IAA biosynthesis under pathogenic attack
Expression data of IAA biosynthetic genes revealed two major IAA biosynthetic pathways in Arabidopsis, predominated by TAA1 and CYP79B3 genes. Due to the down-regulation of TAA1 under the all selected pathogens (Figure 1), it was designated as control pathway. On the other hand, expression of CYP79B3 was up-regulated under pathogenic attack hence, this pathway was referred to as pathogenic pathways (Figure 2). B. cinerea attack induced high level of CYP71A13 transcript in Arabidopsis (Figure 1a-c), modulating IAA biosynthesis pathways to enhance camalexin production on the expense of IAA (Figure 2c). By up-regulating NIT3, E. orontii and P. syringae enhanced IAA production (Figure 1d & 2a). Moreover, IAA biosynthesis was also manipulated in P. infestans infection shifting the balance toward GL biosynthesis by up-regulating expression SUR2 genes (Figure 1b & 2b).
IAA transport under Pathogenic attack
Under pathogen attack, the host expressed and repressed differential genes. Under B. cinerea attack, the host expressed PIN2, PIN5, PIN8 genes, while repressed PIN1, PIN3, PIN4, PIN6, PIN7, AUX1, LAX2, and LAX3 genes an indication of acropetal IAA transport (Figure 3a, 4a). E. orontii induced an exactly opposite response in Arabisopsis by up-regulating expression of PIN3, PIN6, PIN1, PIN4 and LAX2, accompanied by repression of PIN2, PIN4, PIN7, PIN5, PIN8, LAX2, and AUXI which means multidirectional IAA transport (Figure 3d, 4d). P. infestans infection up-regulated PIN3 expression while repressing all other transport genes restricting IAA transport (Figure 3c, 4c). The P. syringae infected Arabidopsis seemed to possess basipetal IAA transport as indicated by up-regulated expression of PIN5, PIN8, LAX2, and PIN2 and a shutdown of the remaining genes
(Figure 3b, 4b).
Gene expression of defensive phyto-signalling molecules
Gene expression data of ACS2, ICS1 and LOX3 (signalling genes for ethylene, salicylic acid, jasmonic acid) with their correlated defense genes such as PDF1.2, PR-1 and VSP2 respectively, showed that LOX3 expression was high in B. cinerea infection as compared to ACS2 and ICS1. Therefore, the expression of VSP2 was high. Interestingly, the expression of PR-1 was also high despite of low expression of its correlated signalling gene. As opposed, E. orontii repressed LOX3 while expressed ICS1 and ACS2 gene. This response was accompanied by up-regulated expression of PR-1 and PDF1.2 and down-regulation of VSP2. P. infestans infection increased the expression of ACS2, followed by up-regulation of PDF1.2. Interestingly, leaves challenged with P. syringae had induced expression of ACS1 and VSP2 genes (Figure 5).
Co-expression of cross linking phyto-signalling molecules
Co-expression of high expressed VSP2 included genes encoding glycosyl hydrolase beta amylase, and cysteine lyase that can be involved in the biosynthesis of cellulose, starch and ethylene precursors. Similarly, arginine amidohydrolase, polygalacturonase inhibiting protein, myrosinase-binding protein and phytosulfokinase were also up-regulated. Moreover, the up-regulated genes including the defense signalling genes BGL1, BAM5, and JAR2 were also the most prominent. The down-regulated genes were those coding for cysteine synthase c1, cyclic nucleotide-gated channel 12 and phosphoinositide phosphatase protein. Genes involved in ions transport such as nitrate transport, and nodulin MtN21-like transporter family were found to be down-regulated (Figure 6a).
Co-expression with upregulated PR-1 comprised genes related to chitin catabolism, glutaredoxin production, and zinc transport. Thaumatin-like protein, vacuolar sorting receptor, glutathione s-transferase production and phosphofructokinase and ADP/ATP carriers were up-regulated for cell signalling purpose. Enzymatic processes such farnesol dehydrogenase and Pumilio (APUM) proteins were also enhanced. Down-regulated genes in high PR-1 expressing Arabidopsis were found to be coding for thylakoidal ascorbate peroxidase, uracil phosphoribosyl transferase and salicylic acid-binding proteins. Some biosynthetic processes in the infection were down-regulated such as starch synthase for normal amylopectin synthesis, lysophosphatidylcholine acyltransferase (Figure 6b).
Genes co-expressed with Up-regulated PDF1.2 included hevein-like pathogenesis-related protein, cytochrome p450 for secondary metabolites and UDP-glucosyl transferase for quercetin 3-O-glucosyltransferase activity. Cell defense proteins such as Transporter k+, glutamate-cysteine ligase, and octadecanoid-responsive Arabidopsis AP2/ERF were also up-regulated. The o-mtase family 3 protein and transcription-coupled nucleotide-excision repair genes were also co-expressed. Contrary to these genes, TAT protein binding, homeobox protein and ATP synthesis coupled electron transport proteins coding genes were not expressed in Arabidopsis with induced expression of PDF1.2. The biosynthetic processes such as cellulose synthase, RNA polymerase beta' subunit-1 and trichome birefringence protein were also down-regulated (Figure 6c).
Phyto-signalling make-up at pathogenic attack
Infection Map 1st
Infection with the biotrophic pathogen, B. cinerea modulated host physiology by inducing the expression of genes involved in biosynthesis of ABA, ET, and JA and degradation of auxins, GA, BR and SA. More notably, the CKs biosynthesis and degradation genes were simultaneously repressed. Expression of genes involved in signal perception (a first step of signalling mechanism that occurs at the cell receptor level) of GAs, CKs, ET and JA was high. Contrary to this, expression of genes responsible for auxins and SA signalling were repressed. The cell signal response (a last step of signalling mechanism, when cell produces protein) was only high for ET, JA and CKs (Figure 7a).
Infection Map 2nd
E. orontii infected leaves were characterized by enhanced expression of genes coding for GAs biosynthesis enzymes and repression of GAs degrading gene (Figure 7b). Genes involved in the biosynthesis and degradation of the rest of the phytohormones showed exact opposite trend. An exception was JA, where the biosynthesis as well as degrading genes were down-regulated. Expression of cell signal perception was very low for SA and GA, while very high for auxins, JA and ET. Interestingly, cell signalling repression to all hormones were very low, while cell signalling response was only low for auxins and SA.
Infection Map 3rd
In P. infestans exposed Arabidopsis thaliana, auxins, SA, ET, ABA, GAs and CKs biosynthesis and degradation genes were up and down regulated respectively. Interestingly, only auxins cell signalling perception genes was found to have low transcripts while those for the rest of phytohormones had high expression. where cell signalling repression of only SA and ABA was high. Moreover, cell signalling response of all were high except ABA (Figure 7c).
Infection Map 4th
Under P. syringae infection, expression of auxins biosynthesis genes was low as compared to its degradation genes. while the rest had high degradation. Genes involved in cell signal perception for all phytohormones was high except JA and ET while cell signal repression genes had high expression only for ABA and SA and high for the rest. Cell signalling response was found to be high only for GAs and CKs (Figure 7d.