The Arabidopsis Protein CONSERVED ONLY IN THE GREEN LINEAGE160 Promotes the Assembly of the Membranous Part of the Chloroplast ATP Synthase

The chloroplast F₁F_o-ATP synthase/ATPase (cpATPase) couples ATP synthesis to the light-driven electrochemical proton gradient. The cpATPase is a multiprotein complex and consists of a membrane-spanning protein channel (comprising subunit types a, b, b′, and c) and a peripheral domain (subunits α, β, γ, δ, and ε). We report the characterization of the Arabidopsis (Arabidopsis thaliana) CONSERVED ONLY IN THE GREEN LINEAGE160 (AtCGL160) protein (AtCGL160), conserved in green algae and plants. AtCGL160 is an integral thylakoid protein, and its carboxyl-terminal portion is distantly related to prokaryotic ATP SYNTHASE PROTEIN1 (Atp1/UncI) proteins that are thought to function in ATP synthase assembly. Plants without AtCGL160 display an increase in xanthophyll cycle activity and energy-dependent nonphotochemical quenching. These photosynthetic perturbations can be attributed to a severe reduction in cpATPase levels that result in increased acidification of the thylakoid lumen. AtCGL160 is not an integral cpATPase component but is specifically required for the efficient incorporation of the c-subunit into the cpATPase. AtCGL160, as well as a chimeric protein containing the amino-terminal part of AtCGL160 and Synechocystis sp. PCC6803 Atp1, physically interact with the c-subunit. We conclude that AtCGL160 and Atp1 facilitate the assembly of the membranous part of the cpATPase in their hosts, but loss of their functions provokes a unique compensatory response in each organism.The majority of cellular energy is stored in the form of ATP synthesized by the ubiquitous F₁F_o-ATP synthase (F₁ stands for coupling factor 1, F_o for coupling factor o), which is found in the energy-transducing membranes of bacteria, mitochondria, and chloroplasts. The chloroplast F₁F_o-ATP synthase/ATPase (cpATPase) is a rotary motor that is responsible for coupling ATP synthesis (and hydrolysis) to the light-driven electrochemical proton gradient. The cpATPase comprises two physically separable parts, chloroplast coupling factor o (CF_o), which is an integral membrane-spanning proton channel, and chloroplast coupling factor 1 (CF₁), which is located peripheral to the membrane and contains the catalytic site(s) for reversible ATP synthesis (for review, see von Ballmoos et al., 2009). CF_o comprises four different subunit types, designated b (synonymously, I or AtpF), b′ (II or AtpG), c (III or AtpH), and a (IV or AtpI), and contains one each of subunits a, b, and b′ and a ring made up of 14 copies of subunit c. CF₁ comprises five different subunits, α (AtpA), β (AtpB), γ (AtpC), δ (AtpD), and ε (AtpE), and its subunit composition is α₃β₃γδε (for review, see von Ballmoos et al., 2009).The passage of protons through the CF_o motor drives rotation of the ring of c-subunits, which together form a rotor. The c-ring is connected to subunit γ, and rotation of γ causes conformational changes in the catalytic nucleotide-binding sites of the CF₁ motor, resulting in the synthesis and release of ATP (for review, see Okuno et al., 2011). This process is made possible by the fact that CF₁ and CF_o are physically connected by two stalks, a central one containing the ε- and γ-subunits and a peripheral one made up of δ, b, and b′ (for review, see Böttcher and Gräber, 2000; Weber, 2007). There are six nucleotide-binding sites in CF₁, one at each of the αβ-subunit interfaces about halfway along the vertical axis of the hexamer. Three of the sites are located primarily on the β-subunits and are catalytic; the other three are noncatalytic and probably regulatory. While the three-dimensional structure of the α₃β₃ hexamer in chloroplasts has been solved to a resolution of 3.2 Å (Groth and Pohl, 2001), the structure of the entire CF_o has not yet been determined. However, the conformation of the ring-forming part of CF_o from spinach (Spinacia oleracea) chloroplasts has been defined and found to consist of 14 c-units (Vollmar et al., 2009), whereas the c-ring of the ATP synthase from the cyanobacterium Spirulina platensis contains 15 units (Pogoryelov et al., 2009).Similar to other thylakoid multiprotein complexes like PSII and PSI as well as the cytochrome b₆f complex (Cyt b₆f), the assembly of the ATP synthase must be tightly regulated. Moreover, the variable stoichiometry of the constituents of F₁ (three α/β-subunits versus one each of γ, δ, and ε) and F_o (10–15 c-subunits versus one each of a, b, and b′) requires coordination of the expression of the corresponding genes. This is particularly important in eukaryotes, where the genes are located in different compartments, for instance, in the case of the cpATPase, in the plastid (for α, β, ε, a, b, and c) and the nucleus (for b′, γ, and δ).The assembly of ATP synthase has been most extensively studied in Saccharomyces cerevisiae mitochondria, leading to the identification of several factors involved in this process (for review, see Rak et al., 2009). Thus, three proteins in yeast are known to be involved in the assembly of the α₃β₃ hexamer of F₁. Atp11p (Ackerman and Tzagoloff, 1990a; Wang and Ackerman, 1996) and Atp12p (Ackerman and Tzagoloff, 1990a; Wang and Ackerman, 1998) code for mitochondrial proteins that interact with the β- and α-subunits, respectively, to promote their assembly into the oligomeric F₁-ATPase, and the absence of either protein causes the α- and β-subunits to aggregate into insoluble inclusion bodies in the mitochondrial matrix. Lack of the third protein, FORMATION OF MITOCHONDRIAL COMPLEXES1 (Fmc1p), is associated with aggregation of the α- and β-subunits under heat stress, suggesting that Fmc1p is required for correct folding of Atp12p at elevated temperatures (Lefebvre-Legendre et al., 2001). Originally, the c-ring was assumed to form spontaneously (Arechaga et al., 2002), but subsequent studies have indicated that the assembly of this structural component is also a protein-assisted process. Thus, Atp25p is required for both the synthesis of the c-subunit and its oligomerization into a ring structure of the proper size (Zeng et al., 2008). Moreover, Atp10p (Ackerman and Tzagoloff, 1990b), Atp23p (Osman et al., 2007), and OXIDASE ASSEMBLY1 (Oxa1p) (Jia et al., 2007) are involved in F_o assembly in yeast mitochondria.In prokaryotes, two ATP synthase assembly factors have been described in detail. The membrane protein insertase YidC belongs to the Oxa1 family, is required in vitro for the membrane insertion of subunit c, and assists in the formation of the c-ring from monomers (van der Laan et al., 2004; Kol et al., 2008). In bacterial genomes, the atp1/uncI genes typically precede the genes encoding the structural subunits of the F₁F_o-ATP synthase (for review, see Kol et al., 2008). Moreover, in Synechocystis sp. PCC6803, sll1321/atp1 is coordinately expressed with the seven other genes in the ATP synthase operon (Grossman et al., 2010), implying that Sll1321/Atp1 might have a function associated with the ATP synthase. The genes atp1 and uncI code for small proteins; for instance, Synechocystis sp. PCC6803 Sll1321 has 117 amino acids, and Escherichia coli UncI has 130 amino acids. The function of Atp1/UncI has long remained elusive because deletion of uncI in E. coli results merely in a slightly reduced growth yield (Gay, 1984), indicating that the protein is not essential for the formation of the F₁F_o-ATP synthase complex. Similarly, in the alkaliphilic Bacillus pseudofirmus OF4, Atp1/UncI is not absolutely required for ATP synthase function, and a B. pseudofirmus strain deleted for the atp1 gene could still grow nonfermentatively and its purified ATP synthase had a c-ring of normal size (Liu et al., 2013). Recently, a hybrid F₁F_o (F₁ from Bacillus PS3 and F_o from Propionigenium modestum) was expressed in E. coli. In this system, P. modestum Atp1/UncI was found to be indispensable for c-ring formation and coupled ATPase activity (Suzuki et al., 2007). Similarly, functional production of the Na⁺ F₁F_o-ATP synthase from Acetobacterium woodii in E. coli required the A. woodii atp1/uncI gene for proper assembly (Brandt et al., 2013). Moreover, because subunit c monomers, as well as assembled c-rings, can be copurified together with P. modestum UncI/Atp1 (Suzuki et al., 2007) and the oligomerization of P. modestum c-subunits into c₁₁-rings is mediated by Atp1/UncI in vitro (Ozaki et al., 2008), Atp1/UncI seems to play a role in c-ring assembly for some bacterial ATP synthases.In plants and green algae, regulation of the biogenesis of the cpATPase is well understood at the level of translation of CF₁ subunits (Drapier et al., 2007). Thus, synthesis of the nucleus-encoded subunit γ is required for sustained translation of the chloroplast-encoded subunit β, which in turn transactivates the translation of chloroplast-encoded subunit α. Translational down-regulation of subunit β or α, when not assembled, involves the 5′ untranslated regions (UTRs) of their own mRNAs, pointing to control at the level of translation initiation. In addition, a negative feedback exerted by α/β assembly intermediates on the translation of subunit β can be released when subunit γ assembles with α₃β₃ hexamers.Our knowledge of the nature of true assembly factors for the cpATPase is scarce. So far, only the ALBINO3 homolog Alb4 protein, which can functionally substitute for YidC in E. coli, has been shown to play a role in the biogenesis of the cpATPase, possibly by stabilizing or promoting the assembly of CF₁ during its attachment to the CF_o portion (Benz et al., 2009). Thus, Alb4-Oxa1p-YidC represents an ATP synthase assembly factor family that is conserved between prokaryotes, yeast, and plants. For the bacterial Atp1/UncI protein, one homolog exists in yeast, Vma21p, which is an integral membrane protein localized to the endoplasmic reticulum and is required for vacuolar H⁺-ATPase biogenesis (Graham et al., 1998).In this study, we have identified and characterized a knockout mutant for Arabidopsis (Arabidopsis thaliana) CGL160, a protein that displays moderate similarity to prokaryotic Atp1/UncI proteins in its C-terminal domain. AtCGL160 is required for the efficient assembly of the cpATPase, but lack of AtCGL160 in Arabidopsis has more severe effects on cpATPase assembly than those reported in the literature for inactivation of its prokaryotic relatives and can be located to the assembly of c-subunits into the membranous subcomplex. AtCGL160 physically interacts with the c-subunit of CF_o, and, interestingly, Atp1 can replace the C-terminal part of AtCGL160 in such interactions, indicating that the function of Atp1 and CGL160 proteins is conserved.     

HIV-1 Coreceptor Usage Assessment by Ultra-Deep Pyrosequencing and Response to Maraviroc

Background

Maraviroc is an HIV entry inhibitor that alters the conformation of CCR5 and is poorly efficient in patients infected by viruses that use CXCR4 as an entry coreceptor. The goal of this study was to assess the capacity of ultra-deep pyrosequencing (UDPS) and different data analysis approaches to characterize HIV tropism at baseline and predict the therapeutic outcome on maraviroc treatment.

Methods

113 patients with detectable HIV-1 RNA on HAART were treated with maraviroc. The virological response was assessed at months 1, 3 and 6. The sequence of the HIV V3 loop was determined at baseline and prediction of maraviroc response by different software and interpretation algorithms was analyzed.

Results

UDPS followed by analysis with the Pyrotrop software or geno2pheno algorithm provided better prediction of the response to maraviroc than Sanger sequencing. We also found that the H34Y/S substitution in the V3 loop was the strongest individual predictor of maraviroc response, stronger than substitutions at positions 11 or 25 classically used in interpretation algorithms.

Conclusions

UDPS is a powerful tool that can be used with confidence to predict maraviroc response in HIV-1-infected patients. Improvement of the predictive value of interpretation algorithms is possible and our results suggest that adding the H34S/Y substitution would substantially improve the performance of the 11/25/charge rule.     

Guided bone regeneration produced by new mineralized and reticulated collagen membranes in critical-sized rat calvarial defects

The aim of this study was to evaluate the bone regenerative effect of glutaraldehyde (GA) cross-linking on mineralized polyanionic collagen membranes in critical-sized defects on rat calvarias. Bone calvarial defects were induced in Wistar rats, which were then divided into five groups: a sham group; a control group, which received a commercial membrane; and GA, 25GA, and 75GA groups, which received one of three different polyanionic collagen membranes mineralized by 0, 25, or 75 hydroxyapatite cycles and then cross-linked by GA. Bone formation was evaluated based on digital radiography and computerized tomography. Histological analyses were performed 4 and 12 weeks after the surgical procedure to observe bone formation, membrane resorption, and fibrous tissue surrounding the membranes. Measurement of myeloperoxidase activity, tumor necrosis factor alpha, and interleukin 1beta production was performed 24 h after surgery. The percentage of new bone formation in the GA, 25GA, and 75GA groups was higher compared with the control and sham groups. In the GA and 25 GA groups, the membranes were still in place and were contained in a thick fibrous capsule after 12 weeks. No significant difference was found among the groups regarding myeloperoxidase activity and interleukin 1beta levels, although the GA, 25GA, and 75GA groups presented decreased levels of tumor necrosis factor alpha compared with the control group. These new GA cross-linked membranes accelerated bone healing of the calvarium defects and did not induce inflammation. In addition, unlike the control membrane, the experimental membranes were not absorbed during the analyzed period, so they may offer advantages in large bone defects where prolonged membrane barrier functions are desirable.     

Different properties of ACPA and IgM-RF derived from a large dataset: further evidence of two distinct autoantibody systems

Introduction  

The aim of this study was to examine seroconversion and the relationship with age and inflammation of autoantibodies in a large group of patients attending an outpatient rheumatology clinic.     

Use of methotrexate therapy is not associated with decreased prevalence of metabolic syndrome

With great interest, we read the article by Toms and colleagues [1] in the previous issue of Arthritis Research & Therapy, in which they assessed prevalences of metabolic syndrome (MetS) in rheumatoid arthritis (RA) patients. Moreover, they identified demographic and clinical factors that may be associated with MetS. Toms and colleagues found prevalences of up to 45% of MetS and demonstrated older age and health status (health assessment questionnaire) to be associated with MetS irrespectively of the definition used. Of most interest, an association between methotrexate (MTX) use and decreased presence of MetS was observed in patients more than 60 years of age. The investigators hypothesized that this may be attributed to a drug-specific effect (and not to an anti-inflammatory effect) either by changing levels of adenosine, which is known to interact with glucose and lipid metabolism, or by an indirect effect mediated through concomitant folic acid administration, thereby decreasing homocysteine levels.Recently, we also examined the prevalence of MetS in (a subgroup of) RA patients in the CARRÉ investigation, a prospective cohort study on prevalent and incident cardiovascular disease and its underlying cardiovascular risk factors [2]. The findings of Toms and colleagues stimulated us to perform additional analyses in our total study population (n = 353).The prevalences of MetS were 35% and 25% (Table ​(Table1)1) according to criteria of National Cholesterol Education Program (NCEP) 2004 and NCEP 2001, respectively. In multivariate backward regression analyses, we found significant associations between body mass index, pulse rate, creatinine levels, hypothyroidism and diabetes mellitus and the presence of MetS independently of the criteria used (Table ​(Table2).2). However, an independent association between single use of MTX or use of MTX in combination with other disease-modifying antirheumatic drugs, on the one hand, and a decreased prevalence of MetS, on the other hand, could not be demonstrated (even in the subgroup of patients over the age of 60).

Table 1

Characteristics of the study population

Genome-wide analysis of core promoter elements from conserved human and mouse orthologous pairs

Background  

The canonical core promoter elements consist of the TATA box, initiator (Inr), downstream core promoter element (DPE), TFIIB recognition element (BRE) and the newly-discovered motif 10 element (MTE). The motifs for these core promoter elements are highly degenerate, which tends to lead to a high false discovery rate when attempting to detect them in promoter sequences.     

Kinetic and Spectral Resolution of Multiple Nonphotochemical Quenching Components in Arabidopsis Leaves

Using novel specially designed instrumentation, fluorescence emission spectra were recorded from Arabidopsis (Arabidopsis thaliana) leaves during the induction period of dark to high-light adaptation in order to follow the spectral changes associated with the formation of nonphotochemical quenching. In addition to an overall decrease of photosystem II fluorescence (quenching) across the entire spectrum, high light induced two specific relative changes in the spectra: (1) a decrease of the main emission band at 682 nm relative to the far-red (750–760 nm) part of the spectrum (Δ F₆₈₂); and (2) an increase at 720 to 730 nm (Δ F₇₂₀) relative to 750 to 760 nm. The kinetics of the two relative spectral changes and their dependence on various mutants revealed that they do not originate from the same process but rather from at least two independent processes. The Δ F₇₂₀ change is specifically associated with the rapidly reversible energy-dependent quenching. Comparison of the wild-type Arabidopsis with mutants unable to produce or overexpressing the PsbS subunit of photosystem II showed that PsbS was a necessary component for Δ F₇₂₀. The spectral change Δ F₆₈₂ is induced both by energy-dependent quenching and by PsbS-independent mechanism(s). A third novel quenching process, independent from both PsbS and zeaxanthin, is activated by a high turnover rate of photosystem II. Its induction and relaxation occur on a time scale of a few minutes. Analysis of the spectral inhomogeneity of nonphotochemical quenching allows extraction of mechanistically valuable information from the fluorescence induction kinetics when registered in a spectrally resolved fashion.One of the most important photoprotective mechanisms against high-light (HL) stress in photosynthetic organisms is the nonphotochemical quenching (NPQ) of excitation energy, which is mostly due to thermal deactivation of pigment excited states in the antenna of PSII. There exist a number of literature reviews on the subject (Demmig-Adams and Adams, 1992; Horton et al., 1996; Horton and Ruban, 1999, 2005; Niyogi, 1999, 2000; Müller et al., 2001; Golan et al., 2004; Krause and Jahns, 2004). Chlorophyll (Chl) fluorescence, and in particular pulse amplitude-modulated (PAM) fluorometry as introduced by Schreiber et al. (1986), has become by far the dominant technique to measure NPQ in leaves, chloroplasts, and intact microorganisms (Krause and Weis, 1991; Govindjee, 1995; Maxwell and Johnson, 2000; Krause and Jahns, 2003; Schreiber, 2004), more recently often combined with specific NPQ mutant studies (Golan et al., 2004; Kalituho et al., 2006, 2007; Dall''Osto et al., 2007). In this technique, periodic saturating light pulses are applied, superimposed on the continuous actinic irradiation applied to induce NPQ, in order to transiently close the PSII reaction centers (RCs). Since the photochemistry contribution (photochemical quenching) is thus brought to zero, the method allows us to follow the dynamics of the NPQ development and relaxation by fluorescence in a relatively simple manner (Krause and Jahns, 2003, 2004).Mostly based on its relaxation kinetics, NPQ has been divided technically into the three kinetic components qE, qT, and qI, for the rapid, middle, and slow phases of relaxation (Horton and Hague, 1988), initially attributed to energy-dependent quenching, state transitions, and photoinhibitory quenching (Quick and Stitt, 1989). The rapidly forming and reversible part of NPQ, qE, is the most thoroughly studied. It is well established that this type of quenching is a finely regulated process in which the main governing factors are the proton gradient across the chloroplast thylakoid membrane, Δ pH (Wraight and Crofts, 1970; Briantais et al., 1979), the xanthophyll cycle (i.e. conversion of violaxanthin to antheraxanthin and zeaxanthin [Zx]; Demmig et al., 1987; Demmig-Adams, 1990; Demmig-Adams and Adams, 1992), and the action of the PsbS protein (Funk et al., 1995; Li et al., 2000, 2004; Niyogi et al., 2005). The actual molecular mechanism is still unknown, although there is no shortage of hypotheses and proposed quencher candidates: energy transfer from Chl to Zx in the major light-harvesting complex (LHCII; Frank et al., 2000); electron transfer from a carotenoid to Chl forming a Zx-Chl or lutein-Chl charge-transfer state (Holt et al., 2005; Avenson et al., 2009); direct or indirect quenching by the PsbS protein (Li et al., 2000; Niyogi et al., 2005); energy transfer from Chl to lutein in LHCII (Horton et al., 1991; Ruban et al., 2007) linked to the aggregation of or a conformational change in LHCII; and last but not least, a far-red (FR) light-emitting quenched Chl-Chl charge-transfer state formed by the aggregation of LHCII (Miloslavina et al., 2008). Quenching in the PSII RC has also been proposed (Weis and Berry, 1987; Finazzi et al., 2004; Huner et al., 2005; Ivanov et al., 2008) as an additional type of Zx-independent quenching. Alternatively, it has been suggested that quenching by lutein can complement the Zx-dependent quenching (Niyogi et al., 2001; Li et al. 2009). Johnson et al. (2009) have recently given support to the notion that both Zx-dependent and Zx-independent quenching originate from the same PsbS-dependent mechanism, which is modulated by Zx (Crouchman et al., 2006).While the rapidly relaxing phase qE is now well characterized in its dependence on the various factors, the much slower qT and qI phases are still controversial, and each of them may have contributions from more than one mechanism. The qI component has been traditionally attributed to photoinhibition of PSII (Somersalo and Krause, 1988), associated with coordinated degradation and repair of the photosystem (Powles and Björkman, 1982; Kyle, 1987; Krause, 1988; Aro et al., 1993; Long et al., 1994; Murata et al., 2007). Lately, though, it is more widely accepted that under most conditions the photoinhibition is low and qI, like qE, is a result of thermal deactivation of excited states. Different hypotheses have been put forward to account for its seeming irreversibility: persistent transmembrane Δ pH (Gilmore and Yamamoto, 1992), stable protonation of proteins (Horton et al., 1994), accumulation of inactive PSII reaction centers (Briantais et al., 1992; Schansker and van Rensen, 1999), or stable binding of Zx to CP29 (Färber et al., 1997). The connection of the qT phase with state transitions has been doubted as well, and in fact it is now thought that the fraction of energy redistributed from PSI to PSII under high-light conditions is negligible (Walters and Horton, 1991, 1993) and that the qT must have a different origin or that it has erroneously been ascribed as NPQ (Schansker et al., 2006).Along with the large amount of contradictory evidence on the nature and location of the NPQ quenching site(s), the question of whether the light-induced reversible NPQ represents one single mechanism of deexcitation located in a single site brought about by the combined action of PsbS and Zx (Johnson et al., 2009) or whether it comprises several parallel and largely independent mechanisms acting on different parts of the PSII antenna has not been finally answered. One way to answer this question might be to carefully examine the spectral properties of NPQ-related fluorescence changes. Quenching in different locations of the PSII antenna or with different mechanisms might give rise to a differential quenching in various parts of the PSII antenna that might affect the PSII fluorescence spectra in different ways. This appears possible, since the various pigment-protein complexes of the photosynthetic apparatus have slightly different absorption and emission spectra (Holzwarth, 1991; Holzwarth and Roelofs, 1992). However, in the vast majority of modulated Chl fluorescence instrumentation, including the most widely used PAM fluorometer (Schreiber et al., 1986), the signal is integrated over a broad wavelength range, usually covering the whole range of 710 nm or greater. This integration over the long-wave part of the spectrum has several undesirable consequences and is associated with the unnecessary loss of available information. For example, the fluorescence of PSII peaks in the region of 680 to 685 nm, whereas beyond 700 nm, the PSII fluorescence intensity drops to less than 20% of its peak intensity. In contrast, the fluorescence of intact PSI complexes is dominant in the region above 710 nm (Haehnel et al., 1982; Karukstis and Sauer, 1983; Holzwarth et al., 1985; Holzwarth, 1986; Slavov et al., 2008). Thus, the widely used instrumentation measures the NPQ parameters in a region with reduced PSII contribution and relatively high PSI contribution to total fluorescence, despite the fact that NPQ is generally considered to be primarily a PSII phenomenon. Only in a few studies has the fluorescence in the red and the FR region been separated in order to evaluate the contribution of PSI and its influence on the NPQ parameters (Genty et al., 1990; Peterson et al., 2001). NPQ might also shift the fluorescence properties of the PSII antenna complexes or give rise to entirely new fluorescing components (Miloslavina et al., 2008). This would remain undetected if the NPQ fluorescence changes are not resolved in the spectral domain. It follows from these considerations that a great deal of insight into the NPQ mechanisms and locations may be gained if the spectral dimension is added to the NPQ fluorescence characterization. Among the many advantages of such an approach, one would then be able to distinguish whether NPQ simply leads to a uniform decrease of PSII fluorescence across the emission range or whether this decrease is nonuniform, localized in specific pigment protein complexes, and/or whether new fluorescing species are actually being produced in the NPQ process.The HL-induced NPQ effects on the leaf fluorescence spectra have often been studied also at low temperature, where the differentiation between pigment sites is better (Krause et al., 1983; Demmig and Björkman, 1987; Ruban and Horton, 1994). However, the possibility to resolve the kinetics of NPQ development and relaxation is largely lost when performing the measurements at low temperatures. The 77 K spectra of leaves and thylakoid membranes are characterized by three main peaks, F₆₈₅, F₆₉₅, and F₇₃₀, believed to originate predominantly from Chl a in CP47 of PSII, a specific Chl in CP43 of PSII, and PSI, respectively (Satoh and Butler, 1978; van Dorssen et al., 1987; Andrizhiyevskaya et al., 2005; Komura et al., 2007). Fluorescence from the major LHCII peaks at 680 nm (Rijgersberg and Amesz, 1978) and from the PSII reaction center Chls at 683 nm (Roelofs et al., 1993; Andrizhiyevskaya et al., 2005). Low-temperature studies on the effects of HL irradiation are confined to the changes in the FR-to-red fluorescence ratio, which are the result of the quenching of PSII fluorescence or energy redistribution between the photosystems (state transitions). Ruban and Horton (1994) have shown that photochemical quenching in Guzmania is maximal at 688 nm, whereas nonphotochemical processes quench preferentially at 683 and 698 nm.In this study, we undertook a detailed investigation of the NPQ-associated spectral changes in the fluorescence spectra of Arabidopsis (Arabidopsis thaliana) measured at room temperature (RT) and at 77 K. It follows from the above discussion that deeper insight into the mechanisms of NPQ processes may be gained by combining the kinetic and the spectral information of the fluorescence changes occurring in NPQ. For this purpose, we developed a multiwavelength spectrometer with parallel detection, allowing us to follow the entire time-dependent fluorescence spectra of leaves during the induction and relaxation phases of NPQ with high sensitivity.Specific questions to be addressed in this study are the following. Are there more than one NPQ processes and NPQ sites? Are these processes occurring in a linked fashion or are they independent? How do they depend on the various cofactors known to affect NPQ, in particular regarding the roles of PsbS and Zx? Using this novel approach of adding the spectral information to the NPQ fluorescence changes, we discovered specific spectral changes associated with different NPQ components. By comparing the effects measured on various NPQ mutants of Arabidopsis, it is possible to assign these NPQ components to specific quenching processes. The results provide evidence that the total NPQ is a combination of several parallel and largely independent processes, likely occurring at different locations in the photosynthetic apparatus.     

Chromosome spreading of associated transposable elements and ribosomal DNA in the fish Erythrinus erythrinus. Implications for genome change and karyoevolution in fish

Background  

The fish, Erythrinus erythrinus, shows an interpopulation diversity, with four karyomorphs differing by chromosomal number, chromosomal morphology and heteromorphic sex chromosomes. Karyomorph A has a diploid number of 2n = 54 and does not have differentiated sex chromosomes. Karyomorph D has 2n = 52 chromosomes in females and 2n = 51 in males, and it is most likely derived from karyomorph A by the differentiation of a multiple X₁X₂Y sex chromosome system. In this study, we analyzed karyomorphs A and D by means of cytogenetic approaches to evaluate their evolutionary relationship.     

Levels of anti-citrullinated protein antibodies and IgM rheumatoid factor are not associated with outcome in early arthritis patients: a cohort study

Introduction  

To investigate whether baseline levels of anti-citrullinated protein antibody (ACPA) or IgM rheumatoid factor (IgM-RF) and changes in the year thereafter are associated with disease activity, functional and radiographic outcome in early arthritis patients, and provide additional information over baseline autoantibody status.