Mitochondrial phosphoenolpyruvate carboxylase contributes to carbon fixation in the diatom Phaeodactylum tricornutum at low inorganic carbon concentrations.

Photosynthetic carbon fixation is often limited by CO2 availability, which led to the evolution of CO2 concentrating mechanisms (CCMs). Some diatoms possess CCMs that employ biochemical fixation of bicarbonate, similar to C4 plants, but whether biochemical CCMs are commonly found in diatoms is a subject of debate. In the diatom Phaeodactylum tricornutum, phosphoenolpyruvate carboxylase (PEPC) is present in two isoforms, PEPC1 in the plastids and PEPC2 in the mitochondria. We used real-time quantitative polymerase chain reaction, Western blots, and enzymatic assays to examine PEPC expression and PEPC activity, under low and high concentrations of dissolved inorganic carbon (DIC). We generated and analyzed individual knockout cell lines of PEPC1 and PEPC2, as well as a PEPC1/2 double-knockout strain. While we could not detect an altered phenotype in the PEPC1 knockout strains at ambient, low or high DIC concentrations, PEPC2 and the double-knockout strains grown under ambient air or lower DIC availability conditions showed reduced growth and photosynthetic affinity for DIC while behaving similarly to wild-type (WT) cells at high DIC concentrations. These mutants furthermore exhibited significantly lower 13 C/12 C ratios compared to the WT. Our data imply that in P. tricornutum at least parts of the CCM rely on biochemical bicarbonate fixation catalyzed by the mitochondrial PEPC2.

Reduced vacuolar ß-1,3-glucan synthesis affects carbohydrate metabolism as well as plastid homeostasis and structure in Phaeodactylum tricornutum.

The ß-1,3-glucan chrysolaminarin is the main storage polysaccharide of diatoms. In contrast to plants and green algae, diatoms and most other algal groups do not accumulate storage polysaccharides in their plastids. The diatom Phaeodactylum tricornutum possesses only a single gene encoding a putative ß-1,3-glucan synthase (PtBGS). Here, we characterize this enzyme by expressing GFP fusion proteins in P. tricornutum and by creating and investigating corresponding gene silencing mutants. We demonstrate that PtBGS is a vacuolar protein located in the tonoplast. Metabolite analyses of two mutant strains with reduced amounts of PtBGS reveal a reduction in their chrysolaminarin content and an increase of soluble sugars and lipids. This indicates that carbohydrates are shunted into alternative pathways when chrysolaminarin production is impaired. The mutant strains show reduced growth and lower photosynthetic capacities, while possessing higher photoprotective abilities than WT cells. Interestingly, a strong reduction in PtBGS expression also results in aberrations of the usually very regular thylakoid membrane patterns, including increased thylakoid thickness, reduced numbers of thylakoids per plastid, and increased numbers of lamellae per thylakoid stack. Our data demonstrate the complex intertwinement of carbohydrate storage in the vacuoles with carbohydrate metabolism, photosynthetic homeostasis, and plastid morphology.

The intracellular distribution of inorganic carbon fixing enzymes does not support the presence of a C4 pathway in the diatom Phaeodactylum tricornutum.

Diatoms are unicellular algae and important primary producers. The process of carbon fixation in diatoms is very efficient even though the availability of dissolved CO2 in sea water is very low. The operation of a carbon concentrating mechanism (CCM) also makes the more abundant bicarbonate accessible for photosynthetic carbon fixation. Diatoms possess carbonic anhydrases as well as metabolic enzymes potentially involved in C4 pathways; however, the question as to whether a C4 pathway plays a general role in diatoms is not yet solved. While genome analyses indicate that the diatom Phaeodactylum tricornutum possesses all the enzymes required to operate a C4 pathway, silencing of the pyruvate orthophosphate dikinase (PPDK) in a genetically transformed cell line does not lead to reduced photosynthetic carbon fixation. In this study, we have determined the intracellular location of all enzymes potentially involved in C4-like carbon fixing pathways in P. tricornutum by expression of the respective proteins fused to green fluorescent protein (GFP), followed by fluorescence microscopy. Furthermore, we compared the results to known pathways and locations of enzymes in higher plants performing C3 or C4 photosynthesis. This approach revealed that the intracellular distribution of the investigated enzymes is quite different from the one observed in higher plants. In particular, the apparent lack of a plastidic decarboxylase in P. tricornutum indicates that this diatom does not perform a C4-like CCM.

Genome editing in diatoms: achievements and goals.

Diatoms are major components of phytoplankton and play a key role in the ecology of aquatic ecosystems. These algae are of great scientific importance for a wide variety of research areas, ranging from marine ecology and oceanography to biotechnology. During the last 20 years, the availability of genomic information on selected diatom species and a substantial progress in genetic manipulation, strongly contributed to establishing diatoms as molecular model organisms for marine biology research. Recently, tailored TALEN endonucleases and the CRISPR/Cas9 system were utilized in diatoms, allowing targeted genetic modifications and the generation of knockout strains. These approaches are extremely valuable for diatom research because breeding, forward genetic screens by random insertion, and chemical mutagenesis are not applicable to the available model species Phaeodactylum tricornutum and Thalassiosira pseudonana, which do not cross sexually in the lab. Here, we provide an overview of the genetic toolbox that is currently available for performing stable genetic modifications in diatoms. We also discuss novel challenges that need to be addressed to fully exploit the potential of these technologies for the characterization of diatom biology and for metabolic engineering.

Diatom Vacuolar 1,6-ß-Transglycosylases can Functionally Complement the Respective Yeast Mutants.

Diatoms are unicellular photoautotrophic algae, which can be found in any aquatic habitat. The main storage carbohydrate of diatoms is chrysolaminarin, a nonlinear ß-glucan, consisting of a linear 1,3-ß-chain with 1,6-ß-branches, which is stored in cytoplasmic vacuoles. The metabolic pathways of chrysolaminarin synthesis in diatoms are poorly investigated, therefore we studied two potential 1,6-ß-transglycosylases (TGS) of the diatom Phaeodactylum tricornutum which are similar to yeast Kre6 proteins and which potentially are involved in the branching of 1,3-ß-glucan chains by adding d-glucose as 1,6-side chains. We genetically fused the full-length diatom TGS proteins to GFP and expressed these constructs in P. tricornutum, demonstrating that the enzymes are apparently located in the vacuoles, which indicates that branching of chrysolaminarin may occur in these organelles. Furthermore, we demonstrated the functionality of the diatom enzymes by expressing TGS1 and 2 proteins in yeast, which resulted in a partial complementation of growth deficiencies of a transglycosylase-deficient ∆kre6 yeast strain.

Synthetic polyester from algae oil.

Current efforts to technically use microalgae focus on the generation of fuels with a molecular structure identical to crude oil based products. Here we suggest a different approach for the utilization of algae by translating the unique molecular structures of algae oil fatty acids into higher value chemical intermediates and materials. A crude extract from a microalga, the diatom Phaeodactylum tricornutum, was obtained as a multicomponent mixture containing amongst others unsaturated fatty acid (16:1, 18:1, and 20:5) phosphocholine triglycerides. Exposure of this crude algae oil to CO and methanol with the known catalyst precursor [{1,2-(tBu2 PCH2)2C6H4}Pd(OTf)](OTf) resulted in isomerization/methoxycarbonylation of the unsaturated fatty acids into a mixture of linear 1,17- and 1,19-diesters in high purity (>99 %). Polycondensation with a mixture of the corresponding diols yielded a novel mixed polyester-17/19.17/19 with an advantageously high melting and crystallization temperature.

Peri-interventional LiMAx test for liver ablation - A feasibility study.

BACKGROUND: Local ablation is an important treatment for liver cancer in cases of cirrhosis. Liver failure may occur after ablation, especially in advanced liver diseases. Currently, there is no standardization for peri-interventional risk assessment. The liver maximum capacity test (LiMAx) is used to assess functional liver capacity, but there is a lack of exploration of its use in this context. OBJECTIVE: The aim of this study was to retrospectively evaluate the usefulness of peri-interventional LiMAx measurements in patients with primary or secondary liver cancer who underwent ablation treatment. METHODS: A LiMAx test was performed at 24 hours pre- and postablation in 49 patients. Blood parameters were collected to determine liver function using MELD and ALBI scores. The results of the LiMAx test were related with these scores and to critical postintervention LiMAx values. RESULTS: LiMAx values correlated strongly with MELD and ALBI scores before the intervention and reflected the change in liver function, as shown by an increase in scores after the intervention. Notably, LiMAx values decreased during the intervention. AUC analysis for patients at risk of reaching a critical liver level after the intervention showed a cutoff value of 186µg/kg/h. CONCLUSIONS: The LiMAx test may be a valuable tool in liver ablation for both peri-interventional monitoring of liver function and preintervention risk assessment.

Performance evaluation of two different software programs for automated ASPECTS scoring in patients with suspected stroke.

BACKGROUND: The range of software available to radiologists has increased enormously with the advancement of AI. A good example of this is software to determine ASPECTS in the treatment of potential stroke patients. OBJECTIVE: In this study, two software packages (eASPECTS from Brainomix and VIA_ASPECTS from Siemens) were tested and compared for their performance in the daily clinical routine of a maximum care provider with a 24/7 stroke unit. METHODS: A total of 637 noncontrast CT images were obtained from consecutive patients with suspected stroke, of whom 73 were finally diagnosed with MCA infarction. Differences in agreement and quantification of agreement were analysed, as well as the correlation and sensitivity, specificity and accuracy compared to raters. RESULTS: Compared to VIA_ASPECTS, eASPECTS shows good agreement and strong correlation with the raters. VIA_ASPECTS has lower accuracy and low specificity than eASPECTS but a higher sensitivity. CONCLUSION: Both software products have the potential to be decision support tools for radiologists. There are, however, differences between the two software products in terms of their intended use.

Using AI and Gd-EOB-DTPA-enhanced MR imaging to assess liver function, comparing the MELIF score with the ALBI score.

Monitoring disease progression is particularly important for determining the optimal treatment strategy in patients with liver disease. Especially for patients with diseases that have a reversible course, there is a lack of suitable tools for monitoring liver function. The development and establishment of such tools is very important, especially in view of the expected increase in such diseases in the future. Image-based liver function parameters, such as the T1 relaxometry-based MELIF score, are ideally suited for this purpose. The determination of this new liver function score is fully automated by software developed with AI technology. In this study, the MELIF score is compared with the widely used ALBI score. The ALBI score was used as a benchmark, as it has been shown to better capture the progression of less severe liver disease than the MELD and Child‒Pugh scores. In this study, we retrospectively determined the ALBI and MELIF scores for 150 patients, compared these scores with the corresponding MELD and Child‒Pugh scores (Pearson correlation), and examined the ability of these scores to discriminate between good and impaired liver function (AUC: MELIF 0.8; ALBI 0.77) and to distinguish between patients with and without cirrhosis (AUC: MELIF 0.83, ALBI 0.79). The MELIF score performed more favourably than the ALBI score and may also be suitable for monitoring mild disease progression. Thus, the MELIF score is promising for closing the gap in the available early-stage liver disease monitoring tools (i.e., identification of liver disease at a potentially reversible stage before chronic liver disease develops).

Volume-Assisted Estimation of Remnant Liver Function Based on Gd-EOB-DTPA Enhanced MR Relaxometry: A Prospective Observational Trial.

In the context of liver surgery, predicting postoperative liver dysfunction is essential. This study explored the potential of preoperative liver function assessment by MRI for predicting postoperative liver dysfunction and compared these results with the established indocyanine green (ICG) clearance test. This prospective study included patients undergoing liver resection with preoperative MRI planning. Liver function was quantified using T1 relaxometry and correlated with established liver function scores. The analysis revealed an improved model for predicting postoperative liver dysfunction, exhibiting an accuracy (ACC) of 0.79, surpassing the 0.70 of the preoperative ICG test, alongside a higher area under the curve (0.75). Notably, the proposed model also successfully predicted all cases of liver failure and showed potential in predicting liver synthesis dysfunction (ACC 0.78). This model showed promise in patient survival rates with a Hazard ratio of 0.87, underscoring its potential as a valuable tool for preoperative evaluation. The findings imply that MRI-based assessment of liver function can provide significant benefits in the early identification and management of patients at risk for postoperative liver dysfunction.

Assessment of Liver Function With MRI: Where Do We Stand?

Liver disease and hepatocellular carcinoma (HCC) have become a global health burden. For this reason, the determination of liver function plays a central role in the monitoring of patients with chronic liver disease or HCC. Furthermore, assessment of liver function is important, e.g., before surgery to prevent liver failure after hepatectomy or to monitor the course of treatment. Liver function and disease severity are usually assessed clinically based on clinical symptoms, biopsy, and blood parameters. These are rather static tests that reflect the current state of the liver without considering changes in liver function. With the development of liver-specific contrast agents for MRI, noninvasive dynamic determination of liver function based on signal intensity or using T1 relaxometry has become possible. The advantage of this imaging modality is that it provides additional information about the vascular structure, anatomy, and heterogeneous distribution of liver function. In this review, we summarized and discussed the results published in recent years on this technique. Indeed, recent data show that the T1 reduction rate seems to be the most appropriate value for determining liver function by MRI. Furthermore, attention has been paid to the development of automated tools for image analysis in order to uncover the steps necessary to obtain a complete process flow from image segmentation to image registration to image analysis. In conclusion, the published data show that liver function values obtained from contrast-enhanced MRI images correlate significantly with the global liver function parameters, making it possible to obtain both functional and anatomic information with a single modality.

MELIF, a Fully Automated Liver Function Score Calculated from Gd-EOB-DTPA-Enhanced MR Images: Diagnostic Performance vs. the MELD Score.

In the management of patients with chronic liver disease, the assessment of liver function is essential for treatment planning. Gd-EOB-DTPA-enhanced MRI allows for both the acquisition of anatomical information and regional liver function quantification. The objective of this study was to demonstrate and evaluate the diagnostic performance of two fully automatically generated imaging-based liver function scores that take the whole liver into account. T1 images from the native and hepatobiliary phases and the corresponding T1 maps from 195 patients were analyzed. A novel artificial-intelligence-based software prototype performed image segmentation and registration, calculated the reduction rate of the T1 relaxation time for the whole liver (rrT1liver) and used it to calculate a personalized liver function score, then generated a unified score-the MELIF score-by combining the liver function score with a patient-specific factor that included weight, height and liver volume. Both scores correlated strongly with the MELD score, which is used as a reference for global liver function. However, MELIF showed a stronger correlation than the rrT1liver score. This study demonstrated that the fully automated determination of total liver function, regionally resolved, using MR liver imaging is feasible, providing the opportunity to use the MELIF score as a diagnostic marker in future prospective studies.

Dynamic perfusion analysis in acute ischemic stroke: A comparative study of two different softwares.

BACKGROUND: In clinical practice, decisions often must be made rapidly; therefore, automated software is useful for diagnostic support. Perfusion computed tomography and follow-up evaluation of perfusion data are valuable tools for selecting the optimal recanalization therapy in patients with acute ischemic stroke. OBJECTIVE: This study aimed to compare commercially available software used to evaluate stroke patients prior to thrombectomy. METHODS: The performance of Olea Sphere (OlS) software vs. CT Neuro Perfusion from Syngo (Sy), as well as the electronic Alberta Stroke Program Early Computed Tomography Score (e-ASPECTS) software vs. an experienced radiologist, were compared using descriptive statistics including significance analysis, Spearman's correlation, and the Bland-Altman agreement analysis. For this purpose, 43 data sets of patients with stroke symptoms related to the middle cerebral artery territory were retrospectively post-processed with both tools and analyzed. RESULTS: The automatic e-ASPECTS showed high agreement with an expert rater assessment of the ASPECTS. Using OlS and Sy, we compared the parameters for the ischemic core (relative cerebral blood flow), Time to maximum (Tmax) for the penumbra, and the relative mismatch between these two values. Overall, both software tools achieved good agreement, and their respective values correlated well with each other. However, OlS predicted significantly smaller infarct core volumes compared with Sy. CONCLUSIONS: Although the absolute values have a certain degree of variation, both software programs have good agreement with each other.

Mechanical thrombectomy in ischemic stroke: Parameters affecting the TICI outcome.

BACKGROUND: Mechanical thrombectomy (MT) is a standard stroke treatment for patients with large vessel occlusions (LVOs). A decisive factor for a successful outcome is, among other things, timely treatment. OBJECTIVE: The objective was to analyze several time points in relation to outcomes and/or surrogate parameters. Furthermore, our data was placed in the context of other clinical trial data. METHODS: We retrospectively evaluated 133 data sets from patients who underwent MT. The correlation of various time periods with parameters, such as the ASPECTS, NIHSS, mRS, and, particularly, the TICI score, was investigated. RESULTS: A correlation was found for both the NIHSS score at discharge and the TICI score with the time periods of arrival and/or start to groin puncture as well as with arrival to the end of the intervention and the duration of the intervention. CONCLUSIONS: This retrospective study is consistent with large randomized clinical trials investigating stroke management and provides data from daily clinical practice.

Prediction of transarterial chemoembolization (TACE) outcome by pre- and postinterventional 13C-methacetin breath test.

BACKGROUND AND OBJECTIVE: Liver function is one of the most important parameters for the outcome of transarterial chemoembolization (TACE). The liver maximum capacity (LiMAx) test is a bedside test that provides a real-time option for liver function testing. The objective of this pilot study was to investigate the suitability of the LiMAX test for predicting the TACE outcome. METHODS: 20 patients with intermediate-stage hepatocellular carcinoma (HCC) received a LiMAx test 24 h pre and post TACE. In addition, laboratory values were collected to determine liver function and model for endstage liver disease (MELD) scores. The success of TACE was assessed 6 weeks post intervention by morphological imaging tests using modified response evaluation criteria in solid tumors (mRECIST). RESULTS: Patients with an objective response (OR = CR + PR) according to mRECIST post TACE had significantly higher values in the pre-interventional LiMAx test than patients with a non-OR (PD or SD) post TACE (r(14) = 0.62, p = 0.01). Higher pre-interventional LiMAx values therefore indicate OR. Patients with a disease control (DC = CR + PR + SD) according to mRECIST post TACE had significantly higher values in the pre-interventional LiMAx test than patients with a non-DC (PD) post TACE (r(14) = 0.65, p = 0.01). Higher pre-interventional LiMAx values therefore indicate DC. The point biserial correlations of LiMAx values pre and post TACE with the outcome OR or DC were descriptively stronger than those of MELD with OR or DC. This suggests that the LiMAx test correlates better with the treatment response than the MELD score. CONCLUSIONS: For the first time, we were able to show in our study that patients who are scheduled for TACE could benefit from a LiMAx test to be able to estimate the benefit of TACE. The higher the pre-interventional LiMAx values, the higher the benefit of TACE. On the other hand, laboratory parameters summarized in the form of the MELD score had significantly less descriptive correlation with the TACE outcome.

Blasticidin-S deaminase, a new selection marker for genetic transformation of the diatom Phaeodactylum tricornutum.

Most genetic transformation protocols for the model diatom Phaeodactylum tricornutum rely on one of two available antibiotics as selection markers: Zeocin (a formulation of phleomycin D1) or nourseothricin. This limits the number of possible consecutive genetic transformations that can be performed. In order to expand the biotechnological possibilities for P. tricornutum, we searched for additional antibiotics and corresponding resistance genes that might be suitable for use with this diatom. Among the three different antibiotics tested in this study, blasticidin-S and tunicamycin turned out to be lethal to wild-type cells at low concentrations, while voriconazole had no detectable effect on P. tricornutum. Testing the respective resistance genes, we found that the blasticidin-S deaminase gene (bsr) effectively conferred resistance against blasticidin-S to P. tricornutum. Furthermore, we could show that expression of bsr did not lead to cross-resistances against Zeocin or nourseothricin, and that genetically transformed cell lines with resistance against Zeocin or nourseothricin were not resistant against blasticidin-S. In a proof of concept, we also successfully generated double resistant (against blasticidin-S and nourseothricin) P. tricornutum cell lines by co-delivering the bsr vector with a vector conferring nourseothricin resistance to wild-type cells.

Mitochondrial Glycolysis in a Major Lineage of Eukaryotes.

The establishment of the mitochondrion is seen as a transformational step in the origin of eukaryotes. With the mitochondrion came bioenergetic freedom to explore novel evolutionary space leading to the eukaryotic radiation known today. The tight integration of the bacterial endosymbiont with its archaeal host was accompanied by a massive endosymbiotic gene transfer resulting in a small mitochondrial genome which is just a ghost of the original incoming bacterial genome. This endosymbiotic gene transfer resulted in the loss of many genes, both from the bacterial symbiont as well the archaeal host. Loss of genes encoding redundant functions resulted in a replacement of the bulk of the host's metabolism for those originating from the endosymbiont. Glycolysis is one such metabolic pathway in which the original archaeal enzymes have been replaced by bacterial enzymes from the endosymbiont. Glycolysis is a major catabolic pathway that provides cellular energy from the breakdown of glucose. The glycolytic pathway of eukaryotes appears to be bacterial in origin, and in well-studied model eukaryotes it takes place in the cytosol. In contrast, here we demonstrate that the latter stages of glycolysis take place in the mitochondria of stramenopiles, a diverse and ecologically important lineage of eukaryotes. Although our work is based on a limited sample of stramenopiles, it leaves open the possibility that the mitochondrial targeting of glycolytic enzymes in stramenopiles might represent the ancestral state for eukaryotes.

Plastoglobules in algae: A comprehensive comparative study of the presence of major structural and functional components in complex plastids.

Plastoglobules (PG) are lipophilic droplets attached to thylakoid membranes in higher plants and green algae and are implicated in prenyl lipid biosynthesis. They might also represent a central hub for integration of plastid signals under stress and therefore the adaptation of the thylakoid membrane under such conditions. In Arabidopsis thaliana, PG contain around 30 specific proteins of which Fibrillins (FBN) and Activity of bc1 complex kinases (ABC1K) represent the majority with respect to both number and protein mass. However, nothing is known about the presence of PG in most algal species, which are responsible for about 50% of global primary production. Therefore, we searched the genomes of publicly available algal genomes for components of PG and the associated functional network in order to predict their presence and potential evolutionary conservation of physiological functions. We could identify homologous sequences for core components of PG, like FBN and ABC1K, in most investigated algal species. Furthermore, proteins at central and interesting positions within the PG functional coexpression network were identified. Phylogenetic sequence analysis revealed diversity within FBN and ABC1K sequences among algal species with complex plastids of the red lineage and large differences compared with green lineage species. Two types of FBN were detected that differ in their isoelectric point which seems to correlate with subcellular localization. Subgroups of FBN were shared between many investigated species and modeling of their 3D-structure implied a conserved structure. FBN and ABC1K are essential structural and functional components of PG. Their occurrence in investigated algal species suggests presence of PG therein and functions in prenyl lipid metabolism and adaptation of the thylakoid membrane that are conserved during evolution.

Rapid induction of GFP expression by the nitrate reductase promoter in the diatom Phaeodactylum tricornutum.

An essential prerequisite for a controlled transgene expression is the choice of a suitable promoter. In the model diatom Phaeodactylum tricornutum, the most commonly used promoters for trans-gene expression are the light dependent lhcf1 promoters (derived from two endogenous genes encoding fucoxanthin chlorophyll a/c binding proteins) and the nitrate dependent nr promoter (derived from the endogenous nitrate reductase gene). In this study, we investigated the time dependent expression of the green fluorescent protein (GFP) reporter under control of the nitrate reductase promoter in independently genetically transformed P. tricornutum cell lines following induction of expression by change of the nitrogen source in the medium via flow cytometry, microscopy and western blotting. In all investigated cell lines, GFP fluorescence started to increase 1 h after change of the medium, the fastest increase rates were observed between 2 and 3 h. Fluorescence continued to increase slightly for up to 7 h even after transfer of the cells to ammonium medium. The subsequent decrease of GFP fluorescence was much slower than the increase, probably due to the stability of GFP. The investigation of several cell lines transformed with nr based constructs revealed that, also in the absence of nitrate, the promoter may show residual activity. Furthermore, we observed a strong variation of gene expression between independent cell lines, emphasising the importance of a thorough characterisation of genetically modified cell lines and their individual expression patterns.

Comprehensive computational analysis of leucine-rich repeat (LRR) proteins encoded in the genome of the diatom Phaeodactylum tricornutum.

We have screened the genome of the marine diatom Phaeodactylum tricornutum for gene models encoding proteins exhibiting leucine-rich repeat (LRR) structures. In order to reveal the functionality of these proteins, their amino acid sequences were scanned for known domains and for homologies to other proteins. Additionally, proteins were categorized into different LRR-families according to the variable sequence part of their LRR. This approach enabled us to group proteins with potentially similar functionality and to classify also LRR proteins where no characterized homologues in other organisms exist. Most interestingly, we were able to indentify several transmembrane LRR-proteins, which are likely to function as receptor-like molecules. However, none of them carry additional domains that are typical for mammalian or plant-like receptors. Thus, the respective signal recognition pathways seem to be substantially different in diatoms. Moreover, P. tricornutum encodes a family of secreted LRR proteins likely to function as adhesion or binding proteins as part of the extracellular matrix. Additionally, intracellular LRR-only proteins were divided into proteins similar to RasGTPase activators, regulators of nuclear transport, and mitotic regulation. Our approach allowed us to draw a detailed picture of the conservation and diversification of LRR proteins in the marine diatom P. tricornutum.