Nitrated Cyclic GMP Modulates Guard Cell Signaling in Arabidopsis

Nitric oxide (NO) is a ubiquitous signaling molecule involved in diverse physiological processes, including plant senescence and stomatal closure. The NO and cyclic GMP (cGMP) cascade is the main NO signaling pathway in animals, but whether this pathway operates in plant cells, and the mechanisms of its action, remain unclear. Here, we assessed the possibility that the nitrated cGMP derivative 8-nitro-cGMP functions in guard cell signaling. Mass spectrometry and immunocytochemical analyses showed that abscisic acid and NO induced the synthesis of 8-nitro-cGMP in guard cells in the presence of reactive oxygen species. 8-Nitro-cGMP triggered stomatal closure, but 8-bromoguanosine 3′,5′-cyclic monophosphate (8-bromo-cGMP), a membrane-permeating analog of cGMP, did not. However, in the dark, 8-bromo-cGMP induced stomatal opening but 8-nitro-cGMP did not. Thus, cGMP and its nitrated derivative play different roles in the signaling pathways that lead to stomatal opening and closure. Moreover, inhibitor and genetic studies showed that calcium, cyclic adenosine-5′-diphosphate-ribose, and SLOW ANION CHANNEL1 act downstream of 8-nitro-cGMP. This study therefore demonstrates that 8-nitro-cGMP acts as a guard cell signaling molecule and that a NO/8-nitro-cGMP signaling cascade operates in guard cells.     

Seasonal and site-specific variability in terrigenous particulate organic carbon concentration in near-shore waters of Lake Biwa, Japan

Identifying sources of particulate organic matter (POM) is important for clarifying fundamental mechanisms by which lake food webs are sustained. We determined carbon and nitrogen stable isotope ratios of POM in near-shore waters of Lake Biwa, a large, meso-eutrophic lake in Japan, to estimate relative contributions of terrigenous particulate organic carbon (T-POC), plankton-derived POC (P-POC) and epilithon-derived POC (E-POC) to POC in near-shore waters. Samples were collected during different months (November, February, May and July) at 29 sites located near the mouth of tributary rivers with different discharge and catchment land use. The data revealed that POC mainly consisted of P-POC and T-POC, with relative contributions varying widely over season and among locations. E-POC generally contributed little to the near-shore POC. Path analyses revealed that the concentration of riverine POC whose isotopic signatures were similar to those of rice straws increased with a larger %paddy field area in the catchment of tributary rivers, which subsequently enhanced T-POC inputs to near-shore waters through riverine transportation. Furthermore, our results suggested that T-POC contribution was influenced, with a time lag, by wave-driven turbulence and shore topography, which appear to affect sedimentation and resuspension of T-POC.     

Genetically engineered trees for plantation forests: key considerations for environmental risk assessment

Forests are vital to the world's ecological, social, cultural and economic well‐being yet sustainable provision of goods and services from forests is increasingly challenged by pressures such as growing demand for wood and other forest products, land conversion and degradation, and climate change. Intensively managed, highly productive forestry incorporating the most advanced methods for tree breeding, including the application of genetic engineering (GE), has tremendous potential for producing more wood on less land. However, the deployment of GE trees in plantation forests is a controversial topic and concerns have been particularly expressed about potential harms to the environment. This paper, prepared by an international group of experts in silviculture, forest tree breeding, forest biotechnology and environmental risk assessment (ERA) that met in April 2012, examines how the ERA paradigm used for GE crop plants may be applied to GE trees for use in plantation forests. It emphasizes the importance of differentiating between ERA for confined field trials of GE trees, and ERA for unconfined or commercial‐scale releases. In the case of the latter, particular attention is paid to characteristics of forest trees that distinguish them from shorter‐lived plant species, the temporal and spatial scale of forests, and the biodiversity of the plantation forest as a receiving environment.     

Permeabilization induced by lipid II-targeting lantibiotic nisin and its effect on the bioconversion of vitamin D3 to 25-hydroxyvitamin D3 by Rhodococcus erythropolis

Vitamin D₃ (VD₃) is a fat-soluble prohormone in mammals. VD₃ is inert and must be activated by hydroxylation at the C-25 and C-1α positions to exert its biological activity. We recently accomplished the bioconversion of VD₃ to 25(OH)VD₃ with a recombinant strain of Rhodococcus erythropolis and found that the permeability of VD₃ into the cytoplasm may be the rate-limiting step of 25(OH)VD₃ production (Sallam et al., 2010). When the cells were treated with the lipid II-targeting lantibiotic nisin, the permeability of green chemiluminescent cyclodextrin (GCCD), which is used as a model substrate instead of VD₃-partially methylated-β-cyclodextrin (PMCD) complex, was drastically induced. Nisin also induced VD₃ hydroxylation, and the rate was correlated with the expression levels of Vdh and its redox partner proteins. In the bioconversion reaction, the stability of the redox partner proteins and the additional NADH-regenerating system are crucial for VD₃ hydroxylation. The degradation rate of the [2Fe–2S] cluster of ferredoxin ThcC from R. erythropolis NI86/21 is faster than that of AciB from Acinetobacter sp. OC4. Therefore, the nisin-treated R. erythropolis cells coexpressing Vdh and AciBC (1176.5 μg) exhibited much greater 25(OH)VD₃ production than the cells coexpressing Vdh and ThcCD (431.7 μg) after four consecutive 16 h reactions. These results suggest that nisin forms nisin-lipid II pore complexes in the Rhodococcus membrane that increase the accessibility of VD₃–PMCD complexes to the inside of the cells. Furthermore, nisin-treated Rhodococcus cells can be utilized for the bioconversion of other fat-soluble chemicals.     

Distribution of aromatase and sex steroid receptors in the baculum during the rat life cycle: effects of estrogen during the early development of the baculum

The baculum, also called os penis, plays an important role during copulation. However, the hormonal regulation of its development remains to be elucidated. To determine the direct involvement of sex steroids in the development of the baculum of rats, the distributions of androgen receptors (ARs), aromatase, and estrogen receptor alpha (ESR1) were observed immunohistochemically. On Postnatal Day 1, the rudiment of the baculum expressed ARs, aromatase, and ESR1. In the proximal segment of the baculum of neonatal rats, ARs were expressed in the parosteal layer but not in the periosteum or osteoblasts. Aromatase was expressed from the parosteal layer to the endosteum, particularly in the inner osteogenic layer. ESR1 was also abundantly expressed in almost all cells from the parosteal layer to the endosteum. ARs, aromatase, and ESR1 were all abundantly expressed during the neonatal period in the hyaline cartilage of the proximal segment and in fibrocartilage of the distal segment of the baculum. Expression in all the tissues was attenuated in an age-dependent manner and became quite weak at puberty. To determine the effect of estrogen on the growth of the baculum, the aromatase inhibitor 1,4,6-androstatrien-3,17-dione (ATD) was subcutaneously injected daily into pregnant rats from Days 19 to 23 of gestation and into pups on postnatal Days 1, 3, 5, 7, and 9. On Day 10, the length of the baculum in the ATD-treated rats was significantly shorter than that in the controls, although the body weight did not change. These findings suggest that not only androgen but also locally aromatized estrogen is involved in the early growth and development of the baculum.     

Role of amino-terminal half of the S4-S5 linker in type 1 ryanodine receptor (RyR1) channel gating

The type 1 ryanodine receptor (RyR1) is a Ca(2+) release channel found in the sarcoplasmic reticulum of skeletal muscle and plays a pivotal role in excitation-contraction coupling. The RyR1 channel is activated by a conformational change of the dihydropyridine receptor upon depolarization of the transverse tubule, or by Ca(2+) itself, i.e. Ca(2+)-induced Ca(2+) release (CICR). The molecular events transmitting such signals to the ion gate of the channel are unknown. The S4-S5 linker, a cytosolic loop connecting the S4 and S5 transmembrane segments in six-transmembrane type channels, forms an α-helical structure and mediates signal transmission in a wide variety of channels. To address the role of the S4-S5 linker in RyR1 channel gating, we performed alanine substitution scan of N-terminal half of the putative S4-S5 linker (Thr(4825)-Ser(4829)) that exhibits high helix probability. The mutant RyR1 was expressed in HEK cells, and CICR activity was investigated by caffeine-induced Ca(2+) release, single-channel current recordings, and [(3)H]ryanodine binding. Four mutants (T4825A, I4826A, S4828A, and S4829A) had reduced CICR activity without changing Ca(2+) sensitivity, whereas the L4827A mutant formed a constitutive active channel. T4825I, a disease-associated mutation for malignant hyperthermia, exhibited enhanced CICR activity. An α-helical wheel representation of the N-terminal S4-S5 linker provides a rational explanation to the observed activities of the mutants. These results suggest that N-terminal half of the S4-S5 linker may form an α-helical structure and play an important role in RyR1 channel gating.