Chlamydomonas reinhardtii mutants deficient for Old Yellow Enzyme 3 exhibit increased photooxidative stress.

Old Yellow Enzymes (OYEs) are flavin-containing ene-reductases that have been intensely studied with regard to their biotechnological potential for sustainable chemical syntheses. OYE-encoding genes are found throughout the domains of life, but their physiological role is mostly unknown, one reason for this being the promiscuity of most ene-reductases studied to date. The unicellular green alga Chlamydomonas reinhardtii possesses four genes coding for OYEs, three of which we have analyzed biochemically before. Ene-reductase CrOYE3 stood out in that it showed an unusually narrow substrate scope and converted N-methylmaleimide (NMI) with high rates. This was recapitulated in a C. reinhardtii croye3 mutant that, in contrast to the wild type, hardly degraded externally added NMI. Here we show that CrOYE3-mediated NMI conversion depends on electrons generated photosynthetically by photosystem II (PSII) and that the croye3 mutant exhibits slightly decreased photochemical quenching in high light. Non-photochemical quenching is strongly impaired in this mutant, and it shows enhanced oxidative stress. The phenotypes of the mutant suggest that C. reinhardtii CrOYE3 is involved in the protection against photooxidative stress, possibly by converting reactive carbonyl species derived from lipid peroxides or maleimides from tetrapyrrole degradation.

Bilinguals' inhibitory control and attentional processes in a visual perceptual task.

The aim was to examine theories of bilingual inhibitory control superiority in the visual domain. In an ambiguous figure task, the ability to reverse (switch) interpretations (e.g., duck-rabbit) was examined in 3-5-year-olds bilinguals and monolinguals (N = 67). Bilingualism was no performance predictor in conceptual tasks (Droodle task, false belief task, ambiguous figures production task) that did not pose inhibitory demands. Bilinguals outperformed monolinguals in the ability to reverse, suggesting superior inhibitory capacity per se. Once reversal was experienced there was no difference in the time it took to reverse or reversal frequency between bilinguals and monolinguals. Bayesian analyses confirmed statistical result patterns. Findings support the established view of bilinguals' superior domain-general inhibitory control. This might be brought to bear by attending the environment differently.

Liver MRI susceptibility-weighted imaging (SWI) compared to T2* mapping in the presence of steatosis and fibrosis.

PURPOSE: To show that both susceptibility-weighted imaging (SWI) and T2*-mapping are dependent on liver steatosis, which should be taken into account when using these parameters to grade liver fibrosis and cirrhosis. METHODS: In this prospective study, a total of 174 patients without focal liver disease underwent multiparametric MRI at 3 T including SWI, T1- and T2* mapping, proton density fat fraction (PDFF) quantification and MR elastography. SWI, T2* and T1 were measured in the liver (4 locations), as well as in paraspinal muscles, to calculate the liver-to-muscle ratio (LMR). Liver and LMR values were compared among patients with different steatosis grades (PDFF < 5%, 5-10%, 10-20% and >20%), patients with normal, slightly increased and increased liver stiffness (<2.8 kPa, 2.8-3.5 kPa and >3.5 kPa, respectively). ANOVA with Bonferroni-corrected post hoc tests as well as a multivariate analysis were used to compare values among groups and parameters. RESULTS: SWI and T2* both differed significantly among groups with different steatosis grades (p < 0.001). However, SWI allowed a better differentiation among liver fibrosis grades (p < 0.001) than did T2* (p = 0.05). SWI LMR (p < 0.001) and T2* LMR (p = 0.036) showed a similar performance in differentiating among liver fibrosis grades. CONCLUSION: SWI and T2*-mapping are strongly dependent on the liver steatosis grades. Nevertheless, both parameters are useful predictors for liver fibrosis when using a multiparametric approach.

An investigation of parafoveal masks with the incremental boundary paradigm.

Most of what we know about parafoveal preprocessing during reading is based on the boundary paradigm in combination with parafoveal masks as a presumably neutral baseline condition. Recent evidence questions the neutrality of the baseline condition by showing that parafoveal masks inflict preview costs. Using a novel, incremental boundary paradigm we studied the effect of parafoveal masks. Manipulating the salience of parafoveal previews, we found that increasing salience of the masks resulted in increasingly longer fixation times on target words, but also on pretarget words-suggesting preview costs. We conclude that the hidden preview costs of parafoveal masks in the classical boundary paradigm inflate the processing times for the baseline condition and hence lead to an overestimation of the preview benefit. Thus, the present study questions the validity of some of the conclusions drawn on the basis of the classical boundary paradigm.

CT predicts liver fibrosis: Prospective evaluation of morphology- and attenuation-based quantitative scores in routine portal venous abdominal scans.

OBJECTIVES: Our aim was to prospectively determine whether quantitative computed tomography (CT) scores, consisting of simplified indices for liver remodeling and attenuation, may predict liver fibrosis in abdominal CT scans. MATERIALS AND METHODS: This cross-sectional, prospective study was approved by the local IRB (Kantonale Ethikkommission Bern). Written informed consent was given from all patients undergoing study-MR exams. Between 02/16 and 05/17, four different liver fibrosis scores (CRL-R = caudate-right-lobe ratio, LIMV-, LIMA- and LIMVA-fibrosis score, with "LIM" for liver imaging morphology, "V" for liver vein diameter and "A" for attenuation) were calculated in 1534 consecutive abdominal CT scans, excluding patients with prior liver surgery and liver metastasis. Patients were invited to undergo magnetic resonance (MR) elastography as the non-invasive gold standard to evaluate liver fibrosis. MR elastography shear modulus ≥2.8 kPa was defined as beginning liver fibrosis, while ≥3.5 kPa was defined as significant liver fibrosis (which would correspond to fibrosis stage F2 or higher in histology). Cutoff values, sensitivities and specificities obtained from the receiver operating characteristics (ROC) analysis were then calculated in 141 patients who followed the invitation for MR elastography. To mitigate selection bias, prevalence was estimated in the screened total population (n = 1534) by applying the cutoff values with sensitivities and specificities calculated in the MR elastography sub-group. Positive predictive values (PPV) and negative predictive values (NPV) were then calculated. RESULTS: Fibrosis scores including liver vein attenuation LIMA-FS and LIMVA-FS showed higher areas under the ROC curves (0.96-0.97) than CRL-R (0.82) to detect significant liver fibrosis, while LIMV-FS showed good performance as well (0.92). The prevalence-corrected PPV were 29% for CRL-R, 70% for LIMV-FS, 76% for LIMA-FS and 82% for LIMVA-FS. CONCLUSION: CT fibrosis scores, notably LIMA-FS and LIMVA-FS, may predict significant liver fibrosis on routine abdominal CT scans.

Foveal processing difficulty does not affect parafoveal preprocessing in young readers.

Recent evidence suggested that parafoveal preprocessing develops early during reading acquisition, that is, young readers profit from valid parafoveal information and exhibit a resultant preview benefit. For young readers, however, it is unknown whether the processing demands of the currently fixated word modulate the extent to which the upcoming word is parafoveally preprocessed - as it has been postulated (for adult readers) by the foveal load hypothesis. The present study used the novel incremental boundary technique to assess whether 4th and 6th Graders exhibit an effect of foveal load. Furthermore, we attempted to distinguish the foveal load effect from the spillover effect. These effects are hard to differentiate with respect to the expected pattern of results, but are conceptually different. The foveal load effect is supposed to reflect modulations of the extent of parafoveal preprocessing, whereas the spillover effect reflects the ongoing processing of the previous word whilst the reader's fixation is already on the next word. The findings revealed that the young readers did not exhibit an effect of foveal load, but a substantial spillover effect. The implications for previous studies with adult readers and for models of eye movement control in reading are discussed.

A Ribonucleoprotein Supercomplex Involved in trans-Splicing of Organelle Group II Introns.

In the chloroplast of the green alga Chlamydomonas reinhardtii, two discontinuous group II introns, psaA-i1 and psaA-i2, splice in trans, and thus their excision process resembles the nuclear spliceosomal splicing pathway. Here, we address the question whether fragmentation of trans-acting RNAs is accompanied by the formation of a chloroplast spliceosome-like machinery. Using a combination of liquid chromatography-mass spectrometry (LC-MS), size exclusion chromatography, and quantitative RT-PCR, we provide the first characterization of a high molecular weight ribonucleoprotein apparatus participating in psaA mRNA splicing. This supercomplex contains two subcomplexes (I and II) that are responsible for trans-splicing of either psaA-i1 or psaA-i2. We further demonstrate that both subcomplexes are associated with intron RNA, which is a prerequisite for the correct assembly of subcomplex I. This study contributes further to our view of how the eukaryotic nuclear spliceosome evolved after bacterial endosymbiosis through fragmentation of self-splicing group II introns into a dynamic, protein-rich RNP machinery.

On the Development of Parafoveal Preprocessing: Evidence from the Incremental Boundary Paradigm.

Parafoveal preprocessing of upcoming words and the resultant preview benefit are key aspects of fluent reading. Evidence regarding the development of parafoveal preprocessing during reading acquisition, however, is scarce. The present developmental (cross-sectional) eye tracking study estimated the magnitude of parafoveal preprocessing of beginning readers with a novel variant of the classical boundary paradigm. Additionally, we assessed the association of parafoveal preprocessing with several reading-related psychometric measures. The participants were children learning to read the regular German orthography with about 1, 3, and 5 years of formal reading instruction (Grade 2, 4, and 6, respectively). We found evidence of parafoveal preprocessing in each Grade. However, an effective use of parafoveal information was related to the individual reading fluency of the participants (i.e., the reading rate expressed as words-per-minute) which substantially overlapped between the Grades. The size of the preview benefit was furthermore associated with the children's performance in rapid naming tasks and with their performance in a pseudoword reading task. The latter task assessed the children's efficiency in phonological decoding and our findings show that the best decoders exhibited the largest preview benefit.

Uncoupling of PUMA Expression and Apoptosis Contributes to Functional Heterogeneity in Renal Cell Carcinoma - Prognostic and Translational Implications.

Renal cell carcinoma (RCC) is characterized by a profound disruption of proapoptotic signaling networks leading to chemo- and radioresistance. A key mediator of DNA damage-induced apoptosis is the BH3-only protein PUMA. Given its central role in proapoptotic signaling, we analyzed a series of more than 600 precision-annotated primary RCC specimens for PUMA protein expression. We found a reduced expression of PUMA in 22.6% of RCCs analyzed. Unexpectedly, however, PUMA deficiency was not associated with more aggressive tumor characteristic as expected. Instead, a reduced PUMA expression was associated with a lower TNM stage, lower histopathologic grade, and more favorable cancer-specific patient survival. A direct correlation in a separate patient cohort revealed a profound disconnection between PUMA expression and apoptosis as exemplified by the fact that the tumor with the highest level of apoptotic cells was PUMA deficient. In a series of in vitro studies, we corroborated these results and discovered the highest propensity to undergo apoptosis in an RCC cell line with virtually undetectable PUMA expression. At the same time, PUMA expression was not necessarily associated with stronger apoptosis induction, which underscores the striking functional heterogeneity of PUMA expression and apoptosis in RCC. Collectively, our findings suggest that PUMA-independent mechanisms of cell death exist and may play an important role in suppressing malignant progression. They underscore the functional heterogeneity of RCCs and suggest that PUMA expression alone may not be a suitable predictive biomarker. A better understanding of alternative proapoptotic pathways, however, may help to design novel therapeutic strategies for patients with advanced RCC.

An incremental boundary study on parafoveal preprocessing in children reading aloud: Parafoveal masks overestimate the preview benefit.

Parafoveal preprocessing is an important factor for efficient reading and, in eye-movement studies, is typically investigated by means of parafoveal masking: Valid previews are compared to instances in which masks prevent preprocessing. A long-held assumption was that parafoveal preprocessing, as assessed by this technique, only reflects facilitation (i.e., a preview benefit). Recent studies, however, suggested that the benefit estimate is inflated due to interference of the parafoveal masks, i.e., the masks inflict processing costs. With children from Grades 4 and 6, we administered the novel incremental priming technique. The technique manipulates the salience of the previews by systematically varying its perceptibility (i.e., by visually degrading the previews). This technique does not require a baseline condition, but makes it possible to determine whether a preview induces facilitation or interference. Our salience manipulation of valid previews revealed a preview benefit in the children of both Grades. For two commonly used parafoveal masks, we observed interference corroborating the notion that masks are not a proper baseline. With the novel incremental boundary technique, in contrast, one can achieve an accurate estimate of the preview benefit.

The Octatricopeptide Repeat Protein Raa8 Is Required for Chloroplast trans Splicing.

The mRNA maturation of the tripartite chloroplast psaA gene from the green alga Chlamydomonas reinhardtii depends on various nucleus-encoded factors that participate in trans splicing of two group II introns. Recently, a multiprotein complex was identified that is involved in processing the psaA precursor mRNA. Using coupled tandem affinity purification (TAP) and mass spectrometry analyses with the trans-splicing factor Raa4 as a bait protein, we recently identified a multisubunit ribonucleoprotein (RNP) complex comprising the previously characterized trans-splicing factors Raa1, Raa3, Raa4, and Rat2 plus novel components. Raa1 and Rat2 share a structural motif, an octatricopeptide repeat (OPR), that presumably functions as an RNA interaction module. Two of the novel RNP complex components also exhibit a predicted OPR motif and were therefore considered potential trans-splicing factors. In this study, we selected bacterial artificial chromosome (BAC) clones encoding these OPR proteins and conducted functional complementation assays using previously generated trans-splicing mutants. Our assay revealed that the trans-splicing defect of mutant F19 was restored by a new factor we named RAA8; molecular characterization of complemented strains verified that Raa8 participates in splicing of the first psaA group II intron. Three of six OPR motifs are located in the C-terminal end of Raa8, which was shown to be essential for restoring psaA mRNA trans splicing. Our results support the important role played by OPR proteins in chloroplast RNA metabolism and also demonstrate that combining TAP and mass spectrometry with functional complementation studies represents a vigorous tool for identifying trans-splicing factors.

Inhibitory processes in visual perception: a bilingual advantage.

Bilingual inhibitory control advantages are well established. An open question is whether inhibitory superiority also extends to visual perceptual phenomena that involve inhibitory processes. This research used ambiguous figures to assess inhibitory bilingual superiority in 3-, 4-, and 5-year-old mono- and bilingual children (N=141). Findings show that bilinguals across all ages are superior in inhibiting a prevalent interpretation of an ambiguous figure to perceive the alternative interpretation. In contrast, mono- and bilinguals revealed no differences in understanding that an ambiguous figure can have two distinct referents. Together, these results suggest that early bilingual inhibitory control superiority is also evident in visual perception. Bilinguals' conceptual understanding of figure ambiguity is comparable to that of their monolingual peers.

Identification of a chloroplast ribonucleoprotein complex containing trans-splicing factors, intron RNA, and novel components.

Maturation of chloroplast psaA pre-mRNA from the green alga Chlamydomonas reinhardtii requires the trans-splicing of two split group II introns. Several nuclear-encoded trans-splicing factors are required for the correct processing of psaA mRNA. Among these is the recently identified Raa4 protein, which is involved in splicing of the tripartite intron 1 of the psaA precursor mRNA. Part of this tripartite group II intron is the chloroplast encoded tscA RNA, which is specifically bound by Raa4. Using Raa4 as bait in a combined tandem affinity purification and mass spectrometry approach, we identified core components of a multisubunit ribonucleoprotein complex, including three previously identified trans-splicing factors (Raa1, Raa3, and Rat2). We further detected tscA RNA in the purified protein complex, which seems to be specific for splicing of the tripartite group II intron. A yeast-two hybrid screen and co-immunoprecipitation identified chloroplast-localized Raa4-binding protein 1 (Rab1), which specifically binds tscA RNA from the tripartite psaA group II intron. The yeast-two hybrid system provides evidence in support of direct interactions between Rab1 and four trans-splicing factors. Our findings contribute to our knowledge of chloroplast multisubunit ribonucleoprotein complexes and are discussed in support of the generally accepted view that group II introns are the ancestors of the eukaryotic spliceosomal introns.