dataesr/scientific-paragraphs-categorization huggingface dataset:
Members of this family use a highly nucleophilic serine residue as the key catalyst. We hypothesize that the GCSxG motif found in plant PAE is equivalent to the GDSX found in PAE from CE12. In addition, the 3D structure of a kiwi carboxylesterase (AeCXE1, PDB code: 2O7R), which is a SGNH protein like CE12 and CE13 family proteins, showed that the serine residues located in the GxSxG sequence covalently link the acyl group  [37]. The presence of the serine residue in the conserved GCSxG motif across plant PAEs is probably the nucleophile in the active site. The NxayDxwQ motif could be another important motif involved in the catalytic site. In the human Notum protein (PDB code: 4UYU_A), Asp340 and Gln343 contribute to the active site complex of this palmitoleate carboxylesterase  [30]. When AtPAEs were structurealigned with 4UYU, the D and Q amino acid residues of the NxayDxwQ motif co-aligned with D340xxQ343 of hNotum (Fig.
5  and Additional file 8). Furthermore, the 3D structure of Sinorhizobium meliloti, Sm23, carbohydrate esterase of family 3 (CE3) belonging, like CE13, to the SGNH superfamily, showed that an asparagine residue (N) contributes to stabilizing the active site geometry and is located close to the aspartic amino acid residue involved in the catalytic triad  [40]. Hence, the asparagine residue found in the NxayDxwQ motif could also contribute to the stability of the active site. The last conserved domain at the C-terminus, the HCQ motif, is highly conserved among plant PAEs. The histidine residue is the only conserved histidine in the multiple sequence alignment performed on plant PAEs in our study. In the hNotum (PDB code: 4UYU) whose 3D structure has been characterized, histidine is another important amino acid of the catalytic triad that acts as the base to activate the catalytic Ser residue  [30].
The tight hydrophobic interaction between the pectic polymer and the cluster of aromatic rings would keep the substrate bound to the enzyme  [42, 43]. Since the AtPAE models adopt an α/β hydrolase fold while PMEs adopt a right-handed parallel β-helix structure  [41, 44] , we could not superimpose these two different structures to check common conserved amino acid residues, including aromatic residues found to be involved in the pectin binding site in plant PMEs. Furthermore, when compared to some other carbohydrate acetylesterases, aromatic residues could also contribute to binding by working as anchor residues of the substrate. This has been shown in a carbohydrate acetylesterase, Sm23 (PDB code: 4TX1), from Sinorhizobium meliloti 1021, (CE3) in which phenylalanine residues are involved in the positioning of incoming substrate  [45].
Similar results have been observed in the crystal structure of a bacterial acetylxylan esterase (Bacillus pumilus, BpAXE, PDB code: 3FVR) which is a carbohydrate esterase of family 7 (CE7), in which tyrosine residues accommodate the substrate  [46]. In addition to the presence of aromatic residues within these conserved motifs, some conserved glycine residues were found, which could be oxyanion hole residues as shown in the carbohydrate acetylesterase Est24 from Sinorhizobium meliloti (PDB code: 5HOE) or in a kiwi (Actinidia eriantha) plant carboxylesterase (AeCXE1): the regions containing glycine residues are located on loops involved in substrate recognition  [17, 37, 46]. It is known that glycine residues may provide the flexibility needed for enzyme active sites to change conformation  [47].
When protein sequence alignment was performed between AtPAEs and AeCXE1 (PDB code: 2O7R), the glycine residues involved in the oxyanion hole of AeCXE1 were structure-aligned with the two glycine residues in the GGgwC motif, indicating a putative similar function of these two glycine residues in the conserved GGgwC motif of Arabidopsis PAEs (Additional file  14). It should also be noted that a leucine, which is mostly conserved in the ClDG motif of plant PAEs, can be replaced by a methionine residue in some monocot PAEs. The CMDG motif and the highly conserved tryptophan found in the GGxWC motif could thus be together a signature of monocot PAEs, differentiating them from the dicot PAEs studied (Additional file 3b).
Plant growth and development In the land plant species studied in our work, almost all families except Physcomitrella, which appears to have one single PAE gene, contained multiple PAE genes. This obviously questions the role of such abundance. In Arabidopsis, data-mining from the eFP Browser database revealed the diversity of AtPAE expression patterns. It is noticeable that the AtPAE10 gene expression profile was not available in eFP Browser database. The expression of AtPAE genes from clade 2 including AtPAE7 and AtPAE8 appeared to be highly regulated during plant growth and development (Fig. 6 ). Some other PAE genes from clade 2, including AtPAE8, were expressed at specific stages (i. e. flower stage 15 and rosette leaf 12) which could suggest a more specific role in the control of the degree of acetylation of pectins (Fig.
6 ). AtPAE11 which belongs to clade 2, was less expressed and mostly induced in the early stages of development (cotyledon and hypocotyl) while AtPAE12 which belongs to clade 1 was more abundant in seeds (Fig. 6 ). In contrast, AtPAE2, another AtPAE from clade 1, seemed to be flower-specific (Fig. 6 ). The diversity of AtPAE gene expression profiles is in line with that observed for other pectin remodeling enzymes and regulators including PMEs, PMEIs, PLLs and PGs  [48, 49]. To confirm these data, a quantification of AtPAE4, AtPAE8 and AtPAE10 Fig. 6  PAE gene expression pattern derived from eFP Browser database during development in A. thaliana. The expression profile of all AtPAEs during plant growth and development was retrieved from the eFP Browser database  [79]. Heatmap generation was performed using GraphPad Prism 7.
The color scale below the heat map indicates expression values; green indicates low gene expression while red indicates high gene expression gene expression was carried out by qRT-PCR with specific primers for each gene (Fig. 7 ). AtPAE4, AtPAE8 and AtPAE10 were expressed in all selected organs but with some variation in expression levels. AtPAE4 and AtPAE8 were more abundant in vegetative tissues, including leaves, roots, stem and hypocotyl compared to AtPAE10. In addition, AtPAE8 was more expressed in siliques. AtPAE4 was more abundant during the different stages of root development than AtPAE8 and AtPAE10. These genes seem to be regulated differentially in the plant organs at the expression level during the plant development. To assess a correlation between the abundance of transcripts (transcriptomic data) and the presence of the proteins (proteomic data), the cell wall proteome database WallProtDB was used.
The WallProtDB database aims at collecting cell wall proteomic experimental data  [50]. When using WallProtDB, AtPAE10 was present in roots, while no peptides were identified for AtPAE4 and AtPAE8. But other AtPAEs, including AtPAE2 and AtPAE5 were present in roots. For the other AtPAEs, the Wall-ProtDB database showed that AtPAE7, AtPAE11 and AtPAE12 were present in both leaves and roots. AtPAE9 was found in etiolated hypocotyls and in the stem at late flowering stage, while AtPAE3 was found only in leaves  [50]. All these data suggest that a number of PAEs play a key role in controlling the degree of acetylation with consequent effects on development.
Abiotic stress In Arabidopsis, the data analysis from eFP Browser databases showed that some specific AtPAE genes are differentially regulated in response to diverse abiotic stress. For instance, in shoots, AtPAE2 (clade 1) was highly induced in response to osmotic stress and to a certain extent to salt stress (Fig. 8a ). In roots, AtPAE4 expression (clade 3) was slightly over-expressed in response to salt stress as was AtPAE8 (Fig. 8b ). All these data indicate that the fine-tuning of the degree of pectin acetylation could be of importance in establishing abiotic stress tolerance mechanisms.
Discussion In our study, using bioinformatic approaches, we explored the diversity of plant PAEs in terms of sequence, gene expression and protein structure. Focusing on Arabidopsis PAEs, we believe that we provide novel elements related to this small gene family. AtPAEs do not cluster in a similar manner in the phylogenetic tree of the plant kingdom PAEs and in the phylogenetic tree of AtPAEs We used 72 plant PAEs from the 611 putative PAEs from the plant kingdom available in different databases. A set of criteria was used to select the PAE genes. The protein sequences without their signal peptides were retrieved to build a phylogenetic tree. The phylogenetic tree clustered them into five clades where one clade clustered two PAEs, from Moss and Spike moss. Another spike moss PAE (SemoePAE7) clustered with PAEs belonging mostly to monocots.
This suggests that the proteins encoded by these two genes are very important for plant growth and development  [21]. The phylogenetic tree of plant kingdom PAEs failed to distinguish specific clusters for dicot and monocot PAEs, which was previously observed in a study where all the PAE genes from some grasses, including maize, Sorghum, Brachypodium, rice, and two dicot species (poplar and Arabidopsis) were used to build the tree  [21]. However, these specific clusters were not observed in another phylogenetic analysis  [22]. The discrepancies could arise Fig. 8  PAE gene expression profile derived from the eFP Browser database, in response to different types of abiotic stress in A. thaliana. The AtPAE gene expression data in response to abiotic stress from leaves a, roots b and to different pathogen infections c were retrieved from the eFP Browser database  [79].
A Heatmap of these data was drawn using GraphPad Prism 7. 0 software (GraphPad Software, Inc. ). The color scale below the heat map indicates expression values; green indicates low gene expression while red indicates high gene expression. (P syr: Pseudomonas syringae; B cin: Botrytis cinerea; H ara: Hyaloperonospora arabidopsis and P inf: Phytophthora infestans) from the number of species used, the number of sequences, or the parameters used to generate the tree. 3D homology modeling of AtPAEs from each clade shows that proteins adopt a common α/β hydrolase fold The 3D homology modeling of AtPAE4, AtPAE8 and AtPAE10, which belong to distinct clades, was performed. We could not find any plant PAE structure in the PDB database, but the 3D structure of a human Notum protein, which has sequence similarity to CE13 enzymes, was available.
Since the 3D structure of a fungal PAE, AacRGAE, is available in the PDB database, we could hypothesize that 3D homology modeling of any AtPAE with that template could be performed. The 3D homology modeling of AtPAE failed when using AacR-GAE while when using structure alignment of different PAEs from CE12, the conserved motifs involved in the active site were different from those found in plant PAEs from the CE13 family, indicating that PAEs from CE12 and CE13 do not adopt exactly the same structure. Our 3D models were performed using the human Notum as a template. Like all SGNH family proteins, its structure contains the catalytic triad S, D, H. The 3D homology models obtained with each selected AtPAE had a typical similar α/β hydrolase fold and showed the conserved S, D and H amino acid residues.
This was elucidated when we structure-aligned all the AtPAEs with the 3D structure of the hNotum protein and other esterases, including the 3D structure of a kiwi carboxylesterase (AeCXE1, PDB code: 2O7R), which showed that the serine residues located in the GxSxG sequence of AeCXE1 covalently linked the acyl group  [37]. The serine residue in the conserved GCSxG motif across plant PAEs is probably the nucleophile donor to the substrate. We identified other motifs in all AtPAE proteins, such as CLDG, PxYH, and RYCDG, containing aromatic residues (W, Y) and charged amino acids (D) that could contribute to the substrate binding pocket. The 3D structure of a tomato PME has been solved and has shown that some aromatic acid residues, including tyrosine, phenylalanine and tryptophan, could interact with the pectic substrate  [41, 57].
The structure-alignment between AtPAEs and plant PMEs could not provide any evidence of the co-localization of important aromatic residues since AtPAEs adopt an α/β hydrolase fold while plant PMEs have a β-helix structure, (data not shown)  [57]. Nevertheless, since these two enzyme families act on pectin, we could assume some common interaction mechanism between the protein and the substrate. In addition, the literature data on carbohydrate acetylesterases, including a bacterial acetylxylan esterase, shows that aromatic residues could also contribute to binding by working as anchor residues of the substrate  [46]. However, this suggestion needs to be formally and experimentally demonstrated. The expression of AtPAEs could relate to a diversification of their function. We used the eFP Browser database to determine the expression pattern of AtPAE genes in various biological conditions.
In addition, the cell wall proteomics database, WallProtDB, was used to identify the abundance of the different AtPAEs in various tissues. AtPAE7, AtPAE8 and AtPAE11 from clade 2 appear to be specifically expressed during plant growth and development. The high transcript level of these genes encoding these proteins, compared to the other AtPAEs indicates that proteins encoded by them are essential in Arabidopsis growth and development. The presence of peptides from AtPAE7 and AtPAE11 in roots and leaves identified in the WallProtDB database is in agreement with the data from eFP Browser and our in-house data. No peptides corresponding to AtPAE8 could be identified in the WallProtDB database but peptides mapping AtPAE9 which belongs to the putative pair of AtPAEs (AtPAE8/AtPAE9) important for plant growth could be identified in leaves, inflorescence stem and hypocotyls.
The tandem duplicated genes AtPAE7 (At4g19410)/ AtPAE8 (At4g19420) and the pair AtPAE7 (At4g19410)/ AtPAE11 (At5g45280) which displays 83% pairwise protein sequence identity, strengthen the hypothesis that these genes are essential to maintain growth and development in Arabidopsis. Our qRT-PCR data showed that AtPAE4 is also broadly expressed during plant growth and development. The quantification of AtPAE5 expression by qRT-PCR revealed that AtPAE5 shared the same expression pattern as AtPAE4 but AtPAE5 is more abundant than AtPAE4 (Additional file 15). But, when using the WallProtDB database, no peptides mapping AtPAE4 was found, while peptides assigned to AtPAE5 were detected in root tissue. The discrepancies between the gene expression pattern and the proteomic data suggest that AtPAE5 which belongs to the tandem duplicated AtPAE4/AtPAE5 genes, seems to be essential for plant development.
The acetate content in the pectic extract of the atpae8 mutant and the double mutant atpae8/atpae9 was 20% and 37% higher, respectively, compared to the wild type. Data retrieved from eFP Browser showed that AtPAE2 (clade 1) and AtPAE4, (clade 3) genes were mainly regulated in response to different abiotic and biotic stresses. The regulation depended on the stress and the tissues studied. The cell wall plays an important role in response to abiotic stress (for a review, see  [57] ), but the effect of cell wall acetylation in this context is far from being understood. It has been shown that levels of O-acetylation of xyloglucan and pectin are reduced in a water deficit environment and ozone stress, respectively  [59, 60]. PAE genes are induced in response to aluminum stress in Medicago truncatula, and in Populus tremuloides grown in increased levels of atmospheric CO 2  [61, 62].
Data resources The 611 putative plant PAE sequences were retrieved from Phytozome (https://phytozome. jgi. doe. gov/pz/portal. html), PlantCAZyme (http://cys. bios. niu. edu/plantcazyme/) and Uniprot (http://www. uniprot. org/) databases. The PAE sequences were chosen using different criteria, including extensive sequence annotation of the PAE protein of some genomes, availability of full-length protein sequence, model species as well as relevant literature on plant PAE gene function. PAE sequence annotation based on only ESTs were not selected. For PAE genes displaying alternative splicing, only one isoform was selected from the UniProt database.
3D homology modeling Because no 3D structure of plant PAEs was available in the Protein Data Bank  [74] , homologous sequences with known structures as templates were identified using LOMETS  [29]. AtPAE8 without its signal peptide was modeled using the human Notum (PDB code: 4UYU_A)  [30]. Other good templates, including 3FVR_A, 1L7A_A, and 1VLQ_A, were selected based on sequence identity, secondary structure comparison, protein family structure and good query coverage. Templates selected as target proteins for alignment of the AtPAE8 sequence were threaded with FUGUE  [75]. AtPAE4 and AtPAE10 were modeled with chain_B of the 4UYU structure instead of chain A based on sequence-structure comparison using FUGUE. The tertiary structure was modeled with Modeller 9v16  [76] , based on the sequence-structure alignment obtained from FUGUE.
AtPAEs expression profile from eFP Browser The expression profile of all AtPAEs was retrieved from the eFP Browser database  [79]. In this database, the expression level of the different genes was quantified using normalization methods for Arabidopsis plant development  [80]  and global stress  [81]. These data were imported into GraphPad prism 7. 0 software (GraphPad Software, Inc. ). A Heat Map was drawn for gene expression data.
The qRT-PCR was carried out with the LightCycler® 480 SYBR Green I Master (Roche, Indianapolis, IN, USA; Cat. No. 04887352001) in 384-well plates in the LightCycler® 480 Real-Time PCR System (Roche). The CT values for each sample (crossing threshold values are the number of PCR cycles required for the accumulated fluorescence signal to cross a threshold above the background) were acquired with the Light-Cycler® 480 software (Roche) using the second derivative maximum method. We used geNorm software to select the references genes. TIP41 and Clathrin displayed the most stable expression during the development of various organs tested  [83]. But TIP41 gene was considered as the best reference gene according to geNorm  [83]. TIP41 was used to calculate the relative expression of target genes, according to the method described by  [84].
Total RNA extraction, reverse transcription, and RT-qPCR The mRNA expression of HMGCR, ApoB, SR-B1, LDLR, CYP7A1 and glyceraldehyde phosphate dehydrogenase (GAPDH) were analysed by RT-qPCR. Detailed protocols for RNA extraction and reverse transcription have been published elsewhere  [18]. The primers and procedure of RT-qPCR are shown in Table  1. RT-qPCR was performed using an ABI Step One RT-PCR thermal cycler (ABI Stepone, NY, USA) in a 10-μL reaction mixture. To quantify the gene transcripts more precisely, the mRNA level of the housekeeping gene GAPDH was measured and used as a quantitative control. Each sample was normalized against the GAPDH mRNA content. Statistical analysis SPSS 18 (SPSS Science Inc. , Chicago, Illinois) and Prism (GraphPad Software, La Jolla, CA, USA) were used for data analysis.
Improvements in many of the variables assessed were demonstrated between baseline and the end of the study, such as blood pressure and anthropometric or body composition variables, which was to be expected in response to the 15% caloric restriction provided by both diets. oxLDL in the cocoa group was significantly lower than in the control group. After adjusting the data for weight and total and LDL cholesterol, it was shown that cocoa consumption significantly affected the change in oxLDL in men but not in women. To explain this difference, the authors refer to a previously reported difference in the antioxidant status between men and women, with men exhibiting poorer antioxidant status (i. e. , men have a higher oxidation status than women), making them more susceptible than women to an antioxidant effect of the flavanols (44).
More specifically, we sought to describe the types of purchases made to prevent undernutrition in children and to understand the effects and changes induced by regular cash transfers in the daily lives of women, the household, and the community, with regard to preventing undernutrition in children. For this reason, the qualitative study nested within the MAM'Out trial focused mainly on the cash group, although data were also collected from women in the control group in order to receive their feedback on the program and any unanticipated effects. Given the asymmetry in data collection between the two arms of the study, and the objectives of the qualitative study, the control and intervention groups were not compared in this study.
Ethics statement Qualitative data were collected between July 2013 and November 2014. The qualitative study was performed within the framework of a larger research study (the MAM'Out research project), which received ethical approval from two separate ethics committees. The study was approved in April 2013 by the Ethical Committee of the University Hospital of Ghent, and in May 2013 by the Burkinabe National Ethical Committee. The study was registered with ClinicalTrials. gov (trial number: NCT01866124) on May 7, 2013. Mothers of children included in the MAM'Out study provided written informed consent prior to the enrollment of their children in the program. The different participants involved in the qualitative data collection also provided oral consent.
All families were followed up for two years. The main outcomes were the cumulative incidence of acute malnutrition (or wasting) and the cost-effectiveness of the program. Clinical outcomes of the trial will be reported elsewhere. Quantitative data were collected quarterly from June 2013 to September 2015, based on a conceptual framework of actions of cash transfers to prevent acute malnutrition in children. Qualitative data were also collected. This mixed methods approach will allow for a more comprehensive understanding of the results and will help assess the validity of the findings  [10]. Comparing qualitative findings between the control and the intervention groups is beyond the scope of this article, so the results will focus on the data collected from the intervention group. The important aspects of the qualitative study are reported according to the COREQ checklist  [11].
Thirdly, cash transfers were hypothesized to increase female empowerment through the women's control over the income and increased decision-making power. This theoretical framework was used to analyze and organize the qualitative data collected. Individual interviews and focus group discussions offered an opportunity to evaluate this hypothesized model of the effects of cash transfers on preventing acute malnutrition in children. For this purpose, a semi-structured questionnaire administered in the local language was used to assess the experiences related to all possible cash pathways. Interview guides were designed based on the conceptual framework of how the transfers work, and were improved by adding new themes as they arose. More details on these questionnaires are provided later in the paper.
Participants In total, 375 people from the villages that received cash transfers participated in the qualitative data collection during the first year of the program and 22 people from the control group. The second year, 549 people from the villages that received cash transfers and 19 people from the control villages were involved (Fig. 2 ). The choice of participants depended on the interviewers, who randomly selected families to interview. Lists of all the mothers from the different villages were obtained and completed with additional columns for husbands, cowives, mother-in-law, etc. Participants were randomly selected by trained interviewers on a weekly basis. Once a participant was interviewed, his or her name was marked accordingly, and the lists were updated for reference for further randomization sessions.