Transgenic tobacco plants (Nicotiana tabacum cv. SR1) expressing extracellular pancreatic ribonuclease from Bos taurus and characterized by an increased level of ribonuclease activity in leaf extracts were challenged with tobacco mosaic virus. The transgenic plants exhibited a significantly higher level of protection against the virus infection than the control non-transformed plants. The protection was evidenced by the absence (or significant delay) of the appearance of typical mosaic symptoms and the retarded accumulation of infectious virus and viral antigen. These results demonstrate that modulation of extracellular nuclease expression can be efficiently used in promoting protection against viral diseases.

No abstract

The gene of Zinnia elegans L. coding for S-like extracellular ribonuclease (ZRNase II) was used to produce transgenic tobacco plants with an increased ribonuclease activity. The protein-coding part of ZRNase II included the signal peptide sequence so the transgenic protein was located extracellularly. The cDNA of ZRNase II was cloned under the control of 2'-promoter of the mannopine synthase (MAS 2') gene from Ti-plasmid of Agrobacterium tumefaciens. It was shown that the resultant transgenic plants had an increased ribonuclease activity of the crude extracts and the induction of MAS 2' promoter by wounding additionally increased the activity. The plants of two transforming lines characterized by different ribonuclease activities were used to analyze the transgene influence on plant resistance to tobacco mosaic virus. The plants demonstrated either absence of disease symptoms or a significant delay in their appearance, depending on the virus content in the inoculum and ribonuclease activity.

The isolates of potato virus X causing disease symptoms of different severity in thorn-apple plants (Datura stramonium L.) can be categorized in terms of the increase in antigen accumulation, as follows: strong, medium and weak. Extracts from leaves of healthy plants and of those infected with separate virus isolate did not differ in set of acidic pathogenesis-related (PR) proteins as shown by discontinuous non-denaturating polyacrylamide gel electrophoresis. Within 2 weeks after inoculation the total content of PR-proteins in leaves of plants infected with weak or medium isolates was higher than in the healthy ones but after 3 weeks these differences became negligible. The plants infected with the strong isolate had higher total content of PR-proteins than healthy plants and plants infected with weak or medium isolates during 3 weeks. It is proposed that the greater the damage to the plant caused by virus infection, the more PR-proteins are accumulated.

Extracellular Serratia marcescens nuclease is an extremely active enzyme which non specifically degrades RNA and DNA. Its antiviral activity was previously shown both in animals and in plants when applied exogenously. Transgenic tobacco plants (Nicotiana tabacum L. cv. SR1) expressing S. marcescens chi meric, mutant, and intracellular mutant nuclease gene variants were regenerated and challenged with tobacco mosaic virus. The transgenic plants exhibited a higher level of resistance to the virus infection than the control non transgenic plants. The resistance was evidenced by the delay of the appearance of mosaic symptoms and the retarded accumulation of viral antigen. Thus, these results reveal that modulations of both extracellular nuclease activity and intracellular RNA/DNA binding can protect plants against viral diseases.