Hydrogen sulfide can inhibit and enhance oxygenic photosynthesis in a cyanobacterium from sulfidic springs

We used microsensors to investigate the combinatory effect of hydrogen sulfide (H₂S) and light on oxygenic photosynthesis in biofilms formed by a cyanobacterium from sulfidic springs. We found that photosynthesis was both positively and negatively affected by H₂S: (i) H₂S accelerated the recovery of photosynthesis after prolonged exposure to darkness and anoxia. We suggest that this is possibly due to regulatory effects of H₂S on photosystem I components and/or on the Calvin cycle. (ii) H₂S concentrations of up to 210 μM temporarily enhanced the photosynthetic rates at low irradiance. Modelling showed that this enhancement is plausibly based on changes in the light‐harvesting efficiency. (iii) Above a certain light‐dependent concentration threshold H₂S also acted as an inhibitor. Intriguingly, this inhibition was not instant but occurred only after a specific time interval that decreased with increasing light intensity. That photosynthesis is most sensitive to inhibition at high light intensities suggests that H₂S inactivates an intermediate of the oxygen evolving complex that accumulates with increasing light intensity. We discuss the implications of these three effects of H₂S in the context of cyanobacterial photosynthesis under conditions with diurnally fluctuating light and H₂S concentrations, such as those occurring in microbial mats and biofilms.     

The PsbY protein is not essential for oxygenic photosynthesis in the cyanobacterium Synechocystis sp. PCC 6803

A tetra-manganese cluster in the photosystem II (PSII) pigment-protein complex plays a critical role in the photosynthetic oxygen evolution process. PsbY, a small membrane-spanning polypeptide, has recently been suggested to provide a ligand for manganese in PSII (A.E. Gau, H.H. Thole, A. Sokolenko, L. Altschmied, R.G. Herrmann, E.K. Pistorius [1998] Mol Gen Genet 260: 56-68). We have constructed a mutant strain of the cyanobacterium Synechocystis sp. PCC 6803 with an inactivated psbY gene (sml0007). Southern-blot and polymerase chain reaction analysis showed that the mutant had completely segregated. However, the DeltapsbY mutant cells grew normally under photoautotrophic conditions. Moreover, growth of the wild-type and mutant cells were similar under high-light photoinhibition conditions, as well as in media without any added manganese, calcium, or chloride, three required inorganic cofactors for the oxygen-evolving complex of PSII. Analysis of steady-state and flash-induced oxygen evolution, fluorescence induction, and decay kinetics, and thermoluminescence profiles demonstrated that the DeltapsbY mutant cells have normal photosynthetic activities. We conclude that the PsbY protein in Synechocystis 6803 is not essential for oxygenic photosynthesis and does not provide an important binding site for manganese in the oxygen-evolving complex of PSII.     

Metabolic engineering of the Chl d-dominated cyanobacterium Acaryochloris marina: production of a novel Chl species by the introduction of the chlorophyllide a oxygenase gene

In oxygenic photosynthetic organisms, the properties of photosynthetic reaction systems primarily depend on the Chl species used. Acquisition of new Chl species with unique optical properties may have enabled photosynthetic organisms to adapt to various light environments. The artificial production of a new Chl species in an existing photosynthetic organism by metabolic engineering provides a model system to investigate how an organism responds to a newly acquired pigment. In the current study, we established a transformation system for a Chl d-dominated cyanobacterium, Acaryochloris marina, for the first time. The expression vector (constructed from a broad-host-range plasmid) was introduced into A. marina by conjugal gene transfer. The introduction of a gene for chlorophyllide a oxygenase, which is responsible for Chl b biosynthesis, into A. marina resulted in a transformant that synthesized a novel Chl species instead of Chl b. The content of the novel Chl in the transformant was approximately 10% of the total Chl, but the level of Chl a, another Chl in A. marina, did not change. The chemical structure of the novel Chl was determined to be [7-formyl]-Chl d(P) by mass spectrometry and nuclear magnetic resonance spectroscopy. [7-Formyl]-Chl d(P) is hypothesized to be produced by the combined action of chlorophyllide a oxygenase and enzyme(s) involved in Chl d biosynthesis. These results demonstrate the flexibility of the Chl biosynthetic pathway for the production of novel Chl species, indicating that a new organism with a novel Chl might be discovered in the future.     

Chlorophyll f-driven photosynthesis in a cavernous cyanobacterium

Chlorophyll (Chl) f is the most recently discovered chlorophyll and has only been found in cyanobacteria from wet environments. Although its structure and biophysical properties are resolved, the importance of Chl f as an accessory pigment in photosynthesis remains unresolved. We found Chl f in a cyanobacterium enriched from a cavernous environment and report the first example of Chl f-supported oxygenic photosynthesis in cyanobacteria from such habitats. Pigment extraction, hyperspectral microscopy and transmission electron microscopy demonstrated the presence of Chl a and f in unicellular cyanobacteria found in enrichment cultures. Amplicon sequencing indicated that all oxygenic phototrophs were related to KC1, a Chl f-containing cyanobacterium previously isolated from an aquatic environment. Microsensor measurements on aggregates demonstrated oxygenic photosynthesis at 742 nm and less efficient photosynthesis under 768- and 777-nm light probably because of diminished overlap with the absorption spectrum of Chl f and other far-red absorbing pigments. Our findings suggest the importance of Chl f-containing cyanobacteria in terrestrial habitats.The textbook concept that oxygenic phototrophs primarily use radiation in the visible range (400–700 nm) has been challenged by several findings of unique cyanobacteria and chlorophylls (Chl) over the past two decades (Miyashita et al., 1996; Chen et al., 2010; Croce and van Amerongen, 2014) Unicellular cyanobacteria in the genus Acaryochloris primarily employ Chl d for oxygenic photosynthesis at 700–720 nm (Miyashita et al., 1996) and thrive in shaded habitats with low levels of visible light but replete of near-infrared radiation (NIR, >700 nm, Kühl et al., 2005; Behrendt et al., 2011, 2012). Furthermore, Chl f was recently discovered in filamentous (Chen et al., 2010; Airs et al., 2014; Gan et al., 2014) and unicellular cyanobacteria (Miyashita et al., 2014), enabling light harvesting even further into the NIR region up to ∼740 nm, often aided by employing additional far-red light-absorbing pigments such as Chl d and phycobiliproteins (Gan et al., 2014). Whereas the biochemical structure (Willows et al., 2013) and biophysical properties (Li et al., 2013; Tomo et al., 2014) of Chl f have been studied in detail, the actual importance of this new chlorophyll for photosynthesis is hardly explored (Li et al., 2014).Chlorophyll f has been found in cyanobacteria originating from aquatic/wet environments: the filamentous Halomicronema hongdechloris from stromatolites in Australia (Chen et al., 2012), a unicellar morphotype (Strain KC1) from Lake Biwa in Japan (Akutsu et al., 2011; Miyashita et al., 2014) and a filamentous Leptolyngbya sp. strain (JSC-1, Gan et al., 2014) from a hot-spring and in a unicellular Chlorogloeopsis fritschii strain from rice paddies (Airs et al., 2014). In this study, we report on a unicellular Chl f-containing cyanobacterium originating from a wet cavernous habitat and demonstrate its capability of NIR-driven oxygenic photosynthesis. Enrichments of the new cyanobacterium were obtained from a dense dark green-blackish biofilm dominated by globular morphotypes of Nostocaceae growing on moist limestone outside Jenolan Caves, NSW, Australia. The sampling site was heavily shaded even during mid-day with low irradiance levels of 400- to 700-nm light varying from 0.5 to 5 μmol photons m⁻² s⁻¹. Biofilms were carefully scraped off the substratum and kept in shaded zip-lock bags in a moist atmosphere until further processing. Samples were then incubated at 28 °C in a f/2 medium under NIR at 720 nm (∼10 μmol photons m⁻² s⁻¹) yielding conspicuous green cell aggregates after several months of incubation. Repeated transfer of the aggregates into fresh medium resulted in a culture predominated by green cell clusters (Figure 1a), exhibiting orange-red fluorescence upon excitation with blue light (Figure 1b). Transmission electron microscopy revealed that the green clusters consisted of slightly elongated unicellular cyanobacteria (∼1- to 2-μm wide and ∼2- to 3-μm long), with stacked thylakoids and embedded in a joint polymer matrix (Figure 1c). Hyperspectral microscopy (Kühl and Polerecky, 2008) of the clusters revealed distinct troughs in the transmission spectra at absorption maxima indicative of Chl a (675–680 nm) and Chl f (∼720 nm; Figure 1d, red line). In situ spectral irradiance measurements at the sampling site showed strong depletion of visible wavelengths in the 480- to 710-nm range (Figure 1d, gray line), whereas highest light levels were found in the near-infrared region of the solar spectrum at 710–900 nm. The presence of Chl a and f was further confirmed in enrichment cultures using high-performance liquid chromatography-based pigment analysis (Figure 1e, Supplementary Figure S1), while no Chl d was detected. In addition, weak spectral signatures of carotenoids and phycobilins, with absorption occurring at ∼495 and 665 nm, were evident in the hyperspectral data. Cyanobacteria, including those producing Chl d/f, are known to actively remodel their pigment content in response to the available light spectrum (Stomp et al., 2007; Chen and Scheer, 2013; Gan et al., 2014) and Chl d/f has almost exclusively been found in cyanobacteria grown under far-red light and not under visible light (Kühl et al., 2005; Chen et al., 2010; Airs et al., 2014; Gan et al., 2014; Li et al., 2014; Miyashita et al., 2014). Recent work describes this acclimation response as ‘Far-Red Light photoacclimation'' (FaRLiP), which, in strain JSC-1, comprises a global change in gene expression and structural remodeling of the PSII/PSI core proteins and phycobilisome constituents (Gan et al., 2014). The extent to which this arrangement results in optimized photosynthetic performance is only known for the NIR (=710 nm)-acclimated strain JSC-1, where exposure to wavelengths >695 nm resulted in 40% higher O₂ evolution rates as compared with cells that were previously adapted to red light (645 nm; Gan et al., 2014). Yet the discrimination of actinic wavelengths and their relative effect on gross photosynthesis in Chl f-containing cells needs further investigation. Using an O₂ microsensor and the light–dark shift method (Revsbech et al., 1983) on embedded Chl f-containing aggregates, we found maximal gross photosynthesis rates (∼1.06 μmol O₂ cm⁻³ s⁻¹) to occur at irradiances of ∼250 μmol photons m⁻² s⁻¹ of 742 nm (half-bandwidth, HBW, 25 nm, Figures 2a and b) with light saturation to occur very early at ∼35 μmol photons m⁻² s⁻¹. Further red-shifted actinic light, that is, 768 nm (HBW 28 nm) and 777 nm (HBW 30 nm), yielded lower O₂ evolution rates, which, in all likelihood, are an effect of the diminished overlap with far-red light-absorbing pigments, including Chl f (Figures 2a and b). As O₂ evolution rates were measured on non-axenic cell aggregates, 16S rDNA amplicon sequencing was employed to determine the microbial diversity found within the enrichment culture. This revealed the presence of a variety of bacterial types, including anoxygenic phototrophs, yet all sequences for known oxygenic phototrophs in the data set (∼9.3% of all reads on the order level, Supplementary Figure S2) formed a single operational taxonomic unit (OTU) closely affiliated with the Chl f-containing strain KC1 (Miyashita et al., 2014, Figure 2c).Open in a separate windowFigure 1Imaging and pigment analysis of Chl f-containing cyanobacteria isolated from a cavernous low-light environment. (a) Representative bright field microscope image of cultured cells grown under 720 nm NIR. (b) Fluorescence image of the same cells as in a, excited at 450–490 nm, with emission being detected at >510 nm. (c) Transmission electron microscopy of a Chl f-containing cyanobacterium with densely stacked thylakoid membranes. (d) Transmittance spectrum of cell aggregate determined by hyperspectral imaging (red line). Ambient light conditions at the site of isolation (gray line), as measured by a spectroradiometer. Note the Chl f-specific in vivo absorption at ∼720 nm in the transmittance spectrum (dotted line). Small insert picture denotes the cells and area of interest (black arrow) from which the spectrum was taken. (e) In vitro absorption spectrum of Chl f extracted from enrichment cultures and analyzed via high-performance liquid chromatography. The two Chl f-specific absorption peaks (404 and 704 nm in acetone:MeOH solvent) are indicated.Open in a separate windowFigure 2Taxonomic affiliation and O₂ evolution of Chl f-containing cells as determined by O₂ microelectrode measurements and 16 S rDNA amplicon sequencing. (a) Emission spectra of narrow-band light-emitting diodes (LEDs) used in this study, with peak emissions at 742, 768 and 777 nm indicated by a–c, respectively. (b) Gross photosynthesis measured via an O₂ microsensor placed in a clump of agarose-embedded Chl f-containing cells. Different NIR irradiance was administered by the LEDs in a and by altering the distance of the LEDs to the embedded cells. (c) Phylogenetic affiliation of known Chl f and/or Chl d-containing cyanobacteria (highlighted in gray) and their respective habitat/place of isolation. Taxonomy was determined by clustering all known oxygenic phototrophs found in enrichment cultures from this study (at order level) into a single OTU (=292 bp length, see Supplementary Materials for details). Phylogeny was inferred using Maximum-likelihood in conjunction with the GTR +I +G nucleotide substitution model, tree stability was tested using bootstrapping with 100 replicates. The analysis involved 39 nucleotide sequences each truncated to a length of 292 bp. Here, the green-sulphur bacterium Chlorobium tepidum TLS was chosen as the outgroup.This advocates that cells from our enrichment culture are related to KC1 cells and supports, in conjunction with further morphological-, physiological- and ultrastructural evidence, that Chl f is extending the usable light spectrum for oxygenic photosynthesis in a cavernous low-light environment. Given the lifestyle and known habitats of recognized Chl d/f-producing cyanobacteria (Figure 2c), we propose that many, if not all, surface-associated cyanobacteria are intrinsically capable of producing far-red light-absorbing pigments and to actively employ them in oxygenic photosynthesis as a result of FaRLiP or similar, yet unknown, mechanisms.     

Uterine artery remodeling in pseudopregnancy is comparable to that in early pregnancy

During pregnancy, the lumenal diameter and wall mass of the uterine artery (UA) increase, most likely in response to the increased hemodynamic strain resulting from the chronically elevated uterine blood flow (UBF). In this remodeling process, the phenotype of vascular smooth-muscle cells (VSMC) is transiently altered to enable VSMC proliferation. These phenomena are already seen during early pregnancy, when the rise in UBF is still modest. This raises the question whether the newly instituted endocrine environment of pregnancy is involved in the onset of the pregnancy-related UA remodeling. We tested the hypothesis that the conceptus is not essential for the onset of UA remodeling of pregnancy. Six control and 18 pseudopregnant (Postcopulation Days 5, 11, and 17; n = 6 per subgroup) C57Bl/6 mice were killed and UAs were dissected and processed for either morphometric analysis or immunohistochemistry. The latter consisted of staining UA cross sections for the differentiation markers smooth muscle alpha-actin and smoothelin, and for the proliferation marker MKI67. We analyzed the UA changes in response to pseudopregnancy by ANOVA. Data are presented as mean +/- SD. By Day 11 of pseudopregnancy, the UA lumen was 25% wider and the media cross-sectional area 71% larger than in control mice. These differences were accompanied by reduced smoothelin expression and increased proliferation of UA medial VSMC. All UA morphological differences had returned or were in the process of returning to baseline values by Day 17 of pseudopregnancy. The structural and cellular aspects of UA remodeling as seen at midpregnancy are also seen in pseudopregnancy. These results support the concept that the conceptus does not contribute to the initiation of UA remodeling. We suggest that ovarian hormones trigger the onset of UA remodeling.     

Arabidopsis carotenoid mutants demonstrate that lutein is not essential for photosynthesis in higher plants.

Lutein, a dihydroxy beta, epsilon-carotenoid, is the predominant carotenoid in photosynthetic plant tissue and plays a critical role in light-harvesting complex assembly and function. To further understand lutein synthesis and function, we isolated four lutein-deficient mutants of Arabidopsis that define two loci, lut1 and lut2 (for lutein deficient). These loci are required for lutein biosynthesis but not for the biosynthesis of beta, beta-carotenoids. The lut1 mutations are recessive, accumulate high levels of zeinoxanthin, which is the immediate precursor of lutein, and define lut1 as a disruption in epsilon ring hydroxylation. The lut2 mutations are semidominant, and their biochemical phenotype is consistent with a disruption of epsilon ring cyclization. The lut2 locus cosegregates with the recently isolated epsilon cyclase gene, thus, providing additional evidence that the lut2 alleles are mutations in the epsilon cyclase gene. It appears likely that the epsilon cyclase is a key step in regulating lutein levels and the ratio of lutein to beta,beta-carotenoids. Surprisingly, despite the absence of lutein, neither the lut1 nor lut2 mutation causes a visible deleterious phenotype or altered chlorophyll content, but both mutants have significantly higher levels of beta, beta-carotenoids. In particular, there is a stable increase in the xanthophyll cycle pigments (violaxanthin, antheraxanthin, and zeaxanthin) in both lut1 and lut2 mutants as well as an increase in zeinoxanthin in lut1 and beta-carotene in lut2. The accumulation of specific carotenoids is discussed as it pertains to the regulation of carotenoid biosynthesis and incorporation into the photosynthetic apparatus. Presumably, particular beta, beta-carotenoids are able to compensate functionally and structurally for lutein in the photosystems of Arabidopsis.     

Tryptophan-heme pi-electrostatic interactions in cytochrome f of oxygenic photosynthesis

Cytochrome f of oxygenic photosynthesis has an unprecedented structure, including the N-terminus being a heme ligand. The adjacent N-terminal heme-shielding domain is enriched in aromatic amino acids. The atomic structures of the chloroplast and cyanobacterial cytochromes f were compared to explain spectral and redox differences between them. The conserved aromatic side chain in the N-terminal heme-shielding peptide at position 4, Phe and Tyr in plants and algae, respectively, and Trp in cyanobacteria, is in contact with the heme. Mutagenesis of cytochrome f from the eukaryotic green alga Chlamydomonas reinhardtii showed that a Phe4 --> Trp substitution in the N-terminal domain was unique in causing a red shift of 1 and 2 nm in the cytochrome Soret (gamma) and Q (alpha) visible absorption bands, respectively. The resulting alpha band peak at 556 nm is characteristic of the cyanobacterial cytochrome. Conversely, a Trp4 --> Phe mutation in the expressed cytochrome from the cyanobacterium Phormidium laminosum caused a blue shift to the 554 nm alpha band peak diagnostic of the chloroplast cytochrome. Residue 4 was found to be the sole determinant of this 60 cm(-)(1) spectral shift, and of approximately one-half of the 70 mV redox potential difference between cytochrome f of P. laminosum and C. reinhardtii (E(m7) = 297 and 370 mV, respectively). The proximity of Trp-4 to the heme implies that the spectral and redox potential shifts arise through differential interaction of its sigma- or pi-electrostatic potential with the heme ring and of the pi-potential with the heme Fe orbitals, respectively. The dependence of the visible spectrum and redox potential of cytochrome f on the identity of aromatic residue 4 provides an example of the use of the relatively sharp cytochrome spectrum as a "spectral fingerprint", and of the novel structural connection between the heme and a single nonliganding residue.     

The evolutionary consequences of oxygenic photosynthesis: a body size perspective

The high concentration of molecular oxygen in Earth??s atmosphere is arguably the most conspicuous and geologically important signature of life. Earth??s early atmosphere lacked oxygen; accumulation began after the evolution of oxygenic photosynthesis in cyanobacteria around 3.0?C2.5 billion years ago (Gya). Concentrations of oxygen have since varied, first reaching near-modern values ~600 million years ago (Mya). These fluctuations have been hypothesized to constrain many biological patterns, among them the evolution of body size. Here, we review the state of knowledge relating oxygen availability to body size. Laboratory studies increasingly illuminate the mechanisms by which organisms can adapt physiologically to the variation in oxygen availability, but the extent to which these findings can be extrapolated to evolutionary timescales remains poorly understood. Experiments confirm that animal size is limited by experimental hypoxia, but show that plant vegetative growth is enhanced due to reduced photorespiration at lower O₂:CO₂. Field studies of size distributions across extant higher taxa and individual species in the modern provide qualitative support for a correlation between animal and protist size and oxygen availability, but few allow prediction of maximum or mean size from oxygen concentrations in unstudied regions. There is qualitative support for a link between oxygen availability and body size from the fossil record of protists and animals, but there have been few quantitative analyses confirming or refuting this impression. As oxygen transport limits the thickness or volume-to-surface area ratio??rather than mass or volume??predictions of maximum possible size cannot be constructed simply from metabolic rate and oxygen availability. Thus, it remains difficult to confirm that the largest representatives of fossil or living taxa are limited by oxygen transport rather than other factors. Despite the challenges of integrating findings from experiments on model organisms, comparative observations across living species, and fossil specimens spanning millions to billions of years, numerous tractable avenues of research could greatly improve quantitative constraints on the role of oxygen in the macroevolutionary history of organismal size.     

Stoichiometric determination of pheophytin in photosystem II of oxygenic photosynthesis

Pheophytin and chlorophyll extracted from oxygen-evolving photosystem II particles, chloroplast thylakoids and cyanobacterial cells were separated by column chromatography with DEAE-Toyopearl, and quantitatively determined by spectrophotometry. The molecular ratio of chlorophyll a+b to pheophytin a was about 100 in spinach photosystem II particles and about 140 in spinach thylakoids. Using flash spectrophotometry of P680 and measurement of flash-induced oxygen yield, the molecular ratio of the chlorophyll to the photochemical reaction center II was determined to be about 200 in the photosystem II particles. These findings suggest that the stoichiometry in photosystem II particles is one reaction center II and two pheophytin a molecules per about 200 chlorophyll molecules. The same stoichiometry for pheophytin to the reaction center II was obtained in the cyanobacteria, Anacystis nidulans and Synechocystis PCC 6714. A quantitative determination of pheophytin a and the electron donor P700 in stroma thylakoids from pokeweed suggests that photosystem I does not contain pheophytin.Dedicated to Prof. L.N.M. Duysens on the occasion of his retirement.     

Total catch of a red-listed marine species is an order of magnitude higher than official data

Accurate information on total catch and effort is essential for successful fisheries management. Officially reported landings, however, may be underestimates of total catch in many fisheries. We investigated the fishery for the nationally red-listed European lobster (Homarus gammarus) in south-eastern Norway. Probability-based strip transect surveys were used to count buoys in the study area in combination with catch per unit effort data obtained independently from volunteer catch diaries, phone interviews, and questionnaires. We estimate that recreational catch accounts for 65% of total catch in the study area. Moreover, our results indicate that only a small proportion (24%) of lobsters landed commercially were sold through the legal market and documented. Total estimated lobster catch was nearly 14 times higher than reported officially. Our study highlights the need for adequate catch monitoring and data collection efforts in coastal areas, presents a clear warning to resource managers that illegal, unreported and unregulated (IUU) fisheries in coastal areas should not be ignored, and shows the potential impact of recreational fisheries.     

Formyl group modification of chlorophyll a: a major evolutionary mechanism in oxygenic photosynthesis

We discuss recent advances in chlorophyll research in the context of chlorophyll evolution and conclude that some derivations of the formyl side chain arrangement of the porphyrin ring from that of the Chl a macrocycle can extend the photosynthetic active radiation (PAR) of these molecules, for example, Chl d and Chl f absorb light in the near‐infrared region, up to ～750 nm. Derivations such as this confer a selective advantage in particular niches and may, therefore, be beneficial for photosynthetic organisms thriving in light environments with particular light signatures, such as red‐ and near‐far‐red light‐enriched niches. Modelling of formyl side chain substitutions of Chl a revealed yet unidentified but theoretically possible Chls with a distinct shift of light absorption properties when compared to Chl a.     

Two unique cyanobacteria lead to a traceable approach of the first appearance of oxygenic photosynthesis

The evolutionary route from anoxygenic photosynthetic bacteria to oxygenic cyanobacteria is discontinuous in terms of photochemical/photophysical reaction systems. It is difficult to describe this transition process simply because there are no recognized intermediary organisms between the two bacterial groups. Gloeobacter violaceus PCC 7421 might be a model organism that is suitable for analysis because it still possesses primordial characteristics such as the absence of thylakoid membranes. Whole genome analysis and biochemical and biophysical surveys of G. violaceus have favored the hypothesis that it is an intermediary organism. On the other hand, species differentiation is an evolutionary process that could be driven by changes in a small number of genes, and this process might give fair information more in details by monitoring of those genes. Comparative studies of genes, including those in Acaryochloris marina MBIC 11017, have provided information relevant to species differentiation; in particular, the acquisition of a new pigment, chlorophyll d, and changes in amino acid sequences have been informative. Here, based on experimental evidence from these two species, we discuss some of the evolutionary pathways for the appearance and differentiation of cyanobacteria.     

Energy expenditure of bipedal walking is higher than that of quadrupedal walking in Japanese macaques

The authors previously compared energetic costs of bipedal and quadrupedal walking in bipedally trained macaques used for traditional Japanese monkey performances (Nakatsukasa et al. 2004 Am. J. Phys. Anthropol. 124:248-256). These macaques used inverted pendulum mechanics during bipedal walking, which resulted in an efficient exchange of potential and kinetic energy. Nonetheless, energy expenditure during bipedal walking was significantly higher than that of quadrupedal walking. In Nakatsukasa et al. (2004 Am. J. Phys. Anthropol. 124:248-256), locomotor costs were measured before subjects reached a steady state due to technical limitations. The present investigation reports sequential changes of energy consumption during 15 min of walking in two trained macaques, using carbon dioxide production as a proxy of energy consumption, as in Nakatsukasa et al. (2004 Am. J. Phys. Anthropol. 124:248-256). Although a limited number of sessions were conducted, carbon dioxide production was consistently greater during bipedal walking, with the exception of some irregularity during the first minute. Carbon dioxide production gradually decreased after 1 min, and both subjects reached a steady state within 10 min. Energy expenditure during bipedalism relative to quadrupedalism differed between the two subjects. It was considerably higher (140% of the quadrupedal walking cost) in one subject who walked with more bent-knee, bent-hip gaits. This high cost strongly suggests that ordinary macaques, who adopt further bent-knee, bent-hip gaits, consume a far greater magnitude of energy during bipedal walking.     

Interactions between the nitrogen signal transduction protein PII and N-acetyl glutamate kinase in organisms that perform oxygenic photosynthesis

PII, one of the most conserved signal transduction proteins, is believed to be a key player in the coordination of nitrogen assimilation and carbon metabolism in bacteria, archaea, and plants. However, the identity of PII receptors remains elusive, particularly in photosynthetic organisms. Here we used yeast two-hybrid approaches to identify new PII receptors and to explore the extent of conservation of PII signaling mechanisms between eubacteria and photosynthetic eukaryotes. Screening of Synechococcus sp. strain PCC 7942 libraries with PII as bait resulted in identification of N-acetyl glutamate kinase (NAGK), a key enzyme in the biosynthesis of arginine. The integrity of Ser49, a residue conserved in PII proteins from organisms that perform oxygenic photosynthesis, appears to be essential for NAGK binding. The effect of glnB mutations on NAGK activity is consistent with positive regulation of NAGK by PII. Phylogenetic and yeast two-hybrid analyses strongly suggest that there was conservation of the NAGK-PII regulatory interaction in the evolution of cyanobacteria and chloroplasts, providing insight into the function of eukaryotic PII-like proteins.     

Sensitivity of Leishmania braziliensis promastigotes to meglumine antimoniate (glucantime) is higher than that of other Leishmania species and correlates with response to therapy in American tegumentary leishmaniasis

The first line drugs for the treatment of leishmaniasis are antimonial derivatives. Poor clinical response may be credited to factors linked to the host, the drug, or the parasite. We determined the sensitivity of Leishmania sp. promastigotes and amastigotes by counting parasites exposed to increasing concentrations of meglumine antimoniate (Glucantime). Leishmania braziliensis promastigotes were significantly more sensitive than those belonging to other species. The sensitivity of L. braziliensis isolates from patients with unfavorable clinical outcome, such as therapeutic failure or relapse, was significantly lower than those from patients who had clinical cure. Poor clinical response to therapy (therapeutic failure or relapse) was also associated with inadequate antimonial therapy. We also found a significant and positive correlation between promastigotes and intracellular amastigotes with regard to their in vitro susceptibilities to meglumine antimoniate. Our data provide evidence for an association between the sensitivity of promastigotes to antimonials in vitro and clinical response to therapy in American tegumentary leishmaniasis. The high sensitivity of the local L. braziliensis to meglumine antimoniate in vitro provides an explanation for the good clinical response of cutaneous leishmaniasis in the municipality of Rio de Janeiro, Brazil, even when low-dose regimens are employed.