Molecular Factors Affecting the Accumulation of Recombinant Proteins in the Chlamydomonas reinhardtii Chloroplast

In an effort to develop microalgae as a robust system for the production of valuable proteins, we analyzed some of the factors affecting recombinant protein expression in the chloroplast of the green alga Chlamydomonas reinhardtii. We monitored mRNA accumulation, protein synthesis, and protein turnover for three codon-optimized transgenes including GFP, bacterial luciferase, and a large single chain antibody. GFP and luciferase proteins were quite stable, while the antibody was less so. Measurements of protein synthesis, in contrast, clearly showed that translation of the three chimeric mRNAs was greatly reduced when compared to endogenous mRNAs under control of the same atpA promoter/UTR. Only in a few conditions this could be explained by limited mRNA availability since, in most cases, recombinant mRNAs accumulated quite well when compared to the atpA mRNA. In vitro toeprint and in vivo polysome analyses suggest that reduced ribosome association might contribute to limited translational efficiency. However, when recombinant polysome levels and protein synthesis are analyzed as a whole, it becomes clear that other steps, such as inefficient protein elongation, are likely to have a considerable impact. Taken together, our results point to translation as the main step limiting the expression of heterologous proteins in the C. reinhardtii chloroplast

Natural isoforms of the Photosystem II D1 subunit differ in photoassembly efficiency of the water-oxidizing complex

© 2015 Springer Science+Business Media Dordrecht. Oxygenic photosynthesis efficiency at increasing solar flux is limited by light-induced damage (photoinhibition) of Photosystem II (PSII), primarily targeting the D1 reaction center subunit. Some cyanobacteria contain two natural isoforms of D1 that function better under low light (D1:1) or high light (D1:2). Herein, rates and yields of photoassembly of the Mn4CaO5 water-oxidizing complex (WOC) from the free inorganic cofactors (Mn2+, Ca2+, water, electron acceptor) and apo-WOC-PSII are shown to differ significantly: D1:1 apo-WOC-PSII exhibits a 2.3-fold faster rate-limiting step of photoassembly and up to seven-fold faster rate to the first light-stable Mn3+ intermediate, IM1, but with a much higher rate of photoinhibition than D1:2. Conversely, D1:2 apo-WOC-PSII assembles slower but has up to seven-fold higher yield, achieved by a higher quantum yield of charge separation and slower photoinhibition rate. These results confirm and extend previous observations of the two holoenzymes: D1:2-PSII has a greater quantum yield of primary charge separation, faster [P680+ Q A- ] charge recombination and less photoinhibition that results in a slower rate and higher yield of photoassembly of its apo-WOC-PSII complex. In contrast, D1:1-PSII has a lower quantum yield of primary charge separation, a slower [P680+ Q A- ] charge recombination rate, and faster photoinhibition that together result in higher rate but lower yield of photoassembly at higher light intensities. Cyanobacterial PSII reaction centers that contain the high- and low-light D1 isoforms can tailor performance to optimize photosynthesis at varying light conditions, with similar consequences on their photoassembly kinetics and yield. These different efficiencies of photoassembly versus photoinhibition impose differential costs for biosynthesis as a function of light intensity

The Role of Resilience in Rebuilding Lives of Injured Veterans

The aim of this commentary is to discuss potential clinical implications of introducing resilience building interventions into care for veterans who are living with a war wound. Some war veterans are expected to live with a wound upon discharge from an active military role and also to fit into civilian life. These lifestyle adjustments can tax the person’s coping abilities and in that context may hinder successful adaptation. The experience of living with a wound or wounds, either acute or chronic, is connected to losses, including loss of mobility, loss of financial capacity (unable to work during some of the wound healing period) and losses attached to changed social roles. Psychological stress is also a common experience for veterans returning to civilian life. Psychological stress is associated with impaired healing or dysregulation of a biomarker associated with wound healing. Modern health practice is centred on symptom reduction and working with pathology however, working with people’s adaptive behaviours such as resilience has not been a consideration. Using the resilience model as a conceptual framework healthcare professionals can engage with veterans towards resilience within the context of their personal experience of ill health. Using this contemporary framework for considering these aspects of care has the potential to facilitate resistance to stressors associated with being injured potentially averting quality of life impairments

Natural variants of photosystem II subunit D1 tune photochemical fitness to solar intensity

Background: Cyanobacteria use multiple PSII-D1 isoforms to adapt to environmental conditions. Results: D1:2 achieves higher quantum efficiency of water oxidation and biomass accumulation rate at high light versus D1:1; the latter is more efficient at low light due to less charge recombination. Conclusion: A functional advantage for D1:1 is revealed for the first time. Significance: Improved photochemical efficiency at low light suggests an evolutionary advantage to retain D1:1. © 2013 by The American Society for Biochemistry and Molecular Biology, Inc

Long-Billed Curlew Nest Site Selection and Success in the Intermountain West

Grassland birds have experienced steeper population declines between 1966 and 2015 than any other bird group on the North American continent, and migratory grassland birds may face threats in all stages of their annual cycle. The grassland‐associated long‐billed curlew (Numenius americanus) is experiencing population declines in regional and local portions of their North American breeding range. The nesting period is an important portion of the annual cycle when curlews may face demographic rate limitations from a suite of threats including predators and anthropogenic disturbance. We compared nest sites to random sites within breeding territories to examine nest site selection, and modeled correlates of nesting success for 128 curlew nests at 5 Intermountain West sites. Nest sites were 6 times more likely than random sites to be situated adjacent to existing cowpies. Additionally, curlews selected nest sites with shorter vegetation, and less bare ground, grass, and shrub cover than at random sites within their territories. Nest success varied widely among sites and ranged from 12% to 40% in a season with a mean of 27% for all nests during the 2015 and 2016 seasons. Higher nest success probability was associated with higher curlew densities in the area, greater percent cover of conspicuous objects (cowpies, rocks) near the nest, and higher densities of black‐billed magpies (Pica hudsonia) and American crows (Corvus brachyrhynchos) at the site. We also found increased probability of nesting success with increased distance from a nest to the nearest potential perch in that territory. Given the central role of working lands to curlews in much of the Intermountain West, understanding limitations to nesting success in these diverse landscapes is necessary to guide adaptive management strategies in increasingly human‐modified habitats. We suggest some grazing and irrigation practices already provide suitable nesting conditions for curlews, and others may require only minor temporal shifts to improve compatibility