Lesion mimic mutants commonly display spontaneous cell death in pre-senescent green leaves under normal conditions, without pathogen attack. Despite molecular and phenotypic characterization of several lesion mimic mutants, the mechanisms of the spontaneous formation of cell death lesions remain largely unknown. Here, we examined the rice lesion mimic mutant spotted leaf3 (spl3). In mutants grown under a light/dark cycle, spl3 mutants appeared similar to wild type at early developmental stages, but lesions gradually appeared in the mature leaves close to heading stage. By contrast, in mutants grown under continuous light, severe cell death lesions formed in developing leaves, even at the seedling stage. Histochemical analysis showed that hydrogen peroxide accumulated in the mutants, likely causing the cell death phenotype. By map-based cloning and complementation, we showed that a 1-bp deletion in the first exon of Oryza sativa Mitogen-Activated Protein Kinase Kinase Kinase1 (OsMAPKKK1)/OsEDR1/ OsACDR1 causes the spl3 mutant phenotype. We found that the spl3 mutants were insensitive to abscisic acid (ABA), showing normal root growth in ABA-containing media and delayed leaf yellowing during dark-induced and natural senescence. Expression of ABA signaling-associated genes was also less responsive to ABA treatment in the mutants. Furthermore, the spl3 mutants had lower transcript levels and activities of catalases, which scavenge hydrogen peroxide, probably due to impairment of ABA-responsive signaling. Finally we discuss a possible molecular mechanism of lesion formation in the mature leaves of spl3 mutants.

Floral transition is influenced by environmental factors such as light and temperature. Plants are capable of integrating photoperiod and ambient temperature signaling into their developmental program. Despite extensive investigations on individual genetic pathways, little is known about the molecular components that integrate both pathways. Here, we demonstrate that the RING finger–containing E3 ubiquitin ligase CONSTITUTIVE PHOTOMORPHOGENIC1 (COP1) acts as an integrator of photoperiod and ambient temperature signaling. In addition to the role in photoperiodic destabilization of CONSTANS (CO), COP1 also regulates temperature sensitivity by controlling the degradation of GIGANTEA (GI). COP1-impaired mutants showed reduced sensitivity to low ambient temperature. Notably, COP1 is more stabilized at low temperature and accelerates GI turnover in a 26S proteasome-dependent manner. The direct association of GI with the promoter of FLOWERING LOCUS T (FT) depends on ambient temperature, and thus COP1-triggered GI turnover delays flowering at low temperatures via a CO-independent pathway. Taken together, our findings indicate that environmental conditions regulate the stability of COP1, and conditional specificity of its target selection stimulates proper developmental responses and ensures reproductive success.

In the facultative long-day (LD) plant Arabidopsis thaliana, FLAVIN-BINDING, KELCH REPEAT, F-BOX1 (FKF1) is activated by blue light and promotes flowering through the transcriptional and post-transcriptional regulation of CONSTANS under inductive LD conditions. By contrast, the facultative short day (SD) plant rice (Oryza sativa) flowers early under inductive SD and late under non-inductive LD conditions; the regulatory function of OsFKF1 remains elusive. Here we show that osfkf1 mutants flower late under SD, LD, and natural LD conditions. Transcriptional analysis revealed that OsFKF1 up-regulates expression of the floral activator Ehd2 and down-regulates expression of the floral repressor Ghd7; these regulators up- and down-regulate Ehd1 expression, respectively. Moreover, OsFKF1 can upregulate Ehd1 expression under blue light treatment, without affecting the expression of Ehd2 and Ghd7. In contrast to the LD-specific floral activator Arabidopsis FKF1, OsFKF1 likely acts as an autonomous floral activator because it promotes flowering independent of photoperiod, probably via its distinct roles in controlling expression of rice-specific genes including Ehd2, Ghd7, and Ehd1. Like Arabidopsis FKF1, which interacts with GI and CDF1, OsFKF1 also interacts with OsGI and OsCDF1 (also termed OsDOF12). Thus, we have identified similar and distinct roles of FKF1 in Arabidopsis and rice.

Heading date and photoperiod sensitivity are fundamental traits that determine rice adaptation to a wide range of geographic environments. By quantitative trait locus (QTL) mapping and candidate gene analysis using wholegenome re-sequencing, we found that Oryza sativa Pseudo-Response Regulator37 (OsPRR37; hereafter PRR37) is responsible for the Early heading7-2 (EH7-2)/Heading date2 (Hd2) QTL which was identified from a cross of late-heading rice ‘Milyang23 (M23)’ and early-heading rice ‘H143’. H143 contains a missense mutation of an invariantly conserved amino acid in the CCT (CONSTANS, CO-like, and TOC1) domain of PRR37 protein. In the world rice collection, different types of nonfunctional PRR37 alleles were found in many European and Asian rice cultivars. Notably, the japonica varieties harboring nonfunctional alleles of both Ghd7/Hd4 and PRR37/Hd2 flower extremely early under natural long-day conditions, and are adapted to the northernmost regions of rice cultivation, up to 53° N latitude. Genetic analysis revealed that the effects of PRR37 and Ghd7 alleles on heading date are additive, and PRR37 down-regulates Hd3a expression to suppress flowering under long-day conditions. Our results demonstrate that natural variations in PRR37/Hd2 and Ghd7/Hd4 have contributed to the expansion of rice cultivation to temperate and cooler regions

The timing of flowering, which is of crucial importance for plant growth and survival, is controlled by intricate pathways. To identify heading date-QTL, we constructed high resolution map on chromosome 3 using heterogeneous inbred family-near isogenic lines (H-NILs) derived from F7 RILs generated by the cross of early-heading japonica rice ‘H143’ and middle-late-heading indica-japonica hybrid cultivar ‘Milyang23’. QTL and subsequent sequence analysis using H-NILs revealed that the gene underlining QTL EH3, which is detected in the region of Hd16, is EL1 encoding casein kinase I (CKI). Two types of single amino acid substitutions in Ser/Thr kinase domain of EL1 were found in various cultivars, among which H-NIL(eh3) caused loss of function in EL1 demonstrated by altered heading date and GA response. Moreover, the phosphorylation of EL1 appears to involve in EL1 activity to regulate heading date. Transcriptional analysis clearly indicated that H-NIL(EH3) suppresses heading under LD conditions by down-regulating Ehd1, there by Hd3a and RFT1 expressions were not induced, suggesting that EL1 is photoperiod-sensitive and functions as a LD-specific suppressor of heading. Further characterization suggested that EL1 is likely to involve in anther development and seed settings by regulating GAMYB expression. Our study demonstrated that the genetic basis of natural variation occurred in ‘H143’ was revealed by QTL analysis using H-NILs, and EH3/EL1 function is crucial for heading and development in rice. The genetic natural variation of H-NIL(eh3) may have contributed to adaptation of rice cultivation to the higher regions by regulating the expression of rice flowering activator genes and GA signaling.

In field conditions, the zebra2 (z2) mutant in rice (Oryza sativa) produces leaves with transverse pale-green/yellow stripes. It was recently reported that ZEBRA2 encodes carotenoid isomerase (CRTISO) and that low levels of lutein, an essential carotenoid for non-photochemical quenching, cause leaf variegation in z2 mutants. However, we found that the z2 mutant phenotype was completely suppressed by growth under continuous light (CL; permissive) conditions, with concentrations of chlorophyll, carotenoids and chloroplast proteins at normal levels in z2 mutants under CL. In addition, three types of reactive oxygen species (ROS; superoxide [O2-], hydrogen peroxide [H2O2], and singlet oxygen [1O2]) accumulated to high levels in z2 mutants grown under short-day conditions (SD; alternate 10-h light/14-h dark; restrictive), but do not accumulate under CL conditions. However, the levels of lutein and zeaxanthin in z2 leaves were much lower than normal in both permissive CL and restrictive SD growth conditions, indicating that deficiency of these two carotenoids is not responsible for the leaf variegation phenotype. We found that the CRTISO substrate tetra-cis-lycopene accumulated during the dark periods under SD, but not under CL conditions. Its accumulation was also positively correlated with 1O2 levels generated during the light period, which consequently altered the expression of 1O2-responsive and cell death-related genes in the variegated z2 leaves. Taking these results together, we propose that the z2 leaf variegation can be largely attributed to photoperiodic accumulation of tetra-cis-lycopene and generation of excessive 1O2 under natural day-night conditions.

Loss of leaf green color results from chlorophyll (Chl) degradation in the chloroplasts, but little is known about how Chl catabolism is tightly regulated throughout development. Using the stay-green (sgr) mutant in rice which maintains leaf greenness during senescence, we identified SGR by map-based cloning. SGR is a function-unknown gene encoding senescence-induced chloroplast protein. Transgenic rice overexpressing SGR produced yellow leaves, indicating that SGR regulates Chl degradation at the transcriptional level. Leaf stay-greenness of the sgr mutant is mainly associated with a failure in the destabilization of light-harvesting complexes (LHCs) of thylakoid membranes, which is a prerequisite event for the degradation of Chl and LHCs during leaf senescence. SGR binds to light harvesting complex of photosystem II (LHCII), but its biochemical role is so far unknown. During senescence, Chl should be degraded rapidly and safely because Chl catabolic intermediates producing ROS under light are extremely toxic to the plant cells. For safe and rapid degradation of Chl and its catabolic intermediates, Chl catabolic enzymes (CCEs) must catch the Chl intermediates effectively. In recent years, although molecular functions of SGR and CCEs have been characterized in detail, their biochemical mechanism for Chl detoxification remain elusive. Here we show that all five CCEs also specifically interact with LHCII. In addition, SGR and CCEs interact directly or indirectly with each other at LHCII, and SGR is essential for recruiting CCEs in senescing chloroplasts. These data indicate a predominant role for the SGR-CCE-LHCII protein interaction in the breakdown of LHCII-located Chl, likely to allow metabolic channeling of phototoxic Chl breakdown intermediates upstream of nontoxic pFCC.

Dehydrins (LEA Dll proteins) are one of the typical families of plant proteins that accumulate in response to dehydration, cold stress, abscisic acid, or during seed maturation. A 1.3-kb cDNA was cloned from a cDNA expression library of 5-day-old germinating maize scutellums under drought stress. The deduced protein sequence indicated a dehydrin gene encoding SK3 LEA protein typically expressed during cold acclimation, but not by drought stress in barley and wheat. Thus, it was named maize DEHYDRIN2 (ZmDhn2). It accumulates rapidly and highly in drought-stressed scutellum and leaf tissues at any stage, but not under cold stress. ZmDhn2 gene was transformed into Arabidopsis thaliana for functional analysis under drought condition. From electrolyte leakage test, no significant difference showed between wild type and transformants under normal growth condition, but the leakage level of electrolyte in wild type plants was about 3 times as high as that in the transformed plants under drought stress. It suggests that ZmDHN2 playa role in increasing drought tolerance.

Barley Yellow Dwarf Virus (BYDV) has been a major disease causing a severe loss of yield in winter cereals worldwide. It has been recently reported that BYDV occurs frequently in wheat field and also causes serious yield reduction in Korea. This study was performed to investigate the regional distributions of BYDV strains in Korea and to identify the resistant cultivars or lines of wheat to the predominant BYDV strains, providing basic information for the breeding of BYDV-resistant wheat varieties. Using RT-PCR and EcoRI digestion methods, the regional distribution of BYDV strains in Korea from 1999 to 2000 showed that PAV strain was mainly detected about 65% (Vic-PAV 52.6% ; CN-PAV 47.4%) and MAV strain about 3%. Using ELISA test for the examination of BYDV resistance with 17 cultivars and 4 lines among Korean wheat, three cultivars, Gurumil, Topdongmil, and Olgurumil, were susceptible to BYDV and the others were resistant. In plant growth and yield component responses to BYDV infection, Gurumil showed significant difference between the uninfected and the infected, suggesting the most susceptible to BYDV among Korean wheat, but Eun-pamil and Seohae118 did no difference, an indication that they have the highest resistance.

Barley Yellow Dwarf Virus (BYDV), an aphid-borne luteovirus, is a major plant pathogenic disease causing a huge economic loss in the grain production of a wide range of Gramineae species throughout the world. It has been recently reported that BYDV also occurred frequently in wheat field of Korea. Here, we performed to develop the detection and classification methods of BYDV strains that were accomplished by reverse transcription-polymerase chain reaction (RT-PCR). Since there are high variations among BYDV strains, three pairs of primers were designed to detect BYDV strains such as PAV (Vic-PAV and CN-PAV) and MAV (primer A) simultaneously, specifically Vic-PAV(primer B), and MAV (primer C) based on the genomic RNA sequences of BYDV strains previously published. The validity of the primers was confirmed using several BYDV strains obtained from CIMMYT. Though three BYDV strains were able to be detected using primer A, PCR products were not distinguished between two PAV strains. It was possible to separate them with a restriction enzyme, EcoRI, whose restriction site was present in the amplified DNA fragment from Vic-PAV, but not from CN-PAV.

Soybean is an important crop because its seed has very high protein relative to others. The quality of soy protein is limited by the concentration of the sulfur-containing amino acids in the amino acid profile. Among the supply of various forms of 0.4mM sulfur as S nutrition during seed fill. only 0.4mM L-methionine can inhibit β-subunit synthesis completely and produce the highest glycinin-containing seeds. Compared to 0.4mM sulfate control, seeds supplied by 0.4mM L-methionine have lower α-, no β-subunit, and highly increased glycinin without altering total protein concentration. Supply of 0.2mM cystine (0.4mM S) did not affect the accumulative pattern of seed storage protein (SSP) subunits. In the supply of L-methionine, 0.2mM treatment showed higher glycinin in seeds but 0.05mM resulted in lower glycinin than tile sulfate control. The relative abundance of α`-subunit was not altered by any N or S nutrition. Under 5mM nitrogen, protein concentration was increased about 3-5% by substituting ammonia for nitrate during seed fill independent of nutrition. The increase resulted in the only increase of 7S protein, mainly β-subunit. Our data suggest that the regulatory system of SSP genes responds to the balance between N and S assimilates supplied from mother plant. and controls the di fferential synthesis of their subunits for the maximum protein accumulation in developing soybean seed.