Long-Term Boron-Excess-Induced Alterations of Gene Profiles in Roots of Two Citrus Species Differing in Boron-Tolerance Revealed by cDNA-AFLP

Abstract

Boron (B) toxicity is observed in some citrus orchards in China. However, limited data are available on the molecular mechanisms of citrus B-toxicity and B-tolerance. Using cDNA-AFLP, we identified 20 up- and 52 down-regulated genes, and 44 up- and 66 down-regulated genes from excess B-treated Citrus sinensis and Citrus grandis roots, respectively, thereby demonstrating that gene expression profiles were more affected in the latter. In addition, phosphorus and total soluble protein concentrations were lowered only in excess B-treated C. grandis roots. Apparently, C. sinensis had higher B-tolerance than C. grandis. Our results suggested that the following several aspects were responsible for the difference in the B-tolerance between the two citrus species including: (a) B-excess induced Root Hair Defective 3 expression in C. sinensis roots, and repressed villin4 expression in C. grandis roots; accordingly, root growth was less inhibited by B-excess in the former; (b) antioxidant systems were impaired in excess B-treated C. grandis roots, hence accelerating root senescence; (c) genes related to Ca2+ signals were inhibited (induced) by B-excess in C. grandis (C. sinensis) roots. B-excess-responsive genes related to energy (i.e., alternative oxidase and cytochrome P450), lipid (i.e., Glycerol-3-phosphate acyltransferase 9 and citrus dioxygenase), and nucleic acid (i.e., HDA19, histone 4, and ribonucleotide reductase RNR1 like protein) metabolisms also possibly accounted for the difference in the B-tolerance between the two citrus species. These data increased our understanding of the mechanisms on citrus B-toxicity and B-tolerance at transcriptional level.

Effects of High Toxic Boron Concentration on Protein Profiles in Roots of Two Citrus Species Differing in Boron-Tolerance Revealed by a 2-DE Based MS Approach

“…We obtained two up-accumulated proteasomes (G6 and 13) and two up-accumulated ubiquitin-conjugating enzymes (G48 and 54) from B-toxic C. grandis roots, but only one up-accumulated proteasome (S9) from B-toxic C. sinensis (Tables 2 , 3 ), demonstrating that B-toxicity accelerated proteolysis, especially in the former. This agrees with our data that B-toxicity only decreased total soluble protein concentration in B-toxic C. grandis roots (Guo et al, 2016 ). B-toxicity-induced increase in protein degradation implies that misfolded and damaged proteins were increased in B-toxic C. sinensis and C. grandis roots, especially in the latter.…”

“…In addition, the typical visible B-toxic symptom only occurred in B-toxic C. grandis leaves, but was not found in B-toxic C. sinensis leaves except for very few seedlings (Figure S1 ). Previous studies showed that B-toxicity only decreased the concentrations of phosphorus (P) and total soluble proteins in C. grandis roots (Guo et al, 2016 ). Based on these results, we concluded that C. sinensis had higher B-tolerance than C. grandis .…”

“…The abundances of four protein spots involved in cellular transport were increased in B-toxic C. sinensis (S20) and C. grandis (G18, 20, and 54) roots (Tables 2 , 3 ), as reported on B-toxic leaves of barley (Atik et al, 2011 ), C. sinensis and C. grandis (Sang et al, 2015 ). However, the mRNA levels of all 13 B-toxicity-responsive genes involved in cellular transport were downregulated in C. grandis and C. sinensis roots except for one upregulated H + -ATPase 4 (Guo et al, 2016 ). The difference between protein abundances and gene expression levels implies that post-translational modifications (PTMs) might affect protein levels.…”

“…By contrast, the abundances of the two kinds of proteins were enhanced in B-toxic C. grandis and C. sinensis roots (Tables 2 , 3 ). Interestingly, total soluble protein level was reduced only in B-toxic C. grandis roots and leaves (Sang et al, 2015 ; Guo et al, 2016 ). Thus, the causes for the decrease of total soluble proteins in B-toxic C. grandis roots and leaves were different.…”

“…For example, when C. sinensis and C. grandis seedlings were submitted to 400 μM B for 15 weeks, the typical B-toxic symptom only occurred in the latter (Guo et al, 2014 ; Sang et al, 2015 ). We previously investigated the differences in B-toxicity-induced alterations of gene expression profiles in roots and leaves and of protein profiles in leaves between B-tolerant C. sinensis and B-sensitive C. grandis and revealed some adaptive responses of citrus to B-toxicity (Guo et al, 2014 , 2016 ; Sang et al, 2015 ). Thus, B-toxicity-responsive proteins in roots should be different between C. sinensis and C. grandis .…”