Comparative proteomics analysis reveals an intimate protein network provoked by hydrogen peroxide stress in rice seedling leaves.

Hydrogen peroxide (H2O2) plays a dual role in plants as the toxic by-product of normal cell metabolism and as a regulatory molecule in stress perception and signal transduction. However, a clear inventory as to how this dual function is regulated in plants is far from complete. In particular, how plants maintain survival under oxidative stress via adjustments of the intercellular metabolic network and antioxidative system is largely unknown. To investigate the responses of rice seedlings to H2O2 stress, changes in protein expression were analyzed using a comparative proteomics approach. Treatments with different concentrations of H2O2 for 6 h on 12-day-old rice seedlings resulted in several stressful phenotypes such as rolling leaves, decreased photosynthetic and photorespiratory rates, and elevated H2O2 accumulation. Analysis of approximately 2000 protein spots on each two-dimensional electrophoresis gel revealed 144 differentially expressed proteins. Of them, 65 protein spots were up-regulated, and 79 were down-regulated under at least one of the H2O2 treatment concentrations. Furthermore 129 differentially expressed protein spots were identified by mass spectrometry to match 89 diverse protein species. These identified proteins are involved in different cellular responses and metabolic processes with obvious functional tendencies toward cell defense, redox homeostasis, signal transduction, protein synthesis and degradation, photosynthesis and photorespiration, and carbohydrate/energy metabolism, indicating a good correlation between oxidative stress-responsive proteins and leaf physiological changes. The abundance changes of these proteins, together with their putative functions and participation in physiological reactions, produce an oxidative stress-responsive network at the protein level in H2O2-treated rice seedling leaves. Such a protein network allows us to further understand the possible management strategy of cellular activities occurring in the H2O2-treated rice seedling leaves and provides new insights into oxidative stress responses in plants.

Histidine kinase MHZ1/OsHK1 interacts with ethylene receptors to regulate root growth in rice.

Ethylene plays essential roles during adaptive responses to water-saturating environments in rice, but knowledge of its signaling mechanism remains limited. Here, through an analysis of a rice ethylene-response mutant mhz1, we show that MHZ1 positively modulates root ethylene responses. MHZ1 encodes the rice histidine kinase OsHK1. MHZ1/OsHK1 is autophosphorylated at a conserved histidine residue and can transfer the phosphoryl signal to the response regulator OsRR21 via the phosphotransfer proteins OsAHP1/2. This phosphorelay pathway is required for root ethylene responses. Ethylene receptor OsERS2, via its GAF domain, physically interacts with MHZ1/OsHK1 and inhibits its kinase activity. Genetic analyses suggest that MHZ1/OsHK1 acts at the level of ethylene perception and works together with the OsEIN2-mediated pathway to regulate root growth. Our results suggest that MHZ1/OsHK1 mediates the ethylene response partially independently of OsEIN2, and is directly inhibited by ethylene receptors, thus revealing mechanistic details of ethylene signaling for root growth regulation.

[Physiological function of the qPGWC-9 related to high percentage of rice grains with Chalkiness].

Percentage of grains with chalkiness (PGWC), one of the important traits assessing rice grain appearance quality, belonged to qualitative trait controlled by many genes. Our previous study identified a novel quantitative trait locus (QTL), namely qPGWC-9, related to high PGWC using chromosomal segment substitution line (CSSL) population. qPGWC-9 was shown to be expressed stably in eight environments. AIS82 which carried a IR24 chromosomal segment corresponding to qPGWC-9 in the Asominori genetic background was selected and analyzed to clarify the physiological function of qPGWC-9 from the relationship of source and sink of carbohydrates. It showed that AIS82 had higher PGWC than Asominori (control variety with low PGWC). The net photosynthetic rate of flag leaf of AIS82 showed no significant difference from that of Asominori, so photosynthetic ability in flag leaf was not directly related with high PGWC in AIS82. But, the changes in pattern of activity of the key enzymes associated with starch synthesis were different in these plants. Activities of some key enzymes in starch synthesis in AIS82 changed more radically than those in Asominori. These results suggest that qPGWC-9 might determine the activities of some enzymes associated with starch synthesis and therefore affect the degree of grain chalkiness.

[Stable expression of QTL for grain shape of milled rice (Oryza sativa L. ) using a CSSLs population].

A set of chromosome segment substitution lines (CSSLs), derived from Asominori/IR24 with Asominori as the recurrent parent,was planted and phenotyped for grain length (GL), grain width (GW) and length-width ratio (LWR) of milled rice in two successive years and four sites. QTL for GL,GW and LWR were characterized and stability of their expression was investigated. The phenotypic values for each trait showed a continuous distribution and some transgressive lines were also observed in the CSSLs population. Additionally, a total of 13 QTL for GL, GW and LWR were identified,and six of them were consistently detected in the eight different environments. Phenotypic values were different significantly (P < 0. 001) between the CSSLs harboring any of the six QTL alleles and the genetic background parent, Asominori. Significant phenotypic correlations (r > or = 0.75, r0.05 = 0.67) were detected among different environments for these CSSLs carrying the same target QTL. Also, the results indicated that the six QTL, namely, qGL-3 for GL, qGW-5a and qGW-5b for GW, qLWR-3, qLWR-5a and qLWR-5b for LWR, were stably expressed in different environments. Since QTL qGL-3 and qLWR-3 were mapped in the R19-C1677 interval, qGW-5a and qLWR-5a in the vicinity of RFLP marker C263, qGW-5b and qLWR-5b near R569,the four RFLP markers, R19, C1677, C263 and R569,would be useful for further marker-assisted selection (MAS) in rice quality improvement.

Coordinated transcriptional regulation underlying the circadian clock in Arabidopsis.

The circadian clock controls many metabolic, developmental and physiological processes in a time-of-day-specific manner in both plants and animals. The photoreceptors involved in the perception of light and entrainment of the circadian clock have been well characterized in plants. However, how light signals are transduced from the photoreceptors to the central circadian oscillator, and how the rhythmic expression pattern of a clock gene is generated and maintained by diurnal light signals remain unclear. Here, we show that in Arabidopsis thaliana, FHY3, FAR1 and HY5, three positive regulators of the phytochrome A signalling pathway, directly bind to the promoter of ELF4, a proposed component of the central oscillator, and activate its expression during the day, whereas the circadian-controlled CCA1 and LHY proteins directly suppress ELF4 expression periodically at dawn through physical interactions with these transcription-promoting factors. Our findings provide evidence that a set of light- and circadian-regulated transcription factors act directly and coordinately at the ELF4 promoter to regulate its cyclic expression, and establish a potential molecular link connecting the environmental light-dark cycle to the central oscillator.

Proteomic response of rice seedling leaves to elevated CO2 levels.

Previous investigations of plant responses to higher CO 2 levels were mostly based on physiological measurements and biochemical assays. In this study, a proteomic approach was employed to investigate plant response to higher CO 2 levels using rice as a model. Ten-day-old seedlings were progressively exposed to 760 ppm, 1140 ppm, and 1520 ppm CO 2 concentrations for 24 h each. The net photosynthesis rate ( P n), stomatal conductance ( G s), transpiration rate ( E), and intercellular to ambient CO 2 concentration ratio ( C i/ C a) were measured. P n, G s, and E showed a maximum increase at 1140 ppm CO 2, but further exposure to 1520 ppm for 24 h resulted in down regulation of these. Proteins extracted from leaves were subjected to 2-DE analysis, and 57 spots showing differential expression patterns, as detected by profile analysis, were identified by MALDI-TOF/TOF-MS. Most of the proteins belonged to photosynthesis, carbon metabolism, and energy pathways. Several molecular chaperones and ascorbate peroxidase were also found to respond to higher CO 2 levels. Concomitant with the down regulation of P n and G s, the levels of enzymes of the regeneration phase of the Calvin cycle were decreased. Correlations between the protein profiles and the photosynthetic measurements at the three CO 2 levels were explored.