Time-series expression profiling of sugarcane leaves infected withPuccinia kuehniireveals an ineffective defense system leading to susceptibility

Abstract

Puccinia kuehnii is an obligate biotrophic fungus that infects sugarcane leaves causing a disease called orange rust. It spread out to other countries resulting in reduction of crop yield since its first outbreak in Australia. One of the knowledge gaps of that pathosystem is to understand the molecular mechanisms altered in susceptible plants by the stress induced by P. kuehnii. Here we investigated changes in temporal expression of transcripts in pathways associated with the immune system. To achieve this purpose, we used RNA-Seq to analyze infected leaf samples collected at 0, 12, 24, 48 hours after inoculation (hai), 5 and 12 days after inoculation (dai). A de novo transcriptome was assembled with preprocessed reads, filtering out potential fungal sequences to focus on plant transcripts. Differential expression analyses of adjacent time points revealed substantial changes at 12, 48 hai and 12 dai, coinciding with the events of spore germination, haustoria post-penetration and post-sporulation. During the first 24 hours, a lack of transcripts involved with resistance mechanisms was revealed by underrepresentation of hypersensitive and defense responses. However, two days after inoculation, upregulation of genes involved with immune response regulation provided evidence of some potential defense response. Events related to biotic stress and phenylpropanoid biosynthesis pathways were predominantly downregulated in the initial time points, but expression was later restored to basal levels. Similar waves of expression were apparent for sets of genes in photosynthesis and oxidative processes, whose initial repression could avoid production of signaling molecules. Their subsequent upregulation possibly restored carbohydrate metabolism, ensuring pathogen nutritional requirements were met. Our results support the hypothesis that P. kuehnii initially suppressed sugarcane genes involved in plant defense systems. Late overexpression of specific regulatory pathways also suggests the possibility of an inefficient recognition system by a susceptible sugarcane genotype. March 15, 2019 1/27 1 Sugarcane is currently cultivated in 27 million hectares worldwide, 52% of which are 2 located in America, more expressively in South America (11 million ha). Brazil is the 3 main producer, responsible for 768.68 million tonnes (FAOSTAT, 2016). The term 4 sugarcane includes at least six species of the genus Saccharum [1, 2]. The ancestor 5 species are Saccharum robustum and S. spontaneum, which present variable ploidy 6 levels with chromosome number ranges of 2n = 60 -170 and 2n = 36 -128, respectively 7 [3, 4]. The octoploid (2n = 80) S. officinarum is an ancient cultivated sugarcane with 8 high sugar content that formed the group of noble cultivars [3, 5, 6]. Sugarcane plants 9 currently used for industrial purposes are modern cultivars derived from interspecific 10 crosses, mainly between S. officinarum and S. spontaneum. As a consequence of 11 successive backcrosses, the majority of chromosomes in homology groups are from S. 12 offici...