Ribulose Bisphosphate Carboxylase Small Chain Research Articles

Gerbera POE1 (GhPOE1) is involved in leaf senescence in Arabidopsis

Senescence, the final stage of plant development, is a complex and highly controlled physiological process. Senescence occurs in various tissues and organs, and is especially apparent in leaves. Pollen Ole e 1 (POE1) proteins are known to initiate pollen germination, participate in the response to various stresses, and be involved in root development and seed germination. However, their role in plant senescence is not well defined. Using transcriptome data, we isolated a novel protein from Gerbera hybrida (gerbera) which is an ortholog of AT2G34700 (which encodes a Pollen Ole e 1 domain protein in Arabidopsis thaliana) and named GhPOE1. This protein is localized in the cytoplasm and expresses mainly in disk florets, old leaves, and old roots. We transformed GhPOE1 into wild-type Arabidopsis (Col-0) and obtained homozygous transgenic lines OE2, OE3 and OE5. Overexpression of GhPOE1 in these lines led to yellowing rosette leaves with decreased chlorophyll content and photochemical efficiency. Moreover, senescence-associated genes such as SAG12, SAG13, WRKY53, and SIRK (senescence-induced receptor-like kinase) were significantly up-regulated, and the photosynthesis-associated genes CAB2 (chlorophyll a/b-binding 2) and RBCS (ribulose bisphosphate carboxylase small chain) were down-regulated in the transgenic plants. Our results showed that GhPOE1 is a positive regulator of senescence that can promote leaf senescence in Arabidopsis. GhPOE1 is a new candidate gene for genetic engineering to prolong the vase life of cut gerbera flowers.

RNA-Seq Transcriptome Analysis of Potato with Differential Tolerance to Bentazone Herbicide

Potato (Solanum tuberosum), an important food crop worldwide, is threatened by broadleaf weeds. Bentazone is an effective herbicide for controlling weeds; however, as a photosynthesis inhibitor, it can also affect potato plants. Therefore, screening potato seedlings for bentazone resistance and determining the genes involved is essential. Herein, we selected potato varieties with tolerance and sensitivity to bentazone. The photosynthetic rate of sensitive plants was notably affected by bentazone application, whereas the tolerant plants showed a significantly higher photosynthetic rate. We observed 95.7% bentazone degradation within 24 d after application in the tolerant plants. Transcriptome sequencing revealed that the numbers of differentially expressed genes (DEGs) between the tolerant and sensitive potato seedlings were 2703 and 11,024 before and after bentazone application, respectively. Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis revealed that the majority of DEGs were enriched in metabolic pathways, biosynthesis of secondary metals, carbon metabolism, glutathione metabolism, and photosynthesis. Polyphenol oxidase (PPO), flavonoid 3′,5′-methyltransferase-like (AOMT3), ribulose bisphosphate carboxylase small chain C (RBCS-C), and chalcone synthase 2 (CHS2) were identified as candidates contributing to bentazone tolerance. These results provide a theoretical basis for selecting potato stress-resistant resources in the future.

Potential of Plant Proteins Digested In Silico by Gastrointestinal Enzymes as Nutritional Supplement for COVID-19 Patients.

Currently, no specific drug and vaccine are available for the new coronavirus SARS-CoV-2, and nutritional supplementation should be helpful. This study tried to provide reference for protein supplementation. Specifically, in silico method was employed to simulate protein degradation by gastrointestinal enzymes and to produce a large number of active peptides, then, the binding ability of these peptides to SARS-CoV-2 spike protein receptor-binding domain (RBD) was evaluated. The results showed that wheat-derived alpha/beta-gliadin, oat-derived avenin, and ribulose bisphosphate carboxylase small chain of different origin could be good protein source in generating potent binders to SARS-CoV-2 spike RBD. In addition, some high-affinity oligopeptides (such as PISCR, VQVVN, PQQQF, etc.) were identified as potential binders of SARS-CoV-2 spike RBD. In summary, a number of plant proteins could be helpful for COVID-19 patients when supplemented with these proteins, the identified oligopeptides could be used as lead compound to design potential entry inhibitors against SARS-CoV-2.

Modulation in growth, biochemical attributes and proteome profile of rice cultivars under salt stress

One of the major abiotic stresses that affect productivity of rice is salinity. Rice cultivars showed significant genetic variation in response to salt stress. In the present investigation, differential growth pattern and physio-chemical traits-based screening of high yielding rice cultivars of various salt affected areas of India was carried out, and salt-sensitive and salt-tolerant cultivars were identified. Differential responses of antioxidant enzyme activity and tolerance index at maximum level of salt treatment depicted that Jhelum and Vytilla-4 cultivars of rice were sensitive and tolerant to salt stress, respectively. Analysis of growth, morpho-physiological, and biochemical parameters also confirmed the salt-tolerant and salt-sensitive characters of cv. Vytilla-4 and cv. Jhelum, respectively. Nano-LCMS/MS-based proteome profile of these two cultivars was carried out to find out the mechanism lying behind the salt tolerance. A total number of 514 and 770 protein spots were reported in the most salt-tolerant (cv. Vytilla-4) and salt-sensitive (cv. Jhelum) cultivars, respectively. The differentially expressed proteins (DEPs) were found associated with major metabolic pathways including photosynthesis, energy metabolism, amino acid metabolism, nitrogen assimilation and stress and signalling pathways. The changes in the major proteins like Ribulose bisphosphate carboxylase small chain, chlorophyll a-b binding protein, phosphoglycerate kinase, cytochrome c oxidase subunit 5C, glutamine synthetase, glutathione S-transferase, peroxidase, and thioredoxin elucidated the mechanism activated by salt-tolerant cv. Vytilla-4. The transcriptional validation of some of the differentially expressed proteins through real-time quantitative PCR analysis further validated the proteomic results. Outcomes of this work could help in finding out the potential cross-links of different pathways involved in salt-tolerance mechanisms operating in the studied here rice cultivars under salt stress.

ITRAQ-based quantitative proteomic and physiological analysis of the response to N deficiency and the compensation effect in rice

BackgroundThe crop growth compensation effect is a naturally biological phenomenon, and nitrogen (N) is essential for crop growth and development, especially for yield formation. Little is known about the molecular mechanism of N deficiency and N compensation in rice. Thus, the N-sensitive stage of rice was selected to study N deficiency at the tillering stage and N compensation at the young panicle differentiation stage. In this study, a proteome analysis was performed to analyze leaf differentially expressed proteins (DEPs), and to investigate the leaf physiological characteristics and yield under N deficiency and after N compensation.ResultsThe yield per plant presented an equivalent compensatory effect. The net photosynthetic rate, optimal/maximal quantum yield of photosystem II (Fv/Fm), soil and plant analyzer development (SPAD) value, and glutamic pyruvic transaminase (GPT) activity of T1 (N deficiency at the tillering stage, and N compensation at the young panicle differentiation stage) were lower than those of CK (N at different stages of growth by constant distribution) under N deficiency. However, after N compensation, the net photosynthetic rate, Fv/Fm, SPAD value and GPT activity were increased. Using an iTRAQ-based quantitative approach, a total of 1665 credible proteins were identified in the three 4-plex iTRAQ experiments. Bioinformatics analysis indicated that DEPs were enriched in photosynthesis, photosynthesis-antenna proteins, carbon metabolism and carbon fixation in the photosynthetic organism pathways. Moreover, the photosynthesis-responsive proteins of chlorophyll a-b binding protein, ribulose bisphosphate carboxylase small chain and phosphoglycerate kinase were significantly downregulated under N deficiency. After N compensation, chlorophyll a-b binding protein, NADH dehydrogenase subunit 5, NADH dehydrogenase subunit 7, and peroxidase proteins were significantly upregulated in rice leaves.ConclusionThrough physiological and quantitative proteomic analysis, we concluded that a variety of metabolic pathway changes was induced by N deficiency and N compensation. GO and KEGG enrichment analysis revealed that DEPs were significantly associated with photosynthesis pathway-, energy metabolism pathway- and stress resistance-related proteins. The DEPs play an important role in the regulation of N deficiency and the compensation effect in rice.

A geometric characterization of DNA sequence

The DNA graphical representation which firstly arranges each nucleotide to a numerical vector and then concatenates the vectors into a zigzag curve provides a powerful tool for analyzing genetic sequences. Here we introduce two common geometric quantities, area and curvature, to DNA sequence analysis based on the graphical representation method. Then we derive a higher dimensional vector corresponding to a DNA sequence by the permutation of arrangements between the nucleotides and the representation vectors. After obtaining the geometric descriptors of the sequence, we can perform similarity/dissimilarity analysis of multiple sequences simultaneously. Our approach was examined by three real biological data sets: the coding sequences of beta globin gene of animals, ribulose bisphosphate carboxylase small chain gene of plants, and mitochondrial genome sequences of mammals. The results by our computational approach are consistent with those obtained by other existing methods. Moreover, our approach for computing the descriptors has complexity of linear growth in the length of sequence, which deduces that the algorithm is much faster than traditional multiple sequence alignments. These results indicate that our work has provided a powerful tool to accurately and quickly specify the similarity of two DNA sequences.

Comparative Proteomic Analysis of Coregulation of CIPK14 and WHIRLY1/3 Mediated Pale Yellowing of Leaves in Arabidopsis.

Pale yellowing of leaf variegation is observed in the mutant Arabidopsis lines Calcineurin B-Like-Interacting Protein Kinase14 (CIPK14) overexpression (oeCIPK14) and double-knockout WHIRLY1/WHIRLY3 (why1/3). Further, the relative distribution of WHIRLY1 (WHY1) protein between plastids and the nucleus is affected by the phosphorylation of WHY1 by CIPK14. To elucidate the coregulation of CIPK14 and WHIRLY1/WHIRLY3-mediated pale yellowing of leaves, a differential proteomic analysis was conducted between the oeCIPK14 variegated (oeCIPK14-var) line, why1/3 variegated (why1/3-var) line, and wild type (WT). More than 800 protein spots were resolved on each gel, and 67 differentially abundant proteins (DAPs) were identified by matrix-assisted laser desorption ionization-time of flight/time of flight mass spectrometry (MALDI-TOF/TOF-MS). Of these 67 proteins, 34 DAPs were in the oeCIPK14-var line and 33 DAPs were in the why1/3-var line compared to the WT. Five overlapping proteins were differentially expressed in both the oeCIPK14-var and why1/3-var lines: ATP-dependent Clp protease proteolytic subunit-related protein 3 (ClpR3), Ribulose bisphosphate carboxylase large chain (RBCL), Beta-amylase 3 (BAM3), Ribosome-recycling factor (RRF), and Ribulose bisphosphate carboxylase small chain (RBCS). Bioinformatics analysis showed that most of the DAPs are involved in photosynthesis, defense and antioxidation pathways, protein metabolism, amino acid metabolism, energy metabolism, malate biosynthesis, lipid metabolism, and transcription. Thus, in the why1/3-var and oeCIPK14-var lines, there was a decrease in the photosystem parameters, including the content of chlorophyll, the photochemical efficiency of photosystem (PS II) (Fv/Fm), and electron transport rates (ETRs), but there was an increase in non-photochemical quenching (NPQ). Both mutants showed high sensitivity to intense light. Based on the annotation of the DAPs from both why1/3-var and oeCIPK14-var lines, we conclude that the CIPK14 phosphorylation-mediated WHY1 deficiency in plastids is related to the impairment of protein metabolism, leading to chloroplast dysfunction.

Overexpression of DnaJ-Like Chaperone Enhances Carotenoid Synthesis in Chlamydomonas reinhardtii.

Production of functional carotenoids using microalgae may facilitate the commercialization of anti-aging nutritional supplements. The green alga Chlamydomonas reinhardtii uses a non-mevalonate (MEP) pathway for isopentenyl diphosphate (IPP) synthesis. Two enzymes thought to play important roles in this MEP pathway to IPP synthesis are 1-deoxy-D-xylulose 5-phosphate synthase (DXS) and reductase (DXR). DnaJ-like chaperone (Orange protein) is thought to support phytoene synthase, a key enzyme in plant carotenoid synthesis. Genes for Orange (OR), DXS, and DXR were overexpressed via nuclear transformation into C. reinhardtii. CDS of OR, DXS, and DXR were amplified and connected with dual promoters of heat-shock protein 70A and ribulose bisphosphate carboxylase small chain 2. Compared with the parental strain, transformant CrOR#2 produced increased lutein and β-carotene (1.9-fold and 1.7-fold per cell, respectively). Transformant CrDXS#1 produced lutein and β-carotene at lower per-cell abundances than those for the parental strain. CrDXR#2 transformant produced lutein and β-carotene at higher per-cell abundances than their parental counterpart; however, these transformants produced lutein and β-carotene at lower per-medium abundances than their parental counterparts. These results suggest that OR protein supports phytoene synthase in C. reinhardtii and that the phytoene synthesis step is rate-limiting in carotenoid synthesis.

Silencing SlAGL6, a tomato AGAMOUS-LIKE6 lineage gene, generates fused sepal and green petal.

Silencing SlAGL6 in tomato leads to fused sepal and green petal by influencing the expression of A-, B-class genes. AGAMOUS-LIKE6 (AGL6) lineage is an important clade MADS-box transcription factor and plays essential roles in various developmental programs especially in flower meristem and floral organ development. Here, we isolated a tomato AGL6 lineage gene SlAGL6 and successfully obtained several RNA interference (RNAi) lines. Silencing SlAGL6 led to abnormal fused sepals and light green petals with smaller size. The total chlorophyll content in transgenic petals increased and the morphology of epidermis cells altered. Further analysis showed that A-class gene MACROCALYX (MC) participating in sepal development and a NAC-domain gene GOBLET involving in boundary establishment were down-regulated in transgenic lines. In transgenic petals, two chlorophyll synthesis genes, Golden2-like1 (SlGLK1) and Golden2-like2 (SlGLK2), two photosystem-related genes, ribulose bisphosphate carboxylase small chain 3B (SlrbcS3B) and chlorophyll a/b-binding protein 7 (SlCab-7) were induced and three B-class genes TM6, TAP3 and SlGLO1 were repressed. These results suggest that SlAGL6 involves in tomato sepal and petal development.

RNA-seq assembler artifacts can bias expression counts and differential expression analysis - case study on the chickpea transcriptome emphasizes importance of freely accessible data for reproducibility

The unprecedented volume of genomic and transcriptomic data analyzed by software pipelines makes verification of inferences based on such data, albeit theoretically possible, a challenging proposition. The availability of intermediate data can immensely aid re-validation efforts. One such example is the transcriptome, assembled from raw RNA-seq reads, which is frequently used for annotation and quantification of genes transcribed. The quality of the assembled transcripts influences the accuracy of inferences based on them. Here the publicly available transcriptome from Cicer arietinum (ICC4958; Desi chickpea, http://www.nipgr.res.in/ctdb.html)1 was analyzed using YeATS2. This revealed that a majority of the highly expressed transcripts (HET) encoded multiple genes, strongly indicating that the counts may have been biased by the merging of different transcripts. TC00004 is ranked in the top five HET for all five tissues analyzed here, and encodes both a retinoblastoma-binding-like protein (E-value=0) and a senescence-associated protein (E-value= 5e-108). Fragmented transcripts are another source of error. The ribulose bisphosphate carboxylase small chain (RBCSC) protein is split into two transcripts with an overlapping amino acid sequence "ASNGGRVHC", TC13991 and TC23009, with length 201 and 332 nucleotides and expression counts 17.90 and 1403.8, respectively. The huge difference in counts indicates an erroneous normalization algorithm in determining counts. It is well known that RBCSC is highly expressed and expectedly TC23009 ranks fifth among HETs in the shoot. Furthermore, some transcripts are split into open reading frames that map to the same protein, although this should not have any significant bearing on the counts. It is proposed that studies analyzing differential expression based on the transcriptome should consider these artifacts, and providing intermediate assembled transcriptomes should be mandatory, possibly with a link to the raw sequence data (Bioproject).

RNA-seq assembler artifacts can bias expression counts and differential expression analysis - application of YeATS on the chickpea transcriptome

Background: The unprecedented volume of genomic and transcriptomic data analyzed by software pipelines makes verification of inferences based on such data, albeit theoretically possible, a challenging proposition. The availability of intermediate data can immensely aid re-validation efforts. One such example is the transcriptome, assembled from raw RNA-seq reads, which is frequently used for annotation and quantification of genes transcribed. The quality of the assembled transcripts influences the accuracy of inferences based on them. Method: Here the publicly available transcriptome from Cicer arietinum (ICC4958; Desi chickpea, http://www.nipgr.res.in/ctdb.html) was analyzed using YeATS. Results and Conclusion: The analysis revealed that a majority of the highly expressed transcripts (HET) encoded multiple genes, strongly indicating that the counts may have been biased by the merging of different transcripts. TC00004 is ranked in the top five HET for all five tissues analyzed here, and encodes both a retinoblastoma-binding-like protein (E-value=0) and a senescence-associated protein (E-value= 5e-108). Fragmented transcripts are another source of error. The ribulose bisphosphate carboxylase small chain (RBCSC) protein is split into two transcripts with an overlapping amino acid sequence ”ASNGGRVHC”, TC13991 and TC23009, with length 201 and 332 nucleotides and expression counts 17.90 and 1403.8, respectively. The huge difference in counts indicates an erroneous normalization algorithm in determining counts. It is well known that RBCSC is highly expressed and expectedly TC23009 ranks fifth among HETs in the shoot. Furthermore, some transcripts are split into open reading frames that map to the same protein, although this should not have any significant bearing on the counts. It is proposed that studies analyzing differential expression based on the transcriptome should consider these artifacts, and providing intermediate assembled transcriptomes should be mandatory, possibly with a link to the raw sequence data (Bioproject).

An expressed sequence tags analysis for leaves of Chinese milk vetch (Astragalus sinicus)

Chinese milk vetch (Astragalus sinicus) is one of the most important green manure crops in temperate zone countries. To identify the genes involved in plant growth, a cDNA library was constructed from the mature leaves of Chinese milk vetch, and expressed sequence tag analysis was performed. Sequencing of the cDNA clones generated 1,027 high-quality expressed sequence tags clustered total 693 unigenes which included 110 Contigs and 583 Singlets. A total of 198 unigenes were most similar to Medicago truncatula genes. Genes related to photosynthesis were highly expressed in our library. The most abundant gene which had 42 clones was annotated as Ribulose bisphosphate carboxylase small chain with the function of carbon utilization by fixation of carbon dioxide. The results suggested that this high-quality library could be a good resource for understanding molecular events of Chinese milk vetch leaf growth, and the full-length clones could be used for crop improvement studies in the future.

Identification of Imperative Enzymes by Differential Protein Expression in Picrorhiza kurroa Under Metabolite Accumulating and Non-accumulating Conditions

Picrorhiza kurroa, an endangered medicinal plant found in the North-Western Himalayan region has a number of medicinal properties due to the presence of metabolites picroside-I and picroside-II. It is used in various herbal formulations like Picroliv, Livokin, Picrolax, Livomap, Tefroliv etc. Review of literature revealed that no information is available as of today on the proteome analysis of Picrorhiza kurroa. Hence, we aim to analyse the difference in proteome of Picrorhiza kurroa in response to ~ 17 times higher content of picroside-I at 15°C as compared to its content at 25°C. Thus, differential protein expression was studied. Densitometry analysis of SDS-PAGE gels of samples under two differential conditions of temperature revealed the presence of distinct set of proteins under picroside-I accumulating (15°C) versus non-accumulating (25°C) conditions. Mass spectrometric analysis of these proteins using MALDI-TOF MS followed by protein identification using database search on MASCOT search engine gave interesting results. The significant proteins identified were NAD(P)H-quinone oxidoreductase subunit K, shikimate kinase, ribulose bisphosphate carboxylase small chain and fructokinase fragment. Hence, these findings throw light on the involvement of these enzymes in the crucial physiological processes of Picrorhiza kurroa and can provide an insight into the biosynthesis of picrosides and other secondary metabolites.

Proteomic study of β-aminobutyric acid-mediated cadmium stress alleviation in soybean

The present study highlights the protective role of β-aminobutyric acid (BABA) in alleviating cadmium (Cd) stress in soybean. Proteomic analyses revealed that out of 66 differentially abundant protein spots in response to Cd challenge, 17 were common in the leaves of BABA-primed and non-primed plants. Oxygen-evolving enhancer protein 1 and ribulose bisphosphate carboxylase small chain 1 were detected in increase abundance in both groups of leaves. Among the 15 commonly decreased protein spots, the relative intensity levels of heat shock cognate 70-kDa protein, carbonic anhydrase, methionine synthase, and glycine dehydrogenase were partially restored after BABA treatment. Moreover, BABA priming significantly enhanced the abundance of the defense-related protein peroxiredoxin and glycolytic enzymes in response to Cd exposure. Additionally, the impact of Cd on the physiological state of BABA-primed and non-primed plants was analyzed using a biophoton technique. The finding of comparatively low biophoton emission in BABA-primed leaves under Cd stress indicates that these plants experienced less oxidative damage than that of non-primed plants. Proteomic study coupled with biophoton analysis reveals that BABA pretreatment helps the plants to combat Cd stress by modulating plants' defence mechanism as well as activating cellular detoxification system to protect the cells from Cd induced oxidative stress damages.

Analysis of Differential Expression of Proteins in Rice Leaf Sheath During Grain Filling

To reveal the molecular mechanism of metabolism in rice ( Oryza sativa L.) leaf sheath, differential expression of proteome in rice leaf sheath was investigated with a large panicles and high grain-setting rate cultivar, i.e., Jinhui 809. Through the method of differential proteomics, 23 proteins were detected, and 11 proteins were identified to be the functional proteins. These functional proteins were classified into 6 categories according to their dynamic patterns of expression during grain-filling stage. The first class, such as oxaloacetate decarboxylase alpha subunit, showed down-regulation during grain filling. The second class, which included ribulose bisphosphate carboxylase small chain and ADP-ribosylation factor 1, exhibited up-regulation at first and then down-regulation. The third class, such as auxin response factor, zinc finger, C3HC4 type family protein, vacuolar proton-ATPase, and ribulose bisphosphate carboxylase/oxygenase activase, was down-regulation first and then up-regulation and down-regulation alternatively. The fourth class included Rubisco binding-protein alpha subunit, showing an expression pattern of up and down-regulation alternatively twice during grain filling. The fifth class performed a decreasing at first and then gradually increasing expression pattern. This class included class II metallothionein-like protein 1A and geranylgeranyl diphosphate synthase. The sixth class was protein kinase family protein performing a gradually increasing expression pattern. These 6 classes of proteins were involved in photosynthesis, hormone regulation, substance transportation, resistant response to plant senescence, and cell signal transduction in leaf sheath, respectively. They regulated the transition of sink to source together.