Sorghum stay-green QTL individually reduce post-flowering drought-induced leaf senescence

Sorghum is an important source of food, feed, and biofuel, especially in the semi-arid tropics because this cereal is well adapted to harsh, drought-prone environments. Post-flowering drought adaptation in sorghum is associated with the stay-green phenotype. Alleles that contribute to this complex trait have been mapped to four major QTL, Stg1-Stg4, using a population derived from BTx642 and RTx7000. Near-isogenic RTx7000 lines containing BTx642 DNA spanning one or more of the four stay-green QTL were constructed. The size and location of BTx642 DNA regions in each RTx7000 NIL were analysed using 62 DNA markers spanning the four stay-green QTL. RTx7000 NILs were identified that contained BTx642 DNA completely or partially spanning Stg1, Stg2, Stg3, or Stg4. NILs were also identified that contained sub-portions of each QTL and various combinations of the four major stay-green QTL. Physiological analysis of four RTx7000 NILs containing only Stg1, Stg2, Stg3, or Stg4 showed that BTx642 alleles in each of these loci could contribute to the stay-green phenotype. RTx7000 NILs containing BTx642 DNA corresponding to Stg2 retained more green leaf area at maturity under terminal drought conditions than RTx7000 or the other RTx7000 NILs. Under post-anthesis water deficit, a trend for delayed onset of leaf senescence compared with RTx7000 was also exhibited by the Stg2, Stg3, and Stg4 NILs, while significantly lower rates of leaf senescence in relation to RTx7000 were displayed by all of the Stg NILs to varying degrees, but particularly by the Stg2 NIL. Greener leaves at anthesis relative to RTx7000, indicated by higher SPAD values, were exhibited by the Stg1 and Stg4 NILs. The RTx7000 NILs created in this study provide the starting point for in-depth analysis of stay-green physiology, interaction among stay-green QTL and map-based cloning of the genes that underlie this trait.     

Hormonally regulated programmed cell death in barley aleurone cells

Cell death was studied in barley (cv Himalaya) aleurone cells treated with abscisic acid and gibberellin. Aleurone protoplasts incubated in abscisic acid remained viable in culture for at least 3 weeks, but exposure to gibberellin initiated a series of events that resulted in death. Between 4 and 8 days after incubation in gibberellin, >70% of all protoplasts died. Death, which occurred after cells became highly vacuolated, was manifest by an abrupt loss of plasma membrane integrity followed by rapid shrinkage of the cell corpse. Hydrolysis of DNA began before death and occurred as protoplasts ceased production of alpha-amylase. DNA degradation did not result in the accumulation of discrete low molecular weight fragments. DNA degradation and cell death were prevented by LY83583, an inhibitor of gibberellin signaling in barley aleurone. We conclude that cell death in aleurone cells is hormonally regulated and is the final step of a developmental program that promotes successful seedling establishment.     

Molecular mapping of QTLs conferring stay-green in grain sorghum (Sorghum bicolor L. Moench).

Drought resistance is of enormous importance in crop production. The identification of genetic factors involved in plant response to drought stress provides a strong foundation for improving drought tolerance. Stay-green is a drought resistance trait in sorghum (Sorghum bicolor L. Moench) that gives plants resistance to premature senescence under severe soil moisture stress during the post-flowering stage. The objective of this study was to map quantitative trait loci (QTLs) that control the stay-green and chlorophyll content in sorghum. By using a restriction fragment length polymorphism (RFLP) map, developed from a recombinant inbred line (RIL) population, we identified four stay-green QTLs, located on three linkage groups. The QTLs (Stg1 and Stg2) are on linkage group A, with the other two, Stg3 and Stg4, on linkage groups D and J, respectively. Two stay-green QTLs, Stg1 and Stg2, explaining 13-20% and 20-30% of the phenotypic variability, respectively, were consistently identified in all trials at different locations in two years. Three QTLs for chlorophyll content (Chl1, Chl2, and Chl3), explaining 25-30% of the phenotypic variability were also identified under post-flowering drought stress. All coincided with the three stay-green QTL regions (Stg1, Stg2, and Stg3) accounting for 46% of the phenotypic variation. The Stg1 and Stg2 regions also contain the genes for key photosynthetic enzymes, heat shock proteins, and an abscisic acid (ABA) responsive gene. Such spatial arrangement shows that linkage group A is important for drought- and heat-stress tolerance and yield production in sorghum. High-resolution mapping and cloning of the consistent stay-green QTLs may help to develop drought-resistant hybrids and to understand the mechanism of drought-induced senescence in plants.     

Optimization of protein synthesis in isolated higher plant chloroplasts. Identification of paused translation intermediates

Protein synthesis in isolated, intact pea chloroplasts was optimized and compared to translation within chloroplasts in vivo. Many polypeptides labeled with [35S]methionine in isolated intact chloroplasts did not comigrate with polypeptides which were labeled within chloroplasts in vivo. Antibodies to the large subunit of ribulose-1,5-bisphosphate carboxylase-oxygenase (EC 4.1.1.39) immunoprecipitated [35S]-labeled large subunit plus several lower-molecular-mass translation products of isolated chloroplasts. The lower-molecular-mass soluble translation products synthesized in pulse-labeled chloroplasts were converted into full-length large-subunit polypeptides during a subsequent chase period. This result suggests that many of the polypeptides observed in pulse-labeled chloroplasts are incomplete translation products which are the result of ribosome pausing at discrete points along chloroplast mRNAs. The pulse-chase technique was used to follow synthesis of the 34.5-kDa precursor of the psb A gene product and its processing to the mature 32-kDa polypeptide in isolated chloroplasts. Chloroplast translation profiles obtained using the pulse-chase assay were very similar to translation profiles obtained in vivo thus extending the utility of protein synthesis in isolated chloroplasts.     

Latitudinal clines in body size,but not in thermal tolerance or heat‐shock cognate 70 (HSC70), in the highly‐dispersing intertidal gastropod Littorina keenae (Gastropoda: Littorinidae)

Natural populations of widely‐distributed animals often exhibit clinal variation in phenotypic traits or in allele frequencies of a particular gene over their geographical range. A planktotrophic intertidal snail, Littorina keenae is broadly distributed along the north‐eastern Pacific coast through a large latitudinal range (24°50′N–43°18′N). We tested for latitudinal clines in two complex phenotypic traits – thermal tolerance and body size – and one single locus trait – heat shock cognate 70 (HSC70) – in L. keenae along almost its entire geographical range. We found only weak evidence for a latitudinal cline in the thermal tolerance and no evidence for a cline in allele frequencies at HSC70. However, as predicted by Bergmann's rule, we detected a strong latitudinal cline that accounted for 60% of the variance in body size (R² = 0.598; P < 0.001). In contrast, body size did not significantly affect thermal tolerance. HSC70 showed no genetic differentiation among the populations, supporting our previous mitochondrial gene‐based estimate of high gene flow during this snail's free‐swimming larval stage. Given that L. keenae experiences panmixia along its species range, the observed size cline may be partially or entirely caused by a phenotypically plastic response to local thermal environments rather than by genetic divergence in body size among populations in response to locally optimizing natural selection. © 2010 The Linnean Society of London, Biological Journal of the Linnean Society, 2010, 100 , 494–505.     

Sensitive and specific high performance liquid chromatographic method for methionine and leucine enkephalins

The pentapeptides with oplate-like properties, methionine-enkephalin (ME) and leucine enkephalin (LE) have been demonstrated in the brain and gut of animals from various species by means of radioimmunoassay (RIA), bioassay, radioreceptor binding, and immunohistochemical methods. Methods utilizing reverse phase high performance liquid chromatography (HPLC) to separate and quantitate peptides lacked the sensitivity and/or specificity for the determination of endogenous biological levels. This report provides an improved HPLC method having sufficient sensitivity and specificity to allow the separation and quantitation of biological tissue levels of ME and LE.