Suppressive Subtractive Hybridization Approach Revealed Differential Expression of Hypersensitive Response and Reactive Oxygen Species Production Genes in Tea (Camellia sinensis (L.) O. Kuntze) Leaves during Pestalotiopsis thea Infection

Abstract

Tea (Camellia sinensis (L.) O. Kuntze) is an economically important plant cultivated for its leaves. Infection of Pestalotiopsis theae in leaves causes gray blight disease and enormous loss to the tea industry. We used suppressive subtractive hybridization (SSH) technique to unravel the differential gene expression pattern during gray blight disease development in tea. Complementary DNA from P. theae-infected and uninfected leaves of disease tolerant cultivar UPASI-10 was used as tester and driver populations respectively. Subtraction efficiency was confirmed by comparing abundance of β-actin gene. A total of 377 and 720 clones with insert size >250 bp from forward and reverse library respectively were sequenced and analyzed. Basic Local Alignment Search Tool analysis revealed 17 sequences in forward SSH library have high degree of similarity with disease and hypersensitive response related genes and 20 sequences with hypothetical proteins while in reverse SSH library, 23 sequences have high degree of similarity with disease and stress response-related genes and 15 sequences with hypothetical proteins. Functional analysis indicated unknown (61 and 59 %) or hypothetical functions (23 and 18 %) for most of the differentially regulated genes in forward and reverse SSH library, respectively, while others have important role in different cellular activities. Majority of the upregulated genes are related to hypersensitive response and reactive oxygen species production. Based on these expressed sequence tag data, putative role of differentially expressed genes were discussed in relation to disease. We also demonstrated the efficiency of SSH as a tool in enriching gray blight disease related up- and downregulated genes in tea. The present study revealed that many genes related to disease resistance were suppressed during P. theae infection and enhancing these genes by the application of inducers may impart better disease tolerance to the plants.

Transcriptional analysis and histochemistry reveal that hypersensitive cell death and H2O2 have crucial roles in the resistance of tea plant (Camellia sinensis (L.) O. Kuntze) to anthracnose

“…As a signalling molecule, ROS can regulate PCD in plants during pathogen infection and can mutually regulate MAPK signalling 39 , 46 , 47 . Senthilkumar et al 14 reported that the HR and ROS bursts are involved in tea plant defence against P. theae . In our study, we observed that DEGs were enriched in the H 2 O 2 catabolic process (Fig.…”

“…The resistance mechanism of tea plants has been loosely explained based on the results of several studies. For example, by studying tea plant defence to grey blight disease caused by Pestalotiopsis species, Senthilkumar et al 14 , who used suppressive subtractive hybridisation techniques, suggested that the HR and ROS play crucial roles in tea plant resistance to P. theae . Moreover, Palanisamy and Mandal 15 reported that the antioxidative enzymes associated with ROS in resistant tea cultivars have higher activity than those in susceptible cultivars following Pestalotiopsis sp.…”

Biochemical and molecular studies on the resistance mechanisms in tea [Camellia sinensis (L.) O. Kuntze] against blister blight disease

“…Tea [Camellia sinensis (L.) O. Kuntze] is a perennial crop, with an economic life span of over 60 years. Extensive cultivation, improved technology, nutrition and fertility management, introduction of high yielding clones and longer pruning cycle have helped in production increase but have also encouraged productivity limit due to biotic stresses caused by insects, mite pests and various other pathogens (Senthilkumar et al 2012;Singh et al 2015).…”

Metabolism of Gallic Acid and Its Distributions in Tea (Camellia sinensis) Plants at the Tissue and Subcellular Levels

“…Host plants may have evolved multiple defense mechanisms in their interactions with various pathogens. Many studies have attempted to elucidate the molecular mechanisms of tea plant resistance against pathogens [ 33 , 34 ]. Additionally, plants can produce many chemicals, known as secondary metabolites, to protect themselves.…”