Physiological evidence for plasticity in glycolate/glycerate transport during photorespiration

© 2016, The Author(s). Photorespiration recycles fixed carbon following the oxygenation reaction of Ribulose, 1–5, carboxylase oxygenase (Rubisco). The recycling of photorespiratory C2 to C3 intermediates is not perfectly efficient and reduces photosynthesis in C3 plants. Recently, a plastidic glycolate/glycerate transporter (PLGG1) in photorespiration was identified in Arabidopsis thaliana, but it is not known how critical this transporter is for maintaining photorespiratory efficiency. We examined a mutant deficient in PLGG1 (plgg1-1) using modeling, gas exchange, and Rubisco biochemistry. Under low light (under 65 μmol m−2 s−1 PAR), there was no difference in the quantum efficiency of CO2 assimilation or in the photorespiratory CO2 compensation point of plgg1-1, indicating that photorespiration proceeded with wild-type efficiency under sub-saturating light irradiances. Under saturating light irradiance (1200 μmol m−2 s−1 PAR), plgg1-1 showed decreased CO2 assimilation that was explained by decreases in the maximum rate of Rubisco carboxylation and photosynthetic linear electron transport. Decreased rates of Rubisco carboxylation resulted from probable decreases in the Rubisco activation state. These results suggest that glycolate/glycerate transport during photorespiration can proceed in moderate rates through an alternative transport process with wild-type efficiencies. These findings also suggest that decreases in net CO2 assimilation that occur due to disruption to photorespiration can occur by decreases in Rubisco activity and not necessarily decreases in the recycling efficiency of photorespiration

H3K4 methyltransferase Set1 is involved in maintenance of ergosterol homeostasis and resistance to Brefeldin A

Set1 is a conserved histone H3 lysine 4 (H3K4) methyltransferase that exists as a multisubunit complex. Although H3K4 methylation is located on many actively transcribed genes, few studies have established a direct connection showing that loss of Set1 and H3K4 methylation results in a phenotype caused by disruption of gene expression. In this study, we determined that cells lacking Set1 or Set1 complex members that disrupt H3K4 methylation have a growth defect when grown in the presence of the antifungal drug Brefeldin A (BFA), indicating that H3K4 methylation is needed for BFA resistance. To determine the role of Set1 in BFA resistance, we discovered that Set1 is important for the expression of genes in the ergosterol biosynthetic pathway, including the rate-limiting enzyme HMG-CoA reductase. Consequently, deletion of SET1 leads to a reduction in HMG-CoA reductase protein and total cellular ergosterol. In addition, the lack of Set1 results in an increase in the expression of DAN1 and PDR11, two genes involved in ergosterol uptake. The increase in expression of uptake genes in set1δ cells allows sterols such as cholesterol and ergosterol to be actively taken up under aerobic conditions. Interestingly, when grown in the presence of ergosterol set1δ cells become resistant to BFA, indicating that proper ergosterol levels are needed for antifungal drug resistance. These data show that H3K4 methylation impacts gene expression and output of a biologically and medically relevant pathway and determines why cells lacking H3K4 methylation have antifungal drug sensitivity

Synthetic glycolate metabolism pathways stimulate crop growth and productivity in the field

y © The Authors, some rights reserved Photorespiration is required in C3 plants to metabolize toxic glycolate formed when ribulose-1,5-bisphosphate carboxylase-oxygenase oxygenates rather than carboxylates ribulose-1,5-bisphosphate. Depending on growing temperatures, photorespiration can reduce yields by 20 to 50% in C3 crops. Inspired by earlier work, we installed into tobacco chloroplasts synthetic glycolate metabolic pathways that are thought to be more efficient than the native pathway. Flux through the synthetic pathways was maximized by inhibiting glycolate export from the chloroplast. The synthetic pathways tested improved photosynthetic quantum yield by 20%. Numerous homozygous transgenic lines increased biomass productivity by \u3e40% in replicated field trials. These results show that engineering alternative glycolate metabolic pathways into crop chloroplasts while inhibiting glycolate export into the native pathway can drive increases in C3 crop yield under agricultural field conditions

Immediate chromatin immunoprecipitation and on-bead quantitative PCR analysis: A versatile and rapid ChIP procedure

© The Author(s) 2014. Published by Oxford University Press on behalf of Nucleic Acids Research. Genome-wide chromatin immunoprecipitation (ChIP) studies have brought significant insight into the genomic localization of chromatin-associated proteins and histone modifications. The large amount of data generated by these analyses, however, require approaches that enable rapid validation and analysis of biological relevance. Furthermore, there are still protein and modification targets that are difficult to detect using standard ChIP methods. To address these issues, we developed an immediate chromatin immunoprecipitation procedure which we call ZipChip. ZipChip significantly reduces the time and increases sensitivity allowing for rapid screening of multiple loci. Here we describe how ZipChIP enables detection of histone modifications (H3K4 mono- and trimethylation) and two yeast histone demethylases, Jhd2 and Rph1, which were previously difficult to detect using standard methods. Furthermore, we demonstrate the versatility of ZipChIP by analyzing the enrichment of the histone deacetylase Sir2 at heterochromatin in yeast and enrichment of the chromatin remodeler, PICKLE, at euchromatin in Arabidopsis thaliana

Charge-based interaction conserved within histone H3 lysine 4 (H3K4) methyltransferase complexes is needed for protein stability, histone methylation, and gene expression

Histone H3 lysine 4 (H3K4) methyltransferases are conserved from yeast to humans, assemble in multisubunit complexes, and are needed to regulate gene expression. The yeast H3K4 methyltransferase complex, Set1 complex or complex of proteins associated with Set1 (COMPASS), consists of Set1 and conserved Set1-associated proteins: Swd1, Swd2, Swd3, Spp1, Bre2, Sdc1, and Shg1. The removal of the WD40 domain-containing subunits Swd1 and Swd3 leads to a loss of Set1 protein and consequently a complete loss ofH3K4methylation. However, until now, how these WD40 domain-containing proteins interact with Set1 and contribute to the stability of Set1 and H3K4 methylation has not been determined. In this study, we identified small basic and acidic patches that mediate protein interactions between theC terminus of Swd1 and the nSET domain of Set1. Absence of either the basic or acidic patches of Set1 and Swd1, respectively, disrupts the interaction between Set1 and Swd1, diminishes Set1 protein levels, and abolishesH3K4methylation. Moreover, these basic and acidic patches are also important for cell growth, telomere silencing, and gene expression. We also show that the basic and acidic patches of Set1 and Swd1 are conserved in their human counter-parts SET1A/B and RBBP5, respectively, and are needed for the protein interaction between SET1A and RBBP5. Therefore, this charge-based interaction is likely important for maintaining the protein stability of the human SET1A/B methyltransferase complexes so that proper H3K4 methylation, cell growth, and gene expression can also occur in mammals. © 2012 by The American Society for Biochemistry and Molecular Biology, Inc

Optimizing photorespiration for improved crop productivity

© 2018 Institute of Botany, Chinese Academy of Sciences In C3 plants, photorespiration is an energy-expensive process, including the oxygenation of ribulose-1,5-bisphosphate (RuBP) by ribulose 1,5-bisphosphate carboxylase/oxygenase (Rubisco) and the ensuing multi-organellar photorespiratory pathway required to recycle the toxic byproducts and recapture a portion of the fixed carbon. Photorespiration significantly impacts crop productivity through reducing yields in C3 crops by as much as 50% under severe conditions. Thus, reducing the flux through, or improving the efficiency of photorespiration has the potential of large improvements in C3 crop productivity. Here, we review an array of approaches intended to engineer photorespiration in a range of plant systems with the goal of increasing crop productivity. Approaches include optimizing flux through the native photorespiratory pathway, installing non-native alternative photorespiratory pathways, and lowering or even eliminating Rubisco-catalyzed oxygenation of RuBP to reduce substrate entrance into the photorespiratory cycle. Some proposed designs have been successful at the proof of concept level. A plant systems-engineering approach, based on new opportunities available from synthetic biology to implement in silico designs, holds promise for further progress toward delivering more productive crops to farmer\u27s fields

Bile acid sodium symporter BASS6 can transport glycolate and is involved in photorespiratory metabolism in Arabidopsis thaliana

© 2017, American Society of Plant Biologists. All rights reserved. Photorespiration is an energy-intensive process that recycles 2-phosphoglycolate, a toxic product of the Rubisco oxygenation reaction. The photorespiratory pathway is highly compartmentalized, involving the chloroplast, peroxisome, cytosol, and mitochondria. Though the soluble enzymes involved in photorespiration are well characterized, very few membrane transporters involved in photorespiration have been identified to date. In this work, Arabidopsis thaliana plants containing a T-DNA disruption of the bile acid sodium symporter BASS6 show decreased photosynthesis and slower growth under ambient, but not elevated CO2. Exogenous expression of BASS6 complemented this photorespiration mutant phenotype. In addition, metabolite analysis and genetic complementation of glycolate transport in yeast showed that BASS6 was capable of glycolate transport. This is consistent with its involvement in the photorespiratory export of glycolate from Arabidopsis chloroplasts. An Arabidopsis double knockout line of both BASS6 and the glycolate/glycerate transporter PLGG1 (bass6, plgg1) showed an additive growth defect, an increase in glycolate accumulation, and reductions in photosynthetic rates compared with either single mutant. Our data indicate that BASS6 and PLGG1 partner in glycolate export from the chloroplast, whereas PLGG1 alone accounts for the import of glycerate. BASS6 and PLGG1 therefore balance the export of two glycolate molecules with the import of one glycerate molecule during photorespiration

Overexpressing the H-protein of the glycine cleavage system increases biomass yield in glasshouse and field grown transgenic tobacco plants

Photorespiration is essential for C3 plants, enabling oxygenic photosynthesis through the scavenging of 2‐phosphoglycolate. Previous studies have demonstrated that overexpression of the L‐ and H‐proteins of the photorespiratory glycine cleavage system results in an increase in photosynthesis and growth in Arabidopsis thaliana. Here, we present evidence that under controlled environment conditions an increase in biomass is evident in tobacco plants overexpressing the H‐protein. Importantly, the work in this paper provides a clear demonstration of the potential of this manipulation in tobacco grown in field conditions, in two separate seasons. We also demonstrate the importance of targeted overexpression of the H‐protein using the leaf‐specific promoter ST‐LS1. Although increases in the H‐protein driven by this promoter have a positive impact on biomass, higher levels of overexpression of this protein driven by the constitutive CaMV 35S promoter result in a reduction in the growth of the plants. Furthermore in these constitutive overexpressor plants, carbon allocation between soluble carbohydrates and starch is altered, as is the protein lipoylation of the enzymes pyruvate dehydrogenase and alpha‐ketoglutarate complexes. Our data provide a clear demonstration of the positive effects of overexpression of the H‐protein to improve yield under field conditions

Alternative pathway to photorespiration protects growth and productivity at elevated temperatures in a model crop.

Adapting crops to warmer growing season temperatures is a major challenge in mitigating the impacts of climate change on crop production. Warming temperatures drive greater evaporative demand and can directly interfere with both reproductive and vegetative physiological processes. Most of the world's crop species have C3 photosynthetic metabolism for which increasing temperature means higher rates of photorespiration, wherein the enzyme responsible for fixing CO2 fixes O2 instead followed by an energetically costly recycling pathway that spans several cell compartments. In C3 crops like wheat, rice and soybean, photorespiration translates into large yield losses that are predicted to increase as global temperature warms. Engineering less energy-intensive alternative photorespiratory pathways into crop chloroplasts drives increases in C3 biomass production under agricultural field conditions, but the efficacy of these pathways in mitigating the impact of warmer growing temperatures has not been tested. We grew tobacco plants expressing an alternative photorespiratory pathway under current and elevated temperatures (+5 °C) in agricultural field conditions. Engineered plants exhibited higher photosynthetic quantum efficiency under heated conditions than the control plants, and produced 26% (between 16% and 37%) more total biomass than WT plants under heated conditions, compared to 11% (between 5% and 17%) under ambient conditions. That is, engineered plants sustained 19% (between 11% and 21%) less yield loss under heated conditions compared to non-engineered plants. These results support the theoretical predictions of temperature impacts on photorespiratory losses and provide insight toward the optimisation strategies required to help sustain or improve C3 crop yields in a warming climate

Risk factors for high anti-HHV-8 antibody titers (≥1:51,200) in black, HIV-1 negative South African cancer patients: a case control study

Background: Infection with human herpesvirus 8 (HHV-8) is the necessary causal agent in the development of Kaposi's sarcoma (KS). Infection with HIV-1, male gender and older age all increase risk for KS. However, the geographic distribution of HHV-8 and KS both prior to the HIV/AIDS epidemic and with HIV/AIDS suggest the presence of an additional co-factor in the development of KS. Methods: Between January 1994 and October 1997, we interviewed 2576 black in-patients with cancer in Johannesburg and Soweto, South Africa. Blood was tested for antibodies against HIV-1 and HHV-8 and the study was restricted to 2191 HIV-1 negative patients. Antibodies against the latent nuclear antigen of HHV-8 encoded by orf73 were detected with an indirect immunofluorescence assay. We examined the relationship between high anti-HHV-8 antibody titers (≥1:51,200) and sociodemographic and behavioral factors using unconditional logistic regression models. Variables that were significant at p = 0.10 were included in multivariate analysis. Results: Of the 2191 HIV-1 negative patients who did not have Kaposi's sarcoma, 854 (39.0%) were positive for antibodies against HHV-8 according to the immunofluorescent assay. Among those seropositive for HHV-8, 530 (62.1%) had low titers (1:200), 227 (26.6%) had medium titers (1:51,200) and 97 (11.4%) had highest titers (1:204,800). Among the 2191 HIV-1 negative patients, the prevalence of high anti-HHV-8 antibody titers (≥1:51,200) was independently associated with increasing age (ptrend = 0.04), having a marital status of separated or divorced (p = 0.003), using wood, coal or charcoal as fuel for cooking 20 years ago instead of electricity (p = 0.02) and consuming traditional maize beer more than one time a week (p = 0.02; p-trend for increasing consumption = 0.05) although this may be due to chance given the large number of predictors considered in this analysis. Conclusions: Among HIV-negative subjects, patients with high anti-HHV-8 antibody titers are characterized by older age. Other associations that may be factors in the development of high anti- HHV-8 titers include exposure to poverty or a low socioeconomic status environment and consumption of traditional maize beer. The relationship between these variables and high anti- HHV-8 titers requires further, prospective study