Yiping Tong

Characterization of arsenate reductase in the extract of roots and fronds of Chinese brake fern, an arsenic hyperaccumulator.

Root extracts from the arsenic (As) hyperaccumulating Chinese brake fern (Pteris vittata) were shown to be able to reduce arsenate to arsenite. An arsenate reductase (AR) in the fern showed a reaction mechanism similar to the previously reported Acr2p, an AR from yeast (Saccharomyces cerevisiae), using glutathione as the electron donor. Substrate specificity as well as sensitivity toward inhibitors for the fern AR (phosphate as a competitive inhibitor, arsenite as a noncompetitive inhibitor) was also similar to Acr2p. Kinetic analysis showed that the fern AR had a Michaelis constant value of 2.33 mm for arsenate, 15-fold lower than the purified Acr2p. The AR-specific activity of the fern roots treated with 2 mm arsenate for 9 d was at least 7 times higher than those of roots and shoots of plant species that are known not to tolerate arsenate. A T-DNA knockout mutant of Arabidopsis (Arabidopsis thaliana) with disruption in the putative Acr2 gene had no AR activity. We could not detect AR activity in shoots of the fern. These results indicate that (1) arsenite, the previously reported main storage form of As in the fern fronds, may come mainly from the reduction of arsenate in roots; and (2) AR plays an important role in the detoxification of As in the As hyperaccumulating fern.

Mapping QTLs for nitrogen uptake in relation to the early growth of wheat (Triticum aestivum L.)

The objective of this study was to map QTLs for N uptake (NUP) in wheat, and to investigate factors influencing NUP. Two independent field trials with low N (LN) and high N (HN) treatments were conducted in the growing seasons of 2002–2003 (trial 1) and 2003–2004 (trial 2) to measure NUP per plant (N accumulated in the aerial part at maturity stage) of a doubled haploid (DH) population consisting of 120 DH lines derived from winter wheat varieties Hanxuan 10 and Lumai 14. A hydroponic culture with all nutrients supplied sufficiently was conducted to investigate shoot dry weight (SDW), root dry weight (RDW), tiller number (TN) and NUP (total plant N uptake) per plant of this mapping population at seedling stage. SDW, RDW, TN and NUP investigated in the hydroponic culture were significantly and positively correlated with each other, and with NUP under both LN and HN conditions in the field trials. Nine and eight QTLs for NUP were detected under LN and HN conditions in the field trials, respectively. Four to five QTLs for SDW, RDW, TN and NUP were detected in the hydroponic culture. One SDW QTL, three RDW QTLs, two TN QTLs detected in the hydroponic culture were linked with QTLs for NUP under LN or HN condition in the field trials. The positive correlation and genetic linkage for the traits between the field trials and the hydroponic culture demonstrated that greater seedling vigor of root and shoot is an important factor influencing N uptake in wheat.

Mapping QTLs for Phosphorus-Deficiency Tolerance at Wheat Seedling Stage

Soil phosphorus (P) deficiency is one of the major limiting factors to crop production throughout the world. It is an important strategy to breed varieties with improved P-deficiency tolerance for sustainable agriculture. The objective of this study was to map QTLs for P-deficiency tolerance in wheat, and develop molecular marker assisted selection in breeding wheat with improved P-deficiency tolerance. A doubled haploid (DH) population, consisting of 92 DH lines (DHLs) derived from P-deficiency tolerant wheat variety Lovrin 10 and P-deficiency sensitive variety Chinese Spring, was developed for mapping QTLs for P-deficiency tolerance. A genetic linkage map consisting of 34 linkage groups was constructed using 253 SSR markers. Shoot dry weight (SDW), tiller number (TN), shoot P uptake (SPU), and shoot P utilization efficiency (PUE) were investigated at seedling stage under P deficiency (−P) and sufficiency (+P) condition in two pot trials in 2002 and 2003, respectively. All traits segregated continuously in the population under either −P or +P condition. Significant positive correlations were found in between TN, SDW and SPU, whereas significant negative correlations were observed between PUE and SPU and between PUE and TN. Twenty and 19 QTLs were detected under −P and +P condition, respectively. The 39 QTLs were distributed on 21 chromosomal regions. There were three clusters of QTLs, which were associated with Xgwm25l (on chromosomes 4B), Xgwm271.2 (on chromosome 5A), and Xgwm121 (on chromosome 5D), respectively. Compared to the DHLs with all Chinese Spring alleles at the three loci, those with all Lovrin 10 alleles had, on average, much higher SPU, SDW and TN under −P condition in both trials, suggesting the importance of the three loci in controlling P-deficiency tolerance. It was interesting to find that two of the above three loci were closely linked with vernalization requirement genes VRN-A1 (on chromosome 5A) and VRN-D1 (on chromosome 5D). Potential implication of the linkage between P-deficiency tolerance and VRN genes was discussed.

Tolerance of photosynthesis to photoinhibition, high temperature and drought stress in flag leaves of wheat : A comparison between a hybridization line and its parents grown under field conditions

Photosynthesis and its tolerance to photoinhibition, high temperature and drought stress of flag leaves were investigated in a wheat (Triticum aestivum L.) hybridization line (1-12) and its parents (Jing-411 and Xiaoyan-54). From the beginning of flowering to the 10th day, light-saturated CO(2) assimilation rate (P(max)) showed no significant decrease and P(max) of 1-12 was comparable to that of its parents. From the 20th day, P(max) decreased significantly and this decrease was much less in 1-12 than in its parents, whereas no decrease in chlorophyll content was observed in 1-12 and its parents on the 20th day, indicating that photo-oxidative damage occurred in 1-12 and its parents but 1-12 is more resistant to photo-oxidative damage than its parents. To further characterize photo-oxidative damage, tolerance to photoinhibition, high temperature and drought stress was compared in 1-12 and its parents. When exposed to high light (1400μmolm(-2)s(-1)), the maximal efficiency of PSII photochemistry (F(v)/F(m)) decreased significantly with increasing exposure time and such a decrease was much less in 1-12 than in its parents. When exposed to higher temperatures (30-45°C) for 15min, F(v)/F(m) started to decrease at 42°C in 1-12 and its parents. The greatest decrease in F(v)/F(m) was observed in Jing-411. 1-12 and Xiaoyan-54 showed a comparable decrease in F(v)/F(m). Similar results were also observed in the actual PSII efficiency (Φ(PSII)), photochemical quenching (q(P)) and non-photochemical quenching (q(N)). During exposure of detached leaves to air under room temperature conditions, relative water content decreased with increasing exposure time. However, such a decrease was greatest in Jing-411 followed by Xiaoyan-54, and 1-12. There were neither changes in F(v)/F(m) nor q(N) during water loss in the line and its parents. However, there was a decrease in Φ(PSII) and q(P) and greatest decrease was observed in Jing-411 followed by Xiaoyan-54, and 1-12. Also, the decrease in P(max) was greatest in Jing-411, followed by Xiaoyan-54, and 1-12 during water loss. In addition, the activities of ribulose-1,5-bisphosphate carboxylase, phosphoenolpyruvate carboxylase, pyruvate phosphate dikinase, NADP-malate dehydrogenase and NADP-malate enzyme were significantly higher in 1-12 than in its parents. The results in this study suggest that high resistance to photo-oxidative damage of the flag leaves in 1-12 may be the physiological basis for its high yield when grown in north China. Our results also suggest that parents can be selected for improved biochemical and physiological traits and crossed to high yielding agronomically elite materials which can be selected for higher performance in yield.

A phosphate starvation response regulator Ta-PHR1 is involved in phosphate signalling and increases grain yield in wheat.

Background and Aims Phosphorus deficiency is a major limiting factor for crop yield worldwide. Previous studies revealed that PHR1 and it homologues play a key role in regulating the phosphate starvation response in plants. However, the function of PHR homologues in common wheat (Triticum aestivum) is still not fully understood. The aim of the study was to characterize the function of PHR1 genes in regulating phosphate signalling and plant growth in wheat. Methods Wheat transgenic lines over-expressing a wheat PHR1 gene were generated and evaluated under phosphorus-deficient and -sufficient conditions in hydroponic culture, a soil pot trial and two field experiments. Key Results Three PHR1 homologous genes Ta-PHR1-A1, B1 and D1 were isolated from wheat, and the function of Ta-PHR1-A1 was analysed. The results showed that Ta-PHR1-A1 transcriptionally activated the expression of Ta-PHT1.2 in yeast cells. Over-expressing Ta-PHR1-A1 in wheat upregulated a subset of phosphate starvation response genes, stimulated lateral branching and improved phosphorus uptake when the plants were grown in soil and in nutrient solution. The data from two field trials demonstrated that over-expressing Ta-PHR1-A1 increased grain yield by increasing grain number per spike. Conclusions TaPHR1 is involved in phosphate signalling in wheat, and was valuable in molecular breeding of crops, with improved phosphorus use efficiency and yield performance.

Major quantitative trait loci for seminal root morphology of wheat seedlings

Vigorous early root growth at seedling stage has been shown to be important for efficient acquisition of nutrients in wheat (Triticum aestivum L.). Identifying quantitative trait loci (QTL) for early root growth can facilitate the selection of wheat varieties with efficient nutrient use. A recombinant inbred line population derived from two Chinese wheat varieties, Xiaoyan 54 and Jing 411, was grown hydroponically at seedling stage. The maximum root length (MRL), primary root length (PRL), lateral root length (LRL), total root length (TRL), and root tip number (RN) of seminal roots were measured using the WinRHIZO Root Analyser. Numerous QTL for the investigated root traits were detected with QTL numbers varying from two to six, depending on the traits. Among them, two loci had major effects on primary (MRL and PRL) and lateral (LRL and RN) root parameters, respectively. The QTL (namely qTaLRO-B1) between Xgwm210 and Xbarc1138.2 on chromosome 2B explained 68.0 and 59.0% of phenotypic variations in MRL and PRL, respectively; the major QTL between Xgwm570 and Xgwm169.2 on chromosome 6A explained 30.5 and 24.5% of phenotypic variations in LRL and RN, respectively. These two major loci showed linkage with previous reported QTL for yield component and nutrient uptake. Detailed analysis of qTaLRO-B1 indicated that the positive allele of qTaLRO-B1 showed dominance over the negative allele, which showed impairment in primary root elongation. The existence of major QTL for root trait and their linkage with agronomic traits and nutrient uptake will facilitate the design of root morphology for better yield performance and efficient nutrient use.

Auxin biosynthetic gene TAR2 is involved in low nitrogen-mediated reprogramming of root architecture in Arabidopsis

In plants, the plasticity of root architecture in response to nitrogen availability largely determines nitrogen acquisition efficiency. One poorly understood root growth response to low nitrogen availability is an observed increase in the number and length of lateral roots (LRs). Here, we show that low nitrogen-induced Arabidopsis LR growth depends on the function of the auxin biosynthesis gene TAR2 (tryptophan aminotransferase related 2). TAR2 was expressed in the pericycle and the vasculature of the mature root zone near the root tip, and was induced under low nitrogen conditions. In wild type plants, low nitrogen stimulated auxin accumulation in the non-emerged LR primordia with more than three cell layers and LR emergence. Conversely, these low nitrogen-mediated auxin accumulation and root growth responses were impaired in the tar2-c null mutant. Overexpression of TAR2 increased LR numbers under both high and low nitrogen conditions. Our results suggested that TAR2 is required for reprogramming root architecture in response to low nitrogen conditions. This finding suggests a new strategy for improving nitrogen use efficiency through the engineering of TAR2 expression in roots.

Gene structure and expression of the high-affinity nitrate transport system in rice roots.

Rice has a preference for uptake of ammonium over nitrate and can use ammonium-N efficiently. Consequently, transporters mediating ammonium uptake have been extensively studied, but nitrate transporters have been largely ignored. Recently, some reports have shown that rice also has high capacity to acquire nitrate from growth medium, so understanding the nitrate transport system in rice roots is very important for improving N use efficiency in rice. The present study identified four putative NRT2 and two putative NAR2 genes that encode components of the high-affinity nitrate transport system (HATS) in the rice (Oryza sativa L. subsp. japonica cv. Nipponbare) genome. OsNRT2.1 and OsNRT2.2 share an identical coding region sequence, and their deduced proteins are closely related to those from mono-cotyledonous plants. The two NAR2 proteins are closely related to those from mono-cotyledonous plants as well. However, OsNRT2.3 and OsNRT2.4 are more closely related to Arabidopsis NRT2 proteins. Relative quantitative reverse transcription-polymerase chain reaction analysis showed that all of the six genes were rapidly upregulated and then downregulated in the roots of N-starved rice plants after they were re-supplied with 0.2 mM nitrate, but the response to nitrate differed among gene members. The results from phylogenetic tree, gene structure and expression analysis implied the divergent roles for the individual members of the rice NRT2 and NAR2 families. High-affinity nitrate influx rates associated with nitrate induction in rice roots were investigated and were found to be regulated by external pH. Compared with the nitrate influx rates at pH 6.5, alkaline pH (pH 8.0) inhibited nitrate influx, and acidic pH (pH 5.0) enhanced the nitrate influx in 1 h nitrate induced roots, but did not significantly affect that in 4 to 8 h nitrate induced roots.

Restricted spatial expression of a high-affinity phosphate transporter in potato roots

Phosphorus deficiency limits plant growth, and high-affinity phosphate transporters, of the Pht1 family, facilitate phosphate uptake and translocation. The family is subdivided into root specific, phosphate deprivation induced members and those also expressed in leaves. An antibody to StPT2, a potato root specific transporter, detected two bands (52 kDa and 30 kDa) on western blots of root plasma membrane extracts that were most intense in whole extracts from the root tip and slightly increased throughout the root in response to phosphate depletion. RT-PCR, using StPT2 specific primers, confirmed these findings. Low power confocal immunofluorescent images showed StPT2 expression mainly in the elongation zone at the root tip. By contrast, a vacuolar pyrophosphatase and a plasma membrane ATPase antibody labelled the whole root. High power images showed, by comparison with α-tubulin, cell wall and plasma membrane ATPase labelling, that StPT2 was in the epidermal plasma membrane and restricted to the apical surface. This is the first evidence of polar plasma membrane localisation of a plant nutrient transporter and is consistent with a role for StPT2 in phosphate capture and uptake.

Bioengineering and management for efficient phosphorus utilization in crops and pastures

Phosphorus (P) is one of the three macronutrients for plants. Because of its low mobility and high fixation in soils, low P availability is a worldwide constraint for crop productivity. Molecular biology provides great opportunities to improve P efficiency in plants. However, transgenic plants cannot be commercialized before integrating all the knowledge on bottlenecks for improving P efficiency of crops/pastures. This review intends to summarize the main strategies of bioengineering to improve P efficiency of crops/pastures, including conventional and molecular assisted breeding, identification and application of key genes for biotech plants. It highlights recent advances in the understanding of improving P efficiency through the integration of bioengineering with P fertilization and cultivation management.