The molecular basis of shoot responses of maize seedlings to Trichoderma harzianum T22 inoculation of the root: a proteomic approach

Abstract

Trichoderma spp. are effective biocontrol agents for several soil-borne plant pathogens, and some are also known for their abilities to enhance systemic resistance to plant diseases and overall plant growth. Root colonization with Trichoderma harzianum Rifai strain 22 (T22) induces large changes in the proteome of shoots of maize (Zea mays) seedlings, even though T22 is present only on roots. We chose a proteomic approach to analyze those changes and identify pathways and genes that are involved in these processes. We used two-dimensional gel electrophoresis to identify proteins that are differentially expressed in response to colonization of maize plants with T22. Up- or down-regulated spots were subjected to tryptic digestion followed by identification using matrix-assisted laser desorption/ionization tandem time-of-flight mass spectrometry and nanospray ion-trap tandem mass spectrometry. We identified 91 out of 114 up-regulated and 30 out of 50 down-regulated proteins in the shoots. Classification of these revealed that a large portion of the up-regulated proteins are involved in carbohydrate metabolism and some were photosynthesis or stress related. Increased photosynthesis should have resulted in increased starch accumulation in seedlings and did indeed occur. In addition, numerous proteins induced in response to Trichoderma were those involved in stress and defense responses. Other processes that were up-regulated were amino acid metabolism, cell wall metabolism, and genetic information processing. Conversely, while the proteins involved in the pathways noted above were generally up-regulated, proteins involved in other processes such as secondary metabolism and protein biosynthesis were generally not affected. Up-regulation of carbohydrate metabolism and resistance responses may correspond to the enhanced growth response and induced resistance, respectively, conferred by the Trichoderma inoculation.

Figure 1.

Functional categories of identified proteins. Identified proteins were categorized into functional groups. Proteins involved in more than one process were assigned to more than one categorical group. The number of proteins in each categorical group is presented here. Up-regulated proteins are in hatched bars and down-regulated proteins are in stippled bars.

Figure 2.

Validation of selected genes. Semiquantitative RT-PCR analysis was performed for selected genes using RNA of shoots from control (C) and Trichoderma-treated (T) plants. PCR was conducted for 20 cycles for all genes. 18S was used as a reference gene and 18 cycles were performed on a 10-fold dilution of the RT reaction.