Arindam Ghatak

Pearl millet genome sequence provides a resource to improve agronomic traits in arid environments

Pearl millet [Cenchrus americanus (L.) Morrone] is a staple food for more than 90 million farmers in arid and semi-arid regions of sub-Saharan Africa, India and South Asia. We report the ∼1.79 Gb draft whole genome sequence of reference genotype Tift 23D2B1-P1-P5, which contains an estimated 38,579 genes. We highlight the substantial enrichment for wax biosynthesis genes, which may contribute to heat and drought tolerance in this crop. We resequenced and analyzed 994 pearl millet lines, enabling insights into population structure, genetic diversity and domestication. We use these resequencing data to establish marker trait associations for genomic selection, to define heterotic pools, and to predict hybrid performance. We believe that these resources should empower researchers and breeders to improve this important staple crop.

Heat-Treatment-Responsive Proteins in Different Developmental Stages of Tomato Pollen Detected by Targeted Mass Accuracy Precursor Alignment (tMAPA)

Recently, we have developed a quantitative shotgun proteomics strategy called mass accuracy precursor alignment (MAPA). The MAPA algorithm uses high mass accuracy to bin mass-to-charge (m/z) ratios of precursor ions from LC-MS analyses, determines their intensities, and extracts a quantitative sample versus m/z ratio data alignment matrix from a multitude of samples. Here, we introduce a novel feature of this algorithm that allows the extraction and alignment of proteotypic peptide precursor ions or any other target peptide from complex shotgun proteomics data for accurate quantification of unique proteins. This strategy circumvents the problem of confusing the quantification of proteins due to indistinguishable protein isoforms by a typical shotgun proteomics approach. We applied this strategy to a comparison of control and heat-treated tomato pollen grains at two developmental stages, post-meiotic and mature. Pollen is a temperature-sensitive tissue involved in the reproductive cycle of plants and plays a major role in fruit setting and yield. By LC-MS-based shotgun proteomics, we identified more than 2000 proteins in total for all different tissues. By applying the targeted MAPA data-processing strategy, 51 unique proteins were identified as heat-treatment-responsive protein candidates. The potential function of the identified candidates in a specific developmental stage is discussed.

Pollen proteomics: from stress physiology to developmental priming

Key messagePollen development and stress.AbstractIn angiosperms, pollen or pollen grain (male gametophyte) is a highly reduced two- or three-cell structure which plays a decisive role in plant reproduction. Male gametophyte development takes place in anther locules where diploid sporophytic cells undergo meiotic division followed by two consecutive mitotic processes. A desiccated and metabolically quiescent form of mature pollen is released from the anther which lands on the stigma. Pollen tube growth takes place followed by double fertilization. Apart from its importance in sexual reproduction, pollen is also an interesting model system which integrates fundamental cellular processes like cell division, differentiation, fate determination, polar establishment, cell to cell recognition and communication. Recently, pollen functionality has been studied by multidisciplinary approaches which also include OMICS analyses like transcriptomics, proteomics and metabolomics. Here, we review recent advances in proteomics of pollen development and propose the process of developmental priming playing a key role to guard highly sensitive developmental processes.

The membrane proteome of male gametophyte in Solanum lycopersicum

UNLABELLED Pollen cells possess specialized cellular compartments separated by membranes. Consequently, mature pollen contains proteinaceous factors for inter- and intracellular transport of metabolites or ions to facilitate the upcoming energy exhausting processes - germination and fertilization. Despite the current advancement in the understanding of pollen development little is known about the role and molecular nature of the membrane proteome that participates in functioning and development of male gametophyte. We dissected the membrane proteome of mature pollen from economically important crop Solanum lycopersicum (tomato). Isolated membrane fractions from mature pollen of two tomato cultivars (cv. Moneymaker and cv. Red setter) were subjected to shotgun proteomics (GEL-LC-Orbitrap-MS). The global tomato protein assignment was achieved by mapping the peptides on reference genome (cv. Heinz 1706) and de novo assembled transcriptome based on mRNA sequencing from the respective cultivar. We identified 687 proteins, where 176 were assigned as putative membrane proteins. About 58% of the identified membrane proteins participate in transport processes. In depth analysis revealed proteins corresponding to energy related pathways (Glycolysis and Krebs cycle) as prerequisite for mature pollen, thereby revealing a reliable model of energy reservoir of the male gametophyte. BIOLOGICAL SIGNIFICANCE Mature pollen plays an indispensable role in plant fertility and crop production. To decipher the functionality of pollen global proteomics studies have been undertaken. However, these datasets are deficient in membrane proteins due to their low abundance and solubility. The work presented here provides a comprehensive investigation of membrane proteome of male gametophyte of an agriculturally important crop plant tomato. The analysis of membrane enriched fractions from two tomato cultivars ensured an effective profiling of the pollen membrane proteome. Particularly proteins of the Krebs cycle or the glycolysis process have been detected and thus a model for the energy dynamics and preparedness of the male gametophyte for the upcoming events - germination and fertilization is provided.

Cereal Crop Proteomics: Systemic Analysis of Crop Drought Stress Responses Towards Marker-Assisted Selection Breeding

Sustainable crop production is the major challenge in the current global climate change scenario. Drought stress is one of the most critical abiotic factors which negatively impact crop productivity. In recent years, knowledge about molecular regulation has been generated to understand drought stress responses. For example, information obtained by transcriptome analysis has enhanced our knowledge and facilitated the identification of candidate genes which can be utilized for plant breeding. On the other hand, it becomes more and more evident that the translational and post-translational machinery plays a major role in stress adaptation, especially for immediate molecular processes during stress adaptation. Therefore, it is essential to measure protein levels and post-translational protein modifications to reveal information about stress inducible signal perception and transduction, translational activity and induced protein levels. This information cannot be revealed by genomic or transcriptomic analysis. Eventually, these processes will provide more direct insight into stress perception then genetic markers and might build a complementary basis for future marker-assisted selection of drought resistance. In this review, we survey the role of proteomic studies to illustrate their applications in crop stress adaptation analysis with respect to productivity. Cereal crops such as wheat, rice, maize, barley, sorghum and pearl millet are discussed in detail. We provide a comprehensive and comparative overview of all detected protein changes involved in drought stress in these crops and have summarized existing knowledge into a proposed scheme of drought response. Based on a recent proteome study of pearl millet under drought stress we compare our findings with wheat proteomes and another recent study which defined genetic marker in pearl millet.

Indian Grape Wines: A Potential Source of Phenols, Polyphenols, and Antioxidants

Indian grape wines are analyzed for total phenolic content and antioxidant activity along with other parameters, such as pH, alcohol content, and reducing sugars. Concentration of polyphenols, like tannic acid, catechol, vanillin, caeffic acid, ferullic acid, and resveratrol, was quantified using reverse phase-high performance liquid chromatography and ultra high performance liquid chromatography. The red wines showed the highest concentration of phenolic content (6.5 ± 0.1 mg/ml) and antioxidant activity (84.60 ± 1%) as compared to white and port wines, while red wine R2 showed the highest radical scavenging activity among red wines and R4 showed the lowest total phenolic content. The white wine W3 showed less total phenolic content and antioxidant activity. Further, a positive correlation between phenolic content and antioxidant activity was observed.

Proteomics survey of Solanaceae family: Current status and challenges ahead

Solanaceae is one of the major economically important families of higher plants and has played a central role in human nutrition since the dawn of human civilization. Therefore, researchers have always been interested in understanding the complex behavior of Solanaceae members to identify key transcripts, proteins or metabolites, which are potentially associated with major traits. Proteomics studies have contributed significantly to understanding the physiology of Solanaceae members. A compilation of all the published reports showed that both gel-based (75%) and gel-free (25%) proteomic technologies have been utilized to establish the proteomes of different tissues, organs, and organelles under normal and adverse environmental conditions. Among the Solanaceae members, most of the research has been focused on tomato (42%) followed by potato (28%) and tobacco (20%), owing to their economic importance. This review comprehensively covers the progress made so far in the field of Solanaceae proteomics including novel methods developed to isolate the proteins from different tissues. Moreover, key proteins presented in this review can serve as a resource to select potential targets for crop improvement. We envisage that information presented in this review would enable us to design the stress tolerant plants with enhanced yields.

Comprehensive tissue-specific proteome analysis of drought stress responses in Pennisetum glaucum (L.) R. Br. (Pearl millet)☆

UNLABELLED Pearl millet is the fifth most important cereal crop worldwide and cultivated especially by small holder farmers in arid and semi-arid regions because of its drought and salt tolerance. The molecular mechanisms of drought stress tolerance in Pennisetum remain elusive. We have used a shotgun proteomics approach to investigate protein signatures from different tissues under drought and control conditions. Drought stressed plants showed significant changes in stomatal conductance and increased root growth compared to the control plants. Root, leaf and seed tissues were harvested and 2281 proteins were identified and quantified in total. Leaf tissue showed the largest number of significant changes (120), followed by roots (25) and seeds (10). Increased levels of root proteins involved in cell wall-, lipid-, secondary- and signaling metabolism and the concomitantly observed increased root length point to an impaired shoot-root communication under drought stress. The harvest index (HI) showed a significant reduction under drought stress. Proteins with a high correlation to the HI were identified using sparse partial least square (sPLS) analysis. Considering the importance of Pearl millet as a stress tolerant food crop, this study provides a first reference data set for future investigations of the underlying molecular mechanisms. BIOLOGICAL SIGNIFICANCE Drought stress is the most limiting factor for plant growth and crop production worldwide. At the same time drought susceptible cereal crops are among the largest producers worldwide. In contrast, Pearl millet is a drought and salt tolerant cereal crop especially used in arid and semi-arid regions by small farmers. The multifactorial molecular mechanisms of this unique drought tolerance are not known. Here, we employ shotgun proteomics for a first characterization of the Pearl millet drought stress proteome. The experimental setup and the data set generated from this study reveal comprehensive physiological and proteomic responses of the drought stressed Pearl millet plants. Our study reveals statistically significant tissue-specific protein signatures during the adaptation to drought conditions. Thus, the work provides a first reference study of the drought stress proteome and related drought responsive proteins (DRPs) in Pearl millet.

Metabolomics in Plant Stress Physiology

Metabolomics is an essential technology for functional genomics and systems biology. It plays a key role in functional annotation of genes and understanding towards cellular and molecular, biotic and abiotic stress responses. Different analytical techniques are used to extend the coverage of a full metabolome. The commonly used techniques are NMR, CE-MS, LC-MS, and GC-MS. The choice of a suitable technique depends on the speed, sensitivity, and accuracy. This chapter provides insight into plant metabolomic techniques, databases used in the analysis, data mining and processing, compound identification, and limitations in metabolomics. It also describes the workflow of measuring metabolites in plants. Metabolomic studies in plant responses to stress are a key research topic in many laboratories worldwide. We summarize different approaches and provide a generic overview of stress responsive metabolite markers and processes compiled from a broad range of different studies. Graphical Abstract.

Pollen Metabolome Dynamics: Biochemistry, Regulation and Analysis

The metabolome of an organism represents the readout of its biochemistry comprising numerous and tightly regulated metabolic pathways. Experimental analysis of the metabolome and its interpretation in a biochemically and physiologically meaningful context is focused by the research field of metabolomics which has become an integral part of many systems biological studies. Pollen development, germination and tube growth comprise numerous steps of metabolic regulation resulting in significant metabolome dynamics. To unravel involved regulatory molecular processes and to promote the understanding of developmental reprogramming and stress tolerance mechanisms in pollen, it is crucial to quantitatively resolve dynamics in the pollen metabolome. Since these dynamics affect various substance groups with different physico-chemical properties, different experimental platforms are needed for robust compound identification and quantification. It has been shown that developmentally and stress-induced metabolic reprogramming in pollen significantly affects the redox homeostasis as well as metabolism of carbohydrates, amino acids, lipids, polyamines, flavonoids and phytohormones. In this chapter, mechanisms of metabolic reprogramming are summarized and discussed in the context of pollen development and stress exposure. Finally, it is discussed how these metabolome dynamics can be resolved methodologically in order to unravel molecular physiological mechanisms of pollen development.