the generation and accumulation of D-RNAs. The CCMV did not generate defective molecules in cowpea (Vigna unguiculata) and Nicotiana benthamiana plants after several serial passages or without passages. This is the first time that D-RNAs have been generated in BBMV without passages and BMV with serial passages.Additional key words: BBMV, BMV, CCMV, host effects, temperature conditions. Resumen Análisis de los factores externos que afectan a la formación y acumulación de los D-RNAs en infecciones de BromovirusLos RNAs defectivos (D-RNAs) están siendo estudiados en diversos virus RNA de plantas. En el género Bromovirus se han descrito D-RNAs en el virus del moteado del haba (BBMV), formados mediante una delección interna en el RNA2, que se generan de novo mediante pases seriados del virus sobre un nuevo huésped. En contraste, en el virus del mosaico del bromo (BMV), los D-RNAs se forman por una o dos delecciones en el RNA3 sin necesidad de pases en sus huéspedes. En este trabajo estudiamos algunos factores externos, como los huéspedes, las inoculaciones y las temperaturas de crecimiento en la generación y acumulación de D-RNAs en BBMV, BMV y en el virus del moteado clorótico de la carilla (CCMV). Los D-RNAs de BBMV y BMV fueron generados y acumulados mediante pases seriados y sin pases de los virus en diferentes huéspedes y cultivares. Plantas inoculadas con BBMV y BMV sin D-RNAs, mantenidas a 12, 16, 20 y 24°C en cámaras de crecimiento a temperatura constante o en un invernadero (22 ± 5°C), generaron D-RNAs con o sin pases sobre nuevos huéspedes. Los D-RNAs observados en ambos virus presentan algunas características en común: son formados de novo mediante pases seriados o purificados de hojas infectadas algunas semanas después de la inoculación; los bordes de la delección tie...

Replication of Plant Viruses

“…The DI RNAs retained the 5′-terminal 1152 nucleotides and the 3′-terminal 468 nucleotides; (ii) the overall size. No D RNAs were found in CCMV (Sandoval et al, 2008). The DI RNA retains a deleted form of the 2a ORF that comprises at least 79% of the molecule.…”

“…To complicate the situation, a DI RNA may also be the target of RNA silencing if it produces enough dsRNA target. The DI RNAs have various sized deletions in the 2a ORF and include the GDD motif characteristic of the function of this ORF product as an RdRp (Romero et al, 1993;Pogany et al, 1995;Sandoval et al, 2008) (Figure 7.17). Graves et al, 1996) The DI RNA of BBMV is derived from, and decreases the concentration of, RNA2.…”