Quantifying and monitoring functional Photosystem II and the stoichiometry of the two photosystems in leaf segments: Approaches and approximations

Given its unique function in light-induced water oxidation and its susceptibility to photoinactivation during photosynthesis, photosystem II (PS II) is often the focus of studies of photosynthetic structure and function, particularly in environmental stress conditions. Here we review four approaches for quantifying or monitoring PS II functionality or the stoichiometry of the two photosystems in leaf segments, scrutinizing the approximations in each approach. (1) Chlorophyll fluorescence parameters are convenient to derive, but the information-rich signal suffers from the localized nature of its detection in leaf tissue. (2) The gross O2 yield per single-turnover flash in CO2-enriched air is a more direct measurement of the functional content, assuming that each functional PS II evolves one O2 molecule after four flashes. However, the gross O2 yield per single-turnover flash (multiplied by four) could overestimate the content of functional PS II if mitochondrial respiration is lower in flash illumination than in darkness. (3) The cumulative delivery of electrons from PS II to P700? (oxidized primary donor in PS I) after a flash is added to steady background far-red light is a whole-tissue measurement, such that a single linear correlation with functional PS II applies to leaves of all plant species investigated so far. However, the magnitude obtained in a simple analysis (with the signal normalized to the maximum photo-oxidizable P700 signal), which should equal the ratio of PS II to PS I centers, was too small to match the independently-obtained photosystem stoichiometry. Further, an under-estimation of functional PS II content could occur if some electrons were intercepted before reaching PS I. (4) The electrochromic signal from leaf segments appears to reliably quantify the photosystem stoichiometry, either by progressively photoinactivating PS II or suppressing PS I via photo-oxidation of a known fraction of the P700 with steady far-red light. Together, these approaches have the potential for quantitatively probing PS II in vivo in leaf segments, with prospects for application of the latter two approaches in the field

Inactivation of Photosystem I in Cucumber Leaves Exposed to Paraquat-Induced Oxidative Stress

Cucumber leaves subjected to light chilling stress exhibit a preferential inactivation of photosystem (PS) I relative to PSII, resulting in the photoinhibition of photosynthesis. In light-chilled cucumber leaves, Cu/Zn-Superoxide dismutase (SOD) is regarded as a primary target of the light chilling stress and its inactivation is closely related to the increased production of reactive oxygen species. In the present study, we further explored that inactivation of PSI in cucumber leaves is not a light chilling specific, but general to various oxidative stresses. Oxidative stress in cucumber leaves was induced by treatment of methylviologen (MV), a producer of reactive oxygen species in chloroplasts. MV treatment decreased the maximal photosynthetic O 2 evolution, resulting in the photoinhibition of photosynthesis. The photoinhibition of photosynthesis was attributable to the decline in PSI functionality determined in vivo by monitoring absorption changes around 820 nm. In addition, MV treatment inactivated both antioxidant enzymes Cu/Zn-superoxide dismutase and ascorbate peroxidase known sensitive to reactive oxygen species. From these results, we suggest that chloroplast antioxidant enzymes are the primary targets of photooxidative stress, followed by subsequent inactivation of PSI

Bilateral iliac and popliteal arterial thrombosis in a child with focal segmental glomerulosclerosis

Thromboembolic complications (TECs) are clinically important sequelae of nephrotic syndrome (NS). The incidence of TECs in children is approximately 2%–5%. The veins are the most commonly affected sites, particularly the deep veins in the legs, the inferior vena cava, the superior vena cava, and the renal veins. Arterial thrombosis, which is less common, typically occurs in the cerebral, pulmonary, and femoral arteries, and is associated with the use of steroids and diuretics. Popliteal artery thrombosis in children has been described in cases of traumatic dissection, osteochondroma, Mycoplasma pneumoniae infection, and fibromuscular dysplasia. We report of a 33-month-old girl with bilateral iliac and popliteal arterial thrombosis associated with steroid-resistant NS due to focal segmental glomerulosclerosis. Her treatment involved thrombectomy and intravenous heparinization, followed by oral warfarin for 8 months. Herein, we report a rare case of spontaneous iliac and popliteal arterial thrombosis in a young child with NS

Heterologous overexpression of the cyanobacterial alcohol dehydrogenase sysr1 confers cold tolerance to the oleaginous alga Nannochloropsis salina

Temperature is an important regulator of growth in algae and other photosynthetic organisms. Temperatures above or below the optimal growth temperature could cause oxidative stress to algae through accumulation of oxidizing compounds such as reactive oxygen species (ROS). Thus, algal temperature stress tolerance could be attained by enhancing oxidative stress resistance. In plants, alcohol dehydrogenase (ADH) has been implicated in cold stress tolerance, eliciting a signal for the synthesis of antioxidant enzymes that counteract oxidative damage associated with several abiotic stresses. Little is known whether temperature stress could be alleviated by ADH in algae. Here, we generated transgenic lines of the unicellular oleaginous alga Nannochloropsis salina that heterologously expressed sysr1, which encodes ADH in the cyanobacterium Synechocystis sp. PCC 6906. To drive sysr1 expression, the heat shock protein 70 (HSP70) promoter isolated from N. salina was used, as its transcript levels were significantly increased under either cold or heat stress growth conditions. When subjected to cold stress, transgenic N. salina cells were more cold-tolerant than wild-type cells, showing less ROS production but increased activity of antioxidant enzymes such as superoxide dismutase, ascorbate peroxidase, and catalase. Thus, we suggest that reinforcement of alcohol metabolism could be a target for genetic manipulation to endow algae with cold temperature stress tolerance

A novel α-type carbonic anhydrase associated with the thylakoid membrane in Chlamydomonas reinhardtii is required for growth at ambient CO\u3csub\u3e2\u3c/sub\u3e

A 29.5 kDa intracellular α-type carbonic anhydrase, designated Cah3, from the unicellular green alga Chlamydomonas reinhardtii is the first of this type discovered inside a photosynthetic eukaryote cell. We describe the cloning of a cDNA which encodes the protein. Immunoblot studies with specific antibodies raised against Cah3 demonstrate that the polypeptide is associated exclusively with the thylakoid membrane. The putative transit peptide suggests that Cah3 is directed to the thylakoid lumen, which is confirmed further by the presence of mature sized Cah3 after thermolysin treatment of intact thylakoids. Complementation of the high inorganic carbon concentration-requiring mutant, cia3, with a subcloned cosmid containing the cah3 gene yielded transformants that grew on atmospheric levels of CO2 (0.035%) and contained an active 29.5 kDa α-type carbonic anhydrase. Although, cia3 has reduced internal carbonic anhydrase activity, unexpectedly the level of Cah3 was similar to that of the wild-type, suggesting that the mutant accumulates an inactive Cah3 polypeptide. Genomic sequence analysis of the mutant revealed two amino acid changes in the transit peptide. Results from photosynthesis and chlorophyll a fluorescence parameter measurements show that the cia3 mutant is photosynthetically impaired. Our results indicate that the carbonic anhydrase, extrinsically located within the chloroplast thylakoid lumen, is essential for growth of C.reinhardtii at ambient levels of CO2, and that at these CO2 concentrations the enzyme is required for optimal photosystem II photochemistry

Antitumorigenic effect of atmospheric-pressure dielectric barrier discharge on human colorectal cancer cells via regulation of Sp1 transcription factor

Human colorectal cancer cell lines (HT29 and HCT116) were exposed to dielectric barrier discharge (DBD) plasma at atmospheric pressure to investigate the anticancer capacity of the plasma. The dose- and time-dependent effects of DBDP on cell viability, regulation of transcription factor Sp1, cell-cycle analysis, and colony formation were investigated by means of MTS assay, DAPI staining, propidium iodide staining, annexin V-FITC staining, Western blot analysis, RT-PCR analysis, fluorescence microscopy, and anchorage-independent cell transformation assay. By increasing the duration of plasma dose times, significant reductions in the levels of both Sp1 protein and Sp1 mRNA were observed in both cell lines. Also, expression of negative regulators related to the cell cycle (such as p53, p21, and p27) was increased and of the positive regulator cyclin D1 was decreased, indicating that the plasma treatment led to apoptosis and cell-cycle arrest. In addition, the sizes and quantities of colony formation were significantly suppressed even though two cancer promoters, such as TPA and epidermal growth factor, accompanied the plasma treatment. Thus, plasma treatment inhibited cell viability and colony formation by suppressing Sp1, which induced apoptosis and cell-cycle arrest in these two human colorectal cancer cell lines.1

Inhibition of autophagy promotes salinomycin-induced apoptosis via reactive oxygen species-mediated PI3K/AKT/mTOR and ERK/p38 MAPK-dependent signaling in human prostate cancer cells

Recently, the interplay between autophagy and apoptosis has become an important factor in chemotherapy for cancer treatment. Inhibition of autophagy may be an effective strategy to improve the treatment of chemo-resistant cancer by consistent exposure to chemotherapeutic drugs. However, no reports have clearly elucidated the underlying mechanisms. Therefore, in this study, we assessed whether salinomycin, a promising anticancer drug, induces apoptosis and elucidated potential antitumor mechanisms in chemo-resistant prostate cancer cells. Cell viability assay, Western blot, annexin V/propidium iodide assay, acridine orange (AO) staining, caspase-3 activity assay, reactive oxygen species (ROS) production, and mitochondrial membrane potential were assayed. Our data showed that salinomycin alters the sensitivity of prostate cancer cells to autophagy. Pretreatment with 3-methyladenine (3-MA), an autophagy inhibitor, enhanced the salinomycin-induced apoptosis. Notably, salinomycin decreased phosphorylated of AKT and phosphorylated mammalian target of rapamycin (mTOR) in prostate cancer cells. Pretreatment with LY294002, an autophagy and PI3K inhibitor, enhanced the salinomycin-induced apoptosis by decreasing the AKT and mTOR activities and suppressing autophagy. However, pretreatment with PD98059 and SB203580, an extracellular signal-regulated kinases (ERK), and p38 inhibitors, suppressed the salinomycin-induced autophagy by reversing the upregulation of ERK and p38. In addition, pretreatment with N-acetyl-L-cysteine (NAC), an antioxidant, inhibited salinomycin-induced autophagy by suppressing ROS production. Our results suggested that salinomycin induces apoptosis, which was related to ROS-mediated autophagy through regulation of the PI3K/AKT/mTOR and ERK/p38 MAPK signaling pathways