Low-CO2-inducible bestrophins outside the pyrenoid sustain high photosynthetic efficacy in diatoms.

Anion transporters sustain a variety of physiological states in cells. Bestrophins belong to a Cl- and/or HCO3- transporter family conserved in bacteria, animals, algae, and plants. Recently, putative bestrophins were found in the green alga Chlamydomonas reinhardtii, where they are up-regulated under low CO2 conditions and play an essential role in the CO2-concentrating mechanism (CCM). The putative bestrophin orthologs are also conserved in diatoms, secondary endosymbiotic algae harboring red-type plastids, but their physiological functions are unknown. Here, we characterized the subcellular localization and expression profile of bestrophins (BSTs) in the marine diatoms Phaeodactylum tricornutum (PtBST1-4) and Thalassiosira pseudonana (TpBST1 and 2). PtBST1, PtBST2, and PtBST4 localized at the stroma thylakoid membrane outside of the pyrenoid, and PtBST3 localized in the pyrenoid. Contrarily, TpBST1 and TpBST2 both localized in the pyrenoid. These bestrophin proteins accumulated in cells grown in atmospheric CO2 (LC) but not in 1% CO2 (HC)-grown cells. To assess the physiological functions, we generated knock-out mutants for the PtBST1gene by genome editing. The lack of PtBST1 decreased photosynthetic affinity for dissolved inorganic carbon to the level comparable to the HC-grown wild type. Furthermore, non-photochemical quenching in LC-grown cells was 1.5-2.0 times higher in the mutants than in the wild type. These data suggest that HCO3- transport at the stroma thylakoid membranes by PtBST1 is a critical part of the CO2-evolving machinery of the pyrenoid in the fully induced CCM and that PtBST1 may modulate photoprotection under CO2-limited environments in P. tricornutum.

Denoising parameter dependence of coronary artery depictability in compressed sensing magnetic resonance angiography.

Using numerical indices and visual evaluation, we evaluated the dependence of coronary-artery depictability on the denoising parameter in compressed sensing magnetic resonance angiography (CS-MRA). This study was conducted to clarify the acceleration factor (AF) and denoising factor (DF) dependence of CS-MRA image quality. Vascular phantoms and clinical images were acquired using three-dimensional CS-MRA on a clinical 1.5 T system. For the phantom measurements, we compared the full width at half maximum (FWHM), sharpness, and contrast ratio of the vascular profile curves for various AFs and DFs. In the clinical cases, the FWHM, sharpness, contrast ratio, signal-to-noise ratio, noise level values, and visual evaluation results were compared for various DFs. Phantom image analyses demonstrated that the respective measurements of the FWHM, sharpness, and contrast ratios did not significantly change with an increase in AF. The FWHM and sharpness measurements slightly changed with the DF level. However, the contrast ratio tended to increase with an increase in the DF level. In the clinical cases, the FWHM and sharpness showed no significant differences, even when the DF level was changed. However, the contrast ratio tended to decrease as the DF level increased. When the DF levels of the clinical cases increased, the background signals of the myocardium, fat, and noise levels decreased. We investigated the dependence of the coronary-artery depictability on AF and DF using CS-MRA. Analysis of the coronary-artery profile curves indicated that a better image quality was achieved with a stronger DF on coronary CS-MRA.

Coordinated phosphate uptake by extracellular alkaline phosphatase and solute carrier transporters in marine diatoms.

Marine diatoms express genes encoding potential phosphate transporter and alkaline phosphatase (APase) under phosphate-limited (-P) condition. This indicates that diatoms use high-affinity phosphate uptake system with organic phosphate hydration. The function of molecules playing roles for Pi uptake was determined in this study. Pi uptake and APase activity of two marine diatoms, Phaeodactylum tricornutum and Thalassiosira pseudonana, were monitored during acclimation to -P condition. The transcript levels of Pi transporter were analyzed, and Pi transporters were localized with GFP tagging in diatom cells. KO mutants of plasma membrane solute carrier proteins (SLC34s) or APase were established, and their phenotype was evaluated. Some Na+ /Pi transporter candidates, SLC34s in P. tricornutum and T. pseudonana, increased transcript under -P condition. Whole-cell Pi transport was specifically stimulated by sodium ion but independent of potassium, lithium, or proton. Genome-editing KO of PtSLC34-5 and APase (Pt49678) in P. tricornutum was highly inhibitory for Pi uptake, and KO of TpSLC34-2 was also highly inhibitory for Pi uptake in T. pseudonana. SLC34s and APase were co-expressed under -P conditions in marine diatoms. SLC34s play a major role in the initial acclimation stage of diatom cells to -P condition and APase plays an increasing role in the prolonged Pi-starved condition.

Localization and characterization θ carbonic anhydrases in Thalassiosira pseudonana.

Carbonic anhydrase (CA) is a crucial component for the operation of CO2-concentrating mechanisms (CCMs) in the majority of aquatic photoautotrophs that maintain the global primary production. In the genome of the centric marine diatom, Thalassiosira pseudonana, there are four putative gene sequences that encode θ-type CA, which was a type of CA recently identified in marine diatoms and green algae. In the present study, specific subcellular locations of four θCAs, TpθCA1, TpθCA2, TpθCA3, and TpθCA4 were determined by expressing GFP-fused proteins of these TpθCAs in T. pseudonana. As a result, C-terminal GFP fusion proteins of TpθCA1, TpθCA2, and TpθCA3 were all localized in the chloroplast; TpθCA2 was at the central chloroplast area, and the other two TpθCAs were throughout the chloroplast. Immunogold-labeling transmission electron microscopy was further performed for the transformants expressing TpθCA1:GFP and TpθCA2:GFP with anti-GFP-monoclonal antibody. TpθCA1:GFP was localized in the free stroma area, including the peripheral pyrenoid area. TpθCA2:GFP was clearly located as a lined distribution at the central part of the pyrenoid structure, which was most likely the pyrenoid-penetrating thylakoid. Considering the presence of the sequence encoding the N-terminal thylakoid-targeting domain in the TpθCA2 gene, this localization was likely the lumen of the pyrenoid-penetrating thylakoid. On the other hand, TpθCA4:GFP was localized in the cytoplasm. Transcript analysis of these TpθCAs revealed that TpθCA2 and TpθCA3 were upregulated in atmospheric CO2 (0.04% CO2, LC) levels, while TpθCA1 and TpθCA4 were highly induced under 1% CO2 (HC) condition. The genome-editing knockout (KO) of TpθCA1, by CRISPR/Cas9 nickase, gave a silent phenotype in T. pseudonana under LC-HC conditions, which was in sharp agreement with the case of the previously reported TpθCA3 KO. In sharp contrast, TpθCA2 KO is so far unsuccessful, suggesting a housekeeping role of TpθCA2. The silent phenotype of KO strains of stromal CAs suggests that TpαCA1, TpθCA1, and TpθCA3 may have functional redundancy, but different transcript regulations in response to CO2 of these stromal CAs suggest in part their independent roles.

Excretion of triacylglycerol as a matrix lipid facilitating apoplastic accumulation of a lipophilic metabolite shikonin.

Plants produce a large variety of lipophilic metabolites, many of which are secreted by cells and accumulated in apoplasts. These compounds often play a role to protect plants from environmental stresses. However, little is known about how these lipophilic compounds are secreted into apoplastic spaces. In this study, we used shikonin-producing cultured cells of Lithospermum erythrorhizon as an experimental model system to analyze the secretion of lipophilic metabolites, taking advantage of its high production rate and the clear inducibility in culture. Shikonin derivatives are lipophilic red naphthoquinone compounds that accumulate exclusively in apoplastic spaces of these cells and also in the root epidermis of intact plants. Microscopic analysis showed that shikonin is accumulated in the form of numerous particles on the cell wall. Lipidomic analysis showed that L. erythrorhizon cultured cells secrete an appreciable portion of triacylglycerol (24-38% of total triacylglycerol), composed predominantly of saturated fatty acids. Moreover, in vitro reconstitution assay showed that triacylglycerol encapsulates shikonin derivatives with phospholipids to form lipid droplet-like structures. These findings suggest a novel role for triacylglycerol as a matrix lipid, a molecular component involved in the secretion of specialized lipophilic metabolites.

A YAK1-type protein kinase, triacylglycerol accumulation regulator 1, in the green alga Chlamydomonas reinhardtii is a potential regulator of cell division and differentiation into gametes during photoautotrophic nitrogen deficiency.

Yet another kinase (YAK) 1 is a conserved eukaryotic protein kinase coordinating growth and development. We previously isolated a mutant of Chlamydomonas reinhardtii defective in the YAK1 ortholog triacylglycerol (TAG) accumulation regulator 1 (TAR1). The mutant tar1-1 displayed higher levels of chlorophyll, starch, TAG, and biomass than the parental strain C9 (renamed as C9-3) in photoautotrophic nitrogen (N)-deficient conditions. However, we found that the parental C9-3 showed faster chlorosis upon N-deficiency than the original C9 (C9-1) freshly recovered from cryopreservation, suggesting that C9-3 had acquired particular characteristics during long-term subculturing. To exclude phenotypes dependent on a particular parental strain, we newly created tar1 mutants from two wild-types, C9-1 and CC 125. Like tar1-1, the new tar1 mutants showed higher levels of chlorophyll and TAG/starch than the parental strain. Upon removal of N, Chlamydomonas cells divide once before ceasing further division. Previously, the single division after N-removal was arrested in tar1-1 in photomixotrophic conditions, but this phenotype was not observed in photoautotrophic conditions because of the particular characteristics of the parental C9-3. However, using C9- 1 and CC-125 as parental strains, we showed that cell division after N-removal was impaired in new tar1 mutants in photoautotrophic conditions. Consistent with the view that the division under N-deficiency is necessary for gametic differentiation, new tar1 mutants showed lower mating efficiency than the parental strains. Taken together, TAR1 was suggested to promote differentiation into gametes through the regulation of cell division in response to N-deficiency.

Multiple plasma membrane SLC4s contribute to external HCO3- acquisition during CO2 starvation in the marine diatom Phaeodactylum tricornutum.

The availability of CO2 is one of the restrictions on aquatic photosynthesis. Solute carrier (SLC) 4-2, a plasma membrane HCO3- transporter has previously been identified in the marine diatom Phaeodactylum tricornutum. In this study, we discovered two paralogs, PtSLC4-1 and PtSLC4-4, that are both localized at the plasma membrane. Their overexpression stimulated HCO3- uptake, and this was inhibited by the anion channel blocker 4,4´-diisothiocyanostilbene-2,2´-disulfonic (DIDS). Similarly to SLC4-2, PtSLC4-1 specifically required Na+ of ~100 mM for its maximum HCO3- transport activity. Unlike PtSLC4-1 and PtSLC4-2, the HCO3- transport of PtSLC4-4 depended equally on Na+, K+, or Li+, suggesting its broad selectivity for cations. Transcript analyses indicated that PtSLC4-1 was the most abundant HCO3- transporter under CO2 concentrations below atmospheric levels, while PtSLC4-4 showed little transcript induction under atmospheric CO2 but transient induction to comparable levels to PtSLC4-1 during the initial acclimation stage from high CO2 (1%) to very low CO2 (<0.002%). Our results strongly suggest a major HCO3- transport role of PtSLC4-1 with a relatively minor role of PtSLC4-2, and that PtSLC4-4 operates under severe CO2 limitation unselectively to cations when the other SLC4s do not function to support HCO3- uptake.

Immobilization of a Broad Range of Polypeptides on the Frustule of the Diatom Thalassiosira pseudonana.

Proteins immobilized on biosilica which have superior reactivity and specificity and are innocuous to natural environments could be useful biological materials in industrial processes. One recently developed technique, living diatom silica immobilization (LiDSI), has made it possible to immobilize proteins, including multimeric and redox enzymes, via a cellular excretion system onto the silica frustule of the marine diatom Thalassiosira pseudonana. However, the number of application examples so far is limited, and the type of proteins appropriate for the technique is still enigmatic. Here, we applied LiDSI to six industrially relevant polypeptides, including protamine, metallothionein, phosphotriesterase, choline oxidase, laccase, and polyamine synthase. Protamine and metallothionein were successfully immobilized on the frustule as protein fusions with green fluorescent protein (GFP) at the N terminus, indicating that LiDSI can be used for polypeptides which are rich in arginine and cysteine. In contrast, we obtained mutants for the latter four enzymes in forms without green fluorescent protein. Immobilized phosphotriesterase, choline oxidase, and laccase showed enzyme activities even after the purification of frustule in the presence of 1% (wt/vol) octylphenoxy poly(ethyleneoxy)ethanol. An immobilized branched-chain polyamine synthase changed the intracellular polyamine composition and silica nanomorphology. These results illustrate the possibility of LiDSI for industrial applications. IMPORTANCE Proteins immobilized on biosilica which have superior reactivity and specificity and are innocuous to natural environments could be useful biological materials in industrial processes. Living diatom silica immobilization (LiDSI) is a recently developed technique for in vivo protein immobilization on the diatom frustule. We aimed to explore the possibility of using LiDSI for industrial applications by successfully immobilizing six polypeptides: (i) protamine (Oncorhynchus keta), a stable antibacterial agent; (ii) metallothionein (Saccharomyces cerevisiae), a metal adsorption molecule useful for bioremediation; (iii) phosphotriesterase (Sulfolobus solfataricus), a scavenger for toxic organic phosphates; (iv) choline oxidase (Arthrobacter globiformis), an enhancer for photosynthetic activity and yield of plants; (v) laccase (Bacillus subtilis), a phenol oxidase utilized for delignification of lignocellulosic materials; and (vi) branched-chain polyamine synthase (Thermococcus kodakarensis), which produces branched-chain polyamines important for DNA and RNA stabilization at high temperatures. This study provides new insights into the field of applied biological materials.

Successful termination of ventricular tachycardia with intrinsic anti-tachycardia pacing.

Intrinsic anti-tachycardia pacing (iATP) is a novel automated ATP algorithm that employs post-pacing interval (PPI) to design the next ATP sequence based on an analysis of the prior failed ATP sequence. A patient with hypertrophic cardiomyopathy received an implantable cardioverter-defibrillator (ICD) (Cobalt™ XT DR, Medtronic, Minneapolis, MN, USA) following an episode of syncope due to macro-reentrant ventricular tachycardia (VT) (right bundle branch block configuration, cycle length [CL] 280 ms). The VF zone was set to VTCL <300 ms and iATP therapy was prescribed before and during capacitor charging. The iATP was initiated when VT recurred 3 months later. The first attempt with an assumption of 150 ms propagation time from the pacing site to the VT circuit (9 pulses) could not reset the VT, leaving a PPI of 650 ms. A subsequent attempt involving 20 pulses with an assumption of 250 ms propagation time terminated the VT. Failure to reach the circuit is a major cause of unsuccessful ATP. In this regard, iATP is expected to have theoretical advantages over empirical and traditional ATP therapies. To the best of our knowledge, this is the first intracardiac electrogram illustrating how automated precision ATP terminates VT in a clinical setting.

Characterization of a CO2-Concentrating Mechanism with Low Sodium Dependency in the Centric Diatom Chaetoceros gracilis.

Microalgae induce a CO2-concentrating mechanism (CCM) to overcome CO2-limiting stress in aquatic environments by coordinating inorganic carbon (Ci) transporters and carbonic anhydrases (CAs). Two mechanisms have been suggested to facilitate Ci uptake from aqueous media: Na+-dependent HCO3- uptake by solute carrier (SLC) family transporters and accelerated dehydration of HCO3- to CO2 by external CA in model diatoms. However, studies on ecologically and industrially important diatoms including Chaetoceros gracilis, a common food source in aquacultures, are still limited. Here, we characterized the CCM of C. gracilis using inhibitors and growth dependency on Na+ and CO2. Addition of a membrane-impermeable SLC inhibitor, 4,4'-diisothiocyano-2,2'-stilbenedisulfonic acid (DIDS), or the transient removal of Na+ from the culture medium did not impair photosynthetic affinity for Ci in CO2-limiting stress conditions, but addition of a membrane-impermeable CA inhibitor, acetazolamide, decreased Ci affinity to one-third of control cultures. In culture medium containing 0.23 mM Na+ C. gracilis grew photoautotrophically by aeration with air containing 5% CO2, but not with the air containing 0.04% CO2. These results suggested that C. gracilis utilizes external CAs in its CCM to elevate photosynthetic affinity for Ci rather than plasma-membrane SLC family transporters. In addition, it is possible that low level of Na+ may support the CCM in processes other than Ci-uptake at the plasma membrane specifically in CO2-limiting conditions. Our findings provide insights into the diversity of CCMs among diatoms as well as basic information to optimize culture conditions for industrial applications.

Metabolic Innovations Underpinning the Origin and Diversification of the Diatom Chloroplast.

: Of all the eukaryotic algal groups, diatoms make the most substantial contributions to photosynthesis in the contemporary ocean. Understanding the biological innovations that have occurred in the diatom chloroplast may provide us with explanations to the ecological success of this lineage and clues as to how best to exploit the biology of these organisms for biotechnology. In this paper, we use multi-species transcriptome datasets to compare chloroplast metabolism pathways in diatoms to other algal lineages. We identify possible diatom-specific innovations in chloroplast metabolism, including the completion of tocopherol synthesis via a chloroplast-targeted tocopherol cyclase, a complete chloroplast ornithine cycle, and chloroplast-targeted proteins involved in iron acquisition and CO2 concentration not shared between diatoms and their closest relatives in the stramenopiles. We additionally present a detailed investigation of the chloroplast metabolism of the oil-producing diatom Fistuliferasolaris, which is of industrial interest for biofuel production. These include modified amino acid and pyruvate hub metabolism that might enhance acetyl-coA production for chloroplast lipid biosynthesis and the presence of a chloroplast-localised squalene synthesis pathway unknown in other diatoms. Our data provides valuable insights into the biological adaptations underpinning an ecologically critical lineage, and how chloroplast metabolism can change even at a species level in extant algae.

Cross-compartment metabolic coupling enables flexible photoprotective mechanisms in the diatom Phaeodactylum tricornutum.

Photoacclimation consists of short- and long-term strategies used by photosynthetic organisms to adapt to dynamic light environments. Observable photophysiology changes resulting from these strategies have been used in coarse-grained models to predict light-dependent growth and photosynthetic rates. However, the contribution of the broader metabolic network, relevant to species-specific strategies and fitness, is not accounted for in these simple models. We incorporated photophysiology experimental data with genome-scale modeling to characterize organism-level, light-dependent metabolic changes in the model diatom Phaeodactylum tricornutum. Oxygen evolution and photon absorption rates were combined with condition-specific biomass compositions to predict metabolic pathway usage for cells acclimated to four different light intensities. Photorespiration, an ornithine-glutamine shunt, and branched-chain amino acid metabolism were hypothesized as the primary intercompartment reductant shuttles for mediating excess light energy dissipation. Additionally, simulations suggested that carbon shunted through photorespiration is recycled back to the chloroplast as pyruvate, a mechanism distinct from known strategies in photosynthetic organisms. Our results suggest a flexible metabolic network in P. tricornutum that tunes intercompartment metabolism to optimize energy transport between the organelles, consuming excess energy as needed. Characterization of these intercompartment reductant shuttles broadens our understanding of energy partitioning strategies in this clade of ecologically important primary producers.

Coefficients in the CAVI Equation and the Comparison Between CAVI With and Without the Coefficients Using Clinical Data.

AIM: The Cardio-Ankle Vascular Index (CAVI) is a stiffness index of the arterial tree from the origin of the aorta to the ankle, independent of blood pressure at the time of measurement. The CAVI equation includes the coefficients "a" and "b" to adjust it to the value of Hasegawa's pulse wave velocity, which is compensated for at 80 mmHg of diastolic pressure. To verify this adjustment with the coefficients, the clinical significance of CAVI and CAVI without the coefficients (haß) were compared in both an epidemiological study and an acute clinical study. METHODS: In the epidemiological study, the significances of CAVI and haß among people with or without coronary risks such as hypertension, dyslipidemia, hyperglycemia, and abnormal electrocardiography change, were compared. In the acute clinical study, nitroglycerin was administered to subjects in a control group and to coronary artery disease patients, observing CAVI and haß values over a 20-min period. RESULTS: There was no discrepancy in terms of statistically significant differences between CAVI and haß among subjects with or without risk factors. Furthermore, there was also no discrepancy in terms of statistically significant differences between CAVI and haß during the changes of those values following nitroglycerin administration over a 20-min period. CONCLUSION: In both the epidemiologic and clinical studies, there was no discrepancy in terms of significant differences between CAVI and haß. These results suggest that both are valid as indices of stiffness of the arterial tree from the origin of the aorta to the ankle.

Influence of the reference scan and scan time on the arterial phase of liver magnetic resonance imaging.

The controlled aliasing in parallel imaging results in higher acceleration (CAIPIRINHA) technique can decrease scan time. The purpose of this study was to determine whether an arterial phase scan can be performed in 5 s using the CAIPIRINHA short-scan and a reference scanning technique. The generalized autocalibrating partially parallel acquisition (GRAPPA), the CAIPIRINHA routine (CAIPI-routine), and the CAIPIRINHA short-scanning (CAIPI-short) methods were compared. The scan time for each method was preset to 20 s, 15 s, and 10 s, respectively. The reference scan had a scan time of 5 s. A phantom study was used to compare the influence of artifacts during the reference scan. For comparison, the phantom was moved during the last 5 s. In the clinical studies of suspected chronic liver diseases, magnetic resonance imaging of the liver is usually performed while the patient is breath-hold. The motion artifacts of each method were compared. Artifacts were reduced in reference scans using the CAIPIRINHA method. At 5 s after initiation, the rate of change in the standard deviation value was within 30% compared to that of the original image. Motion artifacts due to the influence of the reference scan when a patient failed to hold their breath did not complicate image evaluation. The proportion of motion artifacts for each sequence was as follows: GRAPPA, 5.8%; CAIPI-routine, 1.9%; and CAIPI-short, 0.7%. The arterial phase can be scanned in 5 s using the CAIPI-short and reference scan techniques.

Mechanisms of carbon dioxide acquisition and CO2 sensing in marine diatoms: a gateway to carbon metabolism.

Diatoms are one of the most successful marine eukaryotic algal groups, responsible for up to 20% of the annual global CO2 fixation. The evolution of a CO2-concentrating mechanism (CCM) allowed diatoms to overcome a number of serious constraints on photosynthesis in the marine environment, particularly low [CO2]aq in seawater relative to concentrations required by the CO2 fixing enzyme, ribulose-1,5-bisphosphate carboxylase/oxygenase (RubisCO), which is partly due to the slow diffusion rate of CO2 in water and a limited CO2 formation rate from [Formula: see text] in seawater. Diatoms use two alternative strategies to take up dissolved inorganic carbon (DIC) from the environment: one primarily relies on the direct uptake of [Formula: see text] through plasma-membrane type solute carrier (SLC) 4 family [Formula: see text] transporters and the other is more reliant on passive diffusion of CO2 formed by an external carbonic anhydrase (CA). Bicarbonate taken up into the cytoplasm is most likely then actively transported into the chloroplast stroma by SLC4-type transporters on the chloroplast membrane system. Bicarbonate in the stroma is converted into CO2 only in close proximity to RubisCO preventing unnecessary CO2 leakage. CAs play significant roles in mobilizing DIC as it is progressively moved towards the site of fixation. However, the evolutionary types and subcellular locations of CAs are not conserved between different diatoms, strongly suggesting that this DIC mobilization strategy likely evolved multiple times with different origins. By contrast, the recent discovery of the thylakoid luminal θ-CA indicates that the strategy to supply CO2 to RubisCO in the pyrenoid may be very similar to that of green algae, and strongly suggests convergent coevolution in CCM function of the thylakoid lumen not only among diatoms but among eukaryotic algae in general. In this review, both experimental and corresponding theoretical models of the diatom CCMs are discussed.This article is part of the themed issue 'The peculiar carbon metabolism in diatoms'.

Molecular aspects of the biophysical CO2-concentrating mechanism and its regulation in marine diatoms.

Diatoms operate a CO2-concentrating mechanism (CCM) that drives upwards of 20% of annual global primary production. Recent progress in CCM research in the marine pennate diatom Phaeodactylum tricornutum revealed that this diatom directly takes up HCO3- from seawater through low-CO2-inducible plasma membrane HCO3- transporters, which belong to the solute carrier (SLC) 4 family. Apart from this, studies of carbonic anhydrases (CAs) in diatoms have revealed considerable diversity in classes and localization among species. This strongly suggests that the CA systems, which control permeability and flux of dissolved inorganic carbon (DIC) by catalysing reversible CO2 hydration, have evolved from diverse origins. Of particular interest is the occurrence of low-CO2-inducible external CAs in the centric marine diatom Thalassiosira pseudonana, offering a strategy of CA-catalysed initial CO2 entry via passive diffusion, contrasting with active DIC transport in P. tricornutum. Molecular mechanisms to transport DIC across chloroplast envelopes are likely also through specific HCO3- transporters, although details have yet to be elucidated. Furthermore, recent discovery of a luminal θ-CA in the diatom thylakoid implied a common strategy in the mechanism to supply CO2 to RubisCO in the pyrenoid, which is conserved among green algae and some heterokontophytes. These results strongly suggest an occurrence of convergent coevolution between the pyrenoid and thylakoid membrane in aquatic photosynthesis.

Noninvasive Assessment of Advanced Fibrosis Based on Hepatic Volume in Patients with Nonalcoholic Fatty Liver Disease.

BACKGROUND/AIMS: Noninvasive liver fibrosis evaluation was performed in patients with nonalcoholic fatty liver disease (NAFLD). We used a quantitative method based on the hepatic volume acquired from gadoxetate disodium-enhanced (Gd-EOB-DTPA-enhanced) magnetic resonance imaging (MRI) for diagnosing advanced fibrosis in patients with NAFLD. METHODS: A total of 130 patients who were diagnosed with NAFLD and underwent Gd-EOB-DTPA-enhanced MRI were retrospectively included. Histological data were available for 118 patients. Hepatic volumetric parameters, including the left hepatic lobe to right hepatic lobe volume ratio (L/R ratio), were measured. The usefulness of the L/R ratio for diagnosing fibrosis ≥F3-4 and F4 was assessed using the area under the receiver operating characteristic (AUROC) curve. Multiple regression analysis was performed to identify variables (age, body mass index, serum fibrosis markers, and histological features) that were associated with the L/R ratio. RESULTS: The L/R ratio demonstrated good performance in differentiating advanced fibrosis (AUROC, 0.80; 95% confidence interval, 0.72 to 0.88) from cirrhosis (AUROC, 0.87; 95% confidence interval, 0.75 to 0.99). Multiple regression analysis showed that only fibrosis was significantly associated with the L/R ratio (coefficient, 0.121; p<0.0001). CONCLUSIONS: The L/R ratio, which is not influenced by pathological parameters other than fibrosis, is useful for diagnosing cirrhosis in patients with NAFLD.

Evolutionarily distinct strategies for the acquisition of inorganic carbon from seawater in marine diatoms.

The acquisition of dissolved inorganic carbon (DIC) in CO2-limited seawater is a central issue to understand in marine primary production. We previously demonstrated the occurrence of direct HCO3- uptake by solute carrier (SLC) 4 transporters in a diatom, a major marine primary producer. Homologs of SLC are found in both centric and pennate marine diatoms, suggesting that SLC transporters are generally conserved. Here, the generality of SLC-mediated DIC uptake in diatoms was examined using an SLC inhibitor, diisothiocyano-2,2'-stilbenedisulfonic acid (DIDS), and an inhibitor of external carbonic anhydrase, acetazolamide. DIDS suppressed high-DIC-affinity photosynthesis in the pennate diatom Phaeodactylum tricornutum and the centric diatom Chaetoceros muelleri, but there was no effect on either the pennate Cylindrotheca fusiformis or the centric Thalassiosira pseudonana. Interestingly, the DIC affinity of DIDS-insensitive strains was sensitive to treatment with up to 100 µM acetazolamide, displaying a 2-4-fold increase in K0.5[DIC]. In contrast, acetazolamide did not affect the DIDS-sensitive group. These results indicate the occurrence of two distinct strategies for DIC uptake-one primarily facilitated by SLC and the other being passive CO2 entry facilitated by external carbonic anhydrase. The phylogenetic independence of these strategies suggests that environmental demands drove the evolution of distinct DIC uptake mechanisms in diatoms.

Hepatic fat quantification using the two-point Dixon method and fat color maps based on non-alcoholic fatty liver disease activity score.

AIM: The two-point Dixon method for magnetic resonance imaging (MRI) is commonly used to non-invasively measure fat deposition in the liver. The aim of the present study was to assess the usefulness of MRI-fat fraction (MRI-FF) using the two-point Dixon method based on the non-alcoholic fatty liver disease activity score. METHODS: This retrospective study included 106 patients who underwent liver MRI and MR spectroscopy, and 201 patients who underwent liver MRI and histological assessment. The relationship between MRI-FF and MR spectroscopy-fat fraction was used to estimate the corrected MRI-FF for hepatic multi-peaks of fat. Then, a color FF map was generated with the corrected MRI-FF based on the non-alcoholic fatty liver disease activity score. We defined FF variability as the standard deviation of FF in regions of interest. Uniformity of hepatic fat was visually graded on a three-point scale using both gray-scale and color FF maps. Confounding effects of histology (iron, inflammation and fibrosis) on corrected MRI-FF were assessed by multiple linear regression. RESULTS: The linear correlations between MRI-FF and MR spectroscopy-fat fraction, and between corrected MRI-FF and histological steatosis were strong (R2 = 0.90 and R2 = 0.88, respectively). Liver fat variability significantly increased with visual fat uniformity grade using both of the maps (ρ = 0.67-0.69, both P < 0.001). Hepatic iron, inflammation and fibrosis had no significant confounding effects on the corrected MRI-FF (all P > 0.05). CONCLUSIONS: The two-point Dixon method and the gray-scale or color FF maps based on the non-alcoholic fatty liver disease activity score were useful for fat quantification in the liver of patients without severe iron deposition.