A comparison between constitutive and inducible transgenic expression of the PhRIP I gene for broad-spectrum resistance against phytopathogens in potato.

OBJECTIVES: To engineer broad spectrum resistance in potato using different expression strategies. RESULTS: The previously identified Ribosome-Inactivating Protein from Phytolacca heterotepala was expressed in potato under a constitutive or a wound-inducible promoter. Leaves and tubers of the plants constitutively expressing the transgene were resistant to Botrytis cinerea and Rhizoctonia solani, respectively. The wound-inducible promoter was useful in driving the expression upon wounding and fungal damage, and conferred resistance to B. cinerea. The observed differences between the expression strategies are discussed considering the benefits and features offered by the two systems. CONCLUSIONS: Evidence is provided of the possible impact of promoter sequences to engineer BSR in plants, highlighting that the selection of a suitable expression strategy has to balance specific needs and target species.

Draft genome sequence of bitter gourd (Momordica charantia), a vegetable and medicinal plant in tropical and subtropical regions.

Bitter gourd (Momordica charantia) is an important vegetable and medicinal plant in tropical and subtropical regions globally. In this study, the draft genome sequence of a monoecious bitter gourd inbred line, OHB3-1, was analyzed. Through Illumina sequencing and de novo assembly, scaffolds of 285.5 Mb in length were generated, corresponding to ∼84% of the estimated genome size of bitter gourd (339 Mb). In this draft genome sequence, 45,859 protein-coding gene loci were identified, and transposable elements accounted for 15.3% of the whole genome. According to synteny mapping and phylogenetic analysis of conserved genes, bitter gourd was more related to watermelon (Citrullus lanatus) than to cucumber (Cucumis sativus) or melon (C. melo). Using RAD-seq analysis, 1507 marker loci were genotyped in an F2 progeny of two bitter gourd lines, resulting in an improved linkage map, comprising 11 linkage groups. By anchoring RAD tag markers, 255 scaffolds were assigned to the linkage map. Comparative analysis of genome sequences and predicted genes determined that putative trypsin-inhibitor and ribosome-inactivating genes were distinctive in the bitter gourd genome. These genes could characterize the bitter gourd as a medicinal plant.

BDP-30, a systemic resistance inducer from Boerhaavia diffusa L., suppresses TMV infection, and displays homology with ribosome-inactivating proteins.

Root extract of Boerhaavia diffusa L. induced systemic resistance in tobacco against Tobacco mosaic virus. A 30 kDa protein was isolated as the active component, called BDP-30 on the basis of the molecular weight and source plant. BDP-30, a glycoprotein, was found to be temperature and protease resistant. It was basic, possessing a pI greater than 9.0. In-gel proteolytic digestion of BDP-30 generated two peptides that possessed the amino acid sequence KLYDIPPLR and KVTLPYSGNYER by LC/MS/MS. Both peptides shared absolute sequence identity with trichosanthin, a ribosome-inactivating protein from Trichosanthes kirilowii, and a 78 percent and 100 percent homology respectively with an RIP from Bryonia dioica, bryodin. Further, effort was made to look at the fate of TMV in induced resistant Nicotiana tabacum cv. Xanthi, a systemic host of the virus, at specified days after inoculation in control and treated plants. TMV coat protein (CP) was detected by immunoblot 7 days post inoculation up to 21 days in the control set, but not in treated resistant plants. TMV RNA was detected by RT-PCR using TMV-CP specific primers. Resistant tobacco did not show presence of TMV RNA up to 21 days of inoculation. This suggests that BDP-30 may be suppressing TMV replication.

Elicitor-dependent expression of the ribosome-inactivating protein beetin is developmentally regulated.

BE27 and BE29 are two forms of beetin, a virus-inducible type 1 ribosome-inactivating protein isolated from leaves of Beta vulgaris L. Western blot analysis revealed the presence of beetin forms in adult plants but not in germ or young plants, indicating that the expression of these proteins is developmentally regulated. While beetins are expressed only in adult plants, their transcripts are present through all stages of development. In addition, the treatment of B. vulgaris leaves with mediators of plant-acquired resistance such as salicylic acid and hydrogen peroxide promoted the expression of beetin by induction of its transcript, but only in adult plants. The plant expresses three mRNAs which differ only in their 3' untranslated region. All these observations suggest a dual regulation of beetin expression, i.e. at the post-transcriptional and transcriptional levels. Additionally, total RNA isolated from leaves treated with hydrogen peroxide, which express high levels of active beetin, is not de-adenylated by endogenous beetin, nor in vitro by the addition of BE27, thus suggesting that sugar beet ribosomes are resistant to beetin.