Activity of XTHs during host plant infection by the parasitic plant Cuscuta.

Abstract

Parasitic plants including Cuscuta develop specialized structures called haustoria during infection. These specialized organs give the parasite the capacity of host attachment, invasion, vasculature connection and material transfer between the host and the parasite. Successful invasion allows organic substances, nutrients and water to flow into the parasite through the host phloem and xylem cells. Invading the host plant requires a number of cell wall modifications and recent findings suggests that, a number of enzymes and cell wall components control the modification process in the formation of haustoria. The cell wall-modifying enzymes xyloglucan endotransglucosylases/hydrolases (XTHs) have been related to Cuscuta reflexa haustoria formation. In the presented thesis, the infection mechanism of other Cuscuta species was examined by investigating the expression and activity of XTHs during host infection. Also, the effect of an XTH enzyme inhibitor on host plant infection was tested. Cuscuta campestris and Cuscuta platyloba were grown on the host plant Pelargonium zonale to quantify the expression of XTHs as well as the XET activities in the haustoria and stems of the parasite. RNA extraction and qPCR analysis were used for quantifying the gene expression of Cr-XTH homologues. An ELISA assay was used to analyze the cell wall component in the Cuscuta species. Furthermore, an inhibition trial on Cuscuta species and the host plant was conducted using Coomassie blue BB-R250 at 5mM concentration. Results showed that the expression of Cp-XTH1 was well regulated in the haustoria of C. platyloba, but not in C. campestris, while the expression of Cc-XTH2 was greatly regulated in C. campestris but not in C. platyloba. XET activity was generally higher in C. campestris than C. platyloba but possessed higher levels of XET in their haustoria and stems than that of C. platyloba per mg tissue sample. Furthermore, the ELISA assay showed that xyloglucan content was high in both the haustoria and the stem. The higher xyloglucan levels in the haustoria section of C. platyloba coordinated with the high expression of XTHs and activity of XET per mg of the tissue samples but vice versa with the activity levels of XET per mg of the protein concentration. The high expression of XTHs and XET was correlated to high xyloglucan levels in the haustoria. The infection trial of C. platyloba infecting P. zonale coated with Coomassie blue BB-R250 revealed that the preventive infection by 5 Mm concentration was 6 times higher than in the absence of dye (0 mM). In conclusion, the expression of XET in Cuscuta species is not dependent on the different groups of XTH. Therefore, once XTH is expressed, then the activity and levels of XET increases irrespective of the groups within the various species. Some preventive penetration was recorded although it was not of high percentage (15.8%) during the inhibition infection trial of Coomassie blue BB-R250. Therefore, the high levels of XTH, XET and xyloglucan work synergistically to modify the parasitic plant leading to the formation of the specialized structures called haustoria.