PfSRPK1 Regulates Asexual Blood Stage Schizogony and Is Essential for Male Gamete Formation.

Serine/arginine-rich protein kinases (SRPKs) are cell cycle-regulated serine/threonine protein kinases and are important regulators of splicing factors. In this study, we functionally characterize SRPK1 of the human malaria parasite Plasmodium falciparum. P. falciparum SRPK1 (PfSRPK1) was expressed in asexual blood-stage and sexual-stage gametocytes. Pfsrpk1- parasites formed asexual schizonts that generated far fewer merozoites than wild-type parasites, causing reduced replication rates. Pfsrpk1- parasites also showed a severe defect in the differentiation of male gametes, causing a complete block in parasite transmission to mosquitoes. RNA sequencing (RNA-seq) analysis of wild-type PfNF54 and Pfsrpk1- stage V gametocytes suggested a role for PfSRPK1 in regulating transcript splicing and transcript abundance of genes coding for (i) microtubule/cilium morphogenesis-related proteins, (ii) proteins involved in cyclic nucleotide metabolic processes, (iii) proteins involved in signaling such as PfMAP2, (iv) lipid metabolism enzymes, (v) proteins of osmophilic bodies, and (vi) crystalloid components. Our study reveals an essential role for PfSRPK1 in parasite cell morphogenesis and suggests this kinase as a target to prevent malaria transmission from humans to mosquitoes. IMPORTANCE Plasmodium sexual stages represent a critical bottleneck in the parasite life cycle. Gametocytes taken up in an infectious blood meal by female anopheline mosquito get activated to form gametes and fuse to form short-lived zygotes, which transform into ookinetes to infect mosquitoes. In the present study, we demonstrate that PfSRPK1 is important for merozoite formation and critical for male gametogenesis and is involved in transcript homeostasis for numerous parasite genes. Targeting PfSRPK1 and its downstream pathways may reduce parasite replication and help achieve effective malaria transmission-blocking strategies.

PfARID Regulates P. falciparum Malaria Parasite Male Gametogenesis and Female Fertility and Is Critical for Parasite Transmission to the Mosquito Vector.

Sexual reproduction of Plasmodium falciparum parasites is critical to the spread of malaria in the human population. The factors that regulate gene expression underlying formation of fertilization-competent gametes, however, remain unknown. Here, we report that P. falciparum expresses a protein with an AT-rich interaction domain (ARID) which, in other organisms, is part of chromatin remodeling complexes. P. falciparum ARID (PfARID) localized to the parasite nucleus and is critical for the formation of male gametes and fertility of female gametes. PfARID gene deletion (Pfarid-) gametocytes showed downregulation of gene expression important for gametogenesis, antigenic variation, and cell signaling and for parasite development in the mosquito. Our study identifies PfARID as a critical nuclear protein involved in regulating the gene expression landscape of mature gametocytes. This establishes fertility and also prepares the parasite for postfertilization events that are essential for infection of the mosquito vector. IMPORTANCE Successful completion of the Plasmodium life cycle requires formation of mature gametocytes and their uptake by the female Anopheles mosquito vector in an infected blood meal. Inside the mosquito midgut the parasite undergoes gametogenesis and sexual reproduction. In the present study, we demonstrate that PfARID is essential for male gametogenesis and female fertility and, thereby, transmission to the mosquito vector. PfARID possibly regulates the chromatin landscape of stage V gametocytes and targeting PfARID function may provide new avenues into designing interventions to prevent malaria transmission.

Comparative transcriptomics of leaves of five mulberry accessions and cataloguing structural and expression variants for future prospects.

Bombyx mori, a monophagous insect, prefers leaves of the certain species of Morus more than others. The preference has been attributed to morphological and anatomical features and biochemical compounds. In the present manuscript a comparison has been made among the transcriptome of leaves of the two preferred cultivated varieties and three wild types species. While assembling, high quality transcriptomes of five genotypes were constructed with a total of 100930, 151245, 89724, 181761 and 102908 transcripts from ML, MN, MS, K2 and V1 samples respectively. Further, to compare them, orthologs were identified from these assembled transcriptome. A total of 22462, 23413, 23685, 24371, 18362, 22326, 20058, 18049, 17567 and 20518 clusters of orthologs were found in one to one comparison in KvsN, KvsL, KvsS, KvsV, LvsN, LvsS, LvsV, NvsS, NvsV, and SvsV respectively. 4236 orthologs with algebraic connectivity of 1.0 were then used to compare and to find out differentially expressed transcripts from all the genotypes. A total of 1037 transcripts expressed that include some of the important morphology, anatomy and biochemical pathways regulating transcription factors (AP2/ERFs and C2H2 Zinc fingers) and signalling components were identified to express differentially. Further, these transcriptomes were used find out markers (SSR) and variants and a total of 1101013, 537245, 970877, 310437, 675772, 338400, 581189, 751477, 514999 and 257107 variants including SNP, MNP, Insertions and deletions were found in one to one comparisons. Taken together, our data could be highly useful for mulberry community worldwide as it could be utilized in mulberry breeding programs.

Updated inventory, evolutionary and expression analyses of G (PDR) type ABC transporter genes of rice.

ABC transporters constitute the largest family of transporter proteins in living organisms and divided into eight subfamilies, from A-H. ABCG members, specific to plants and fungi, belong to subfamily G. In this study, we provide updated inventory, detailed account of phylogeny, gene structure characteristics, and expression profiling during reproductive development, abiotic and biotic stresses of members of ABCG gene family in rice along with reannotation and cloning of FL-cDNA of OsABCG50/PDR23. We observed that of the 22 ABCGs/PDRs, four genes evolved as a result of gene duplication events and their expression pattern changed after duplication. Analysis of expression revealed seed and developmental stage preferential expression of five ABCG/PDR members. Transcript levels of eight ABCGs/PDRs were affected by abiotic and biotic stresses. Expression of seven ABCG/PDR genes was also altered by hormonal elicitors. The modulated expression is nicely correlated with the presence of tissue/stress specific cis-acting elements present in putative promoter region.

Genome-wide identification and expression analysis of Hsp70, Hsp90, and Hsp100 heat shock protein genes in barley under stress conditions and reproductive development.

Abiotic stress including extreme temperature disturbs the plant cellular homeostasis consequently limiting the yield potential of crop plants. Heat shock proteins (Hsps) are part of major rescue machinery of plants which aid to combat these stressed conditions by re-establishing protein homeostasis. Hsps with their chaperone and co-chaperone mechanisms regulate the activity of their substrate proteins in an ATP-dependent manner. In the present investigation, a genome-wide identification, evolutionary relationship, and comprehensive expression analysis of Hsp70, Hsp90, and Hsp100 gene families have been done in barley. The barley genome possesses 13 members of the Hsp70 gene family, along with 4 members of the Hsp110 subfamily, and 6 members of Hsp90 and 8 members of the Hsp100 gene family. Hsp genes are distributed on all 7 chromosomes of barley, and their encoded protein members are predicted to be localized to cell organelles such as cytosol, mitochondria, chloroplast, and ER. Despite a larger genome size, there are lesser members of these Hsp genes in barley, owing to less duplication events. The variable expression pattern obtained for genes encoding proteins localized to the same subcellular compartment suggests their diverse roles and involvement in different cellular responses. Expression profiling of these genes was performed by qRT-PCR in an array of 32 tissues, which showed a differential and tissue-specific expression of various members of Hsp gene families. We found the upregulation of HvHspc70-4, HvHsp70Mt70-2, HvHspc70-5a, HvHspc70-5b, HvHspc70-N1, HvHspc70-N2, HvHsp110-3, HvHsp90-1, HvHsp100-1, and HvHsp100-2 upon exposure to heat stress during reproductive development. Furthermore, their higher expression during heat stress, heavy metal stress, drought, and salinity stress was also observed in a tissue-specific manner.

Genome-wide Analysis of bZIP Transcription Factors in wheat and Functional Characterization of a TabZIP under Abiotic Stress.

The basic leucine zipper (bZIP) represents one of the largest as well as most diverse transcription factor (TFs) families. They are known to play role in both stress as well as in various plant developmental processes. In the present study, a total of 191 bZIP transcription factors have been identified from Triticum aestivum. Expression analysis during various stress conditions, developmental stages, different varieties and gene ontology enrichment analysis suggest their possible roles in abiotic stress as well as in developmental responses. In the current analysis, one of the members named as TabZIP (Traes_7AL_25850F96F.1) was selected for detailed analysis to understand its role under different abiotic stress conditions. Gene expression studies revealed differential expression of TabZIP in various abiotic stress conditions like heat, salinity and dehydration suggesting the possible role of bZIP in various stress mitigation mechanism. Arabidopsis transgenics overexpressing TabZIP showed enhanced tolerance to salinity, drought, heat and oxidative stress. Thus TabZIP (Traes_7AL_25850F96F.1) can serve as a candidate gene for improving heat as well as other abiotic stress tolerance and can be helpful in enhancing the crop productivity under stress conditions.

miRNomes involved in imparting thermotolerance to crop plants.

Thermal stress is one of the challenges to crop plants that negatively impacts crop yield. To overcome this ever-growing problem, utilization of regulatory mechanisms, especially microRNAs (miRNAs), that provide efficient and precise regulation in a targeted manner have been found to play determining roles. Besides their roles in plant growth and development, many recent studies have shown differential regulation of several miRNAs during abiotic stresses including heat stress (HS). Thus, understanding the underlying mechanism of miRNA-mediated gene expression during HS will enable researchers to exploit this regulatory mechanism to address HS responses. This review focuses on the miRNAs and regulatory networks that were involved in physiological, metabolic and morphological adaptations during HS in plant, specifically in crops. Illustrated examples including, the miR156-SPL, miR169-NF-YA5, miR395-APS/AST, miR396-WRKY, etc., have been discussed in specific relation to the crop plants. Further, we have also discussed the available plant miRNA databases and bioinformatics tools useful for miRNA identification and study of their regulatory role in response to HS. Finally, we have briefly discussed the future prospects about the miRNA-related mechanisms of HS for improving thermotolerance in crop plants.

Identification of Triticum aestivum nsLTPs and functional validation of two members in development and stress mitigation roles.

Role of plant nsLTP in biotic stress is well reported; however, their role during abiotic stress is far from clear. This study comprises genome-wide identification of LTPs and characterizes the regulation and function of two Triticum aestivum lipid transfer proteins, TaLTP40 and TaLTP75, under stresses that influence membrane fluidity. A total of 105 LTP gene family members have been identified. The selected LTPs for functional validation were highly expressed during salt, cold and drought stress. Further, selected LTPs showed differential expression thermotolerant and thermosusceptible wheat cultivars. Higher expression of many TaLTPs was observed under different abiotic stresses in thermotolerant wheat cultivars as compared to thermosusceptible cultivars. TaLTPs regulation was correlated with light energy distribution studies under similar stress conditions. Cellular localization revealed localization of different TaLTPs to the tonoplast membrane along with the organelles involved in the secretory pathway. Induction of TaLTPs was observed upon treatment with dimethylsulphoxide. TaLTP40 and TaLTP75 overexpressing transgenic Arabidopsis showed a constitutively enhanced salt tolerance. Both the TaLTP40 and TaLTP75 overexpressing lines performed better in terms of chlorophyll a fluorescence, total chlorophyll content, membrane injury index, total biomass, percentage germination, percentage survival and relative growth rate. Hence, our analyses indicate that TaLTPs expression might be driven by change in membrane fluidity and could be involved in transferring membrane lipids to the biological membranes thus imparting tolerance to various abiotic stresses.

Auxin Response Factor Genes Repertoire in Mulberry: Identification, and Structural, Functional and Evolutionary Analyses.

Auxin Response Factors (ARFs) are at the core of the regulation mechanism for auxin-mediated responses, along with AUX/IAA proteins.They are critical in the auxin-mediated control of various biological responses including development and stress. A wild mulberry species genome has been sequenced and offers an opportunity to investigate this important gene family. A total of 17 ARFs have been identified from mulberry (Morus notabilis) which show a wide range of expression patterns. Of these 17 ARFs, 15 have strong acidic isoelectric point (pI) values and a molecular mass ranging from 52 kDa to 101 kDa. The putative promoters of these ARFs harbour cis motifs related to light-dependent responses, various stress responses and hormone regulations suggestive of their multifactorial regulation. The gene ontology terms for ARFs indicate their role in flower development, stress, root morphology and other such development and stress mitigation related activities. Conserved motif analysis showed the presence of all typical domains in all but four members that lack the PB1 domain and thus represent truncated ARFs. Expression analysis of these ARFs suggests their preferential expression in tissues ranging from leaf, root, winter bud, bark and male flowers. These ARFs showed differential expression in the leaf tissue of M. notabilis, Morus laevigata and Morus serrata. Insights gained from this analysis have implications in mulberry improvement programs.

Major intrinsic proteins repertoire of Morus notabilis and their expression profiles in different species.

Leaf moisture content in Morus is a significant trait regulating the yield of silk production. Studies have shown that fresh leaves or leaves with high water content are preferably eaten by silk worm. Water and certain other molecules transport in plants is known to be regulated by aquaporins or Major Intrinsic Proteins (MIPs). Members of the MIP gene family have also been implicated in plant development and stress responsiveness. To understand how members of MIP gene family are regulated and evolved, we carried out an extensive analysis of the gene family. We identified a total of 36 non redundant MIPs in Morus notabilis genome, belonging to five subfamilies PIPs, TIPs, NIPs, XIPs and SIPs) have been identified. We performed a Gene ontology (GO) term enrichment analysis and looked at distribution of cis elements in their 2K upstream regulatory region to reveal their putative roles in various stresses and developmental aspects. Expression analysis in developmental stages revealed their tissue preferential expression pattern in diverse vegetative and reproductive tissues. Comparison of expression profiles in the leaves of three species including Morus notabilis, Morus serrata and Morus laevigata led to identification of differential expression in these species. In all, this study elaborates a basic insight into the structure, function and evolutionary analysis of MIP gene family in Morus which is hitherto unavailable. Our analysis will provide a ready reference to the mulberry research community involved in the Morus improvement program.

Identification and Expression Profiling of the Lectin Gene Superfamily in Mulberry.

Lectins are a diverse group of ubiquitously present, highly specific sugar-binding proteins. Members of this large gene family have been assigned broad biological functions from defense to acting as storage proteins. Despite possessing several interesting characteristics, their functions remain essentially undefined. Mulberry ( spp.) known for its medicinal benefits is also a rich source of lectins. Using an exhaustive hidden Markov model (HMM)-based search, we identified the lectin gene complement in C.K. Schneid with around 197 members. These putative lectin genes were classified into 12 distinct gene families based on the presence of characteristic sugar-binding domains. Members of this superfamily were assigned varied gene ontologies (GOs) to identify putative functions and determine cellular localizations. Interestingly, characteristic expression patterns were observed across the lectin superfamily in response to a variety of environmental cues. This is suggestive of specialized functions under diverse conditions possibly by linking the specificity of sugar recognition with mediating precise stress responses in plants. The identification of putative gene family members from the genus developed in this study can find wide applicability in lectin gene identification and characterization. It can also contribute immensely in the understanding of lectins from mulberry with potential medicinal uses.

Genome-wide Identification and Structural, Functional and Evolutionary Analysis of WRKY Components of Mulberry.

Mulberry is known to be sensitive to several biotic and abiotic stresses, which in turn have a direct impact on the yield of silk, because it is the sole food source for the silk worm. WRKYs are a family of transcription factors, which play an important role in combating various biotic and abiotic stresses. In this study, we identified 54 genes with conserved WRKY motifs in the Morus notabilis genome. Motif searches coupled with a phylogenetic analysis revealed seven sub-groups as well as the absence of members of Group Ib in mulberry. Analyses of the 2K upstream region in addition to a gene ontology terms enrichment analysis revealed putative functions of mulberry WRKYs under biotic and abiotic stresses. An RNA-seq-based analysis showed that several of the identified WRKYs have shown preferential expression in the leaf, bark, root, male flower, and winter bud of M. notabilis. Finally, expression analysis by qPCR under different stress and hormone treatments revealed genotype-specific responses. Taken together, our results briefs about the genome-wide identification of WRKYs as well as their differential response to stresses and hormones. Importantly, these data can also be utilized to identify potential molecular targets for conferring tolerance to various stresses in mulberry.

Genome-wide analysis, expression dynamics and varietal comparison of NAC gene family at various developmental stages in Morus notabilis.

NAC genes are important transcription factors and forms a large family in plants. They have shown to play an important role in growth and development and have also been shown to involve in regulation of stress-responsive genes. In the present study, a repertoire of NAC genes in recently published mulberry genome has been identified which consists of a total of 79 members. Structural analysis revealed that most of the NAC genes in mulberry contain two introns. The proteins encoded by them show a wide range of isoelectric points suggestive of their varied roles in varying microcellular environment. Phylogenetic and conserved motif analysis elucidate the presence of 15 sub-groups of these genes along with two novel sub-groups having distinct conserved motifs which are not present in Arabidopsis. Gene ontology term enrichment analysis and cis-element identification from their putative 1 K upstream regulatory region indicates their possible role in important biological processes like organ formation, meristem establishment, senescence, and various biotic and abiotic stresses. Expression analysis across various developmental stages led to identification of their preferential expression in diverse tissues. Taken together, this work provides a solid background information related to structure, function, expression and evolution of NAC gene family in mulberry.

Comparative transcriptomics and comprehensive marker resource development in mulberry.

BACKGROUND: High potential of Morus laevigata and Morus serrata has been proposed in the breeding programs for Morus sp. However, due to the lack of dense molecular markers this goal is still in its nascent stage and not yet realized. We thus, sequenced the transcriptomes of these two wild Morus species and utilized the data for marker development. RESULTS: We generated 87.0 and 80.3 Mb of transcriptome data from M. laevigata and M. serrata, respectively. The transcriptomes from M. laevigata and M. serrata, were assembled into 95,181 and 85,269 transcripts, respectively, and annotated. We identified around 24,049 Simple Sequence Repeats (SSRs), 1,201,326 Single Nucleotide Polymorphisms (SNPs) and 67,875 Insertion-Deletions (InDels). The variants having a higher impact were also identified and their effect was further investigated. CONCLUSIONS: The transcriptome resource from the wildly growing mulberry species developed in this study can find wide applicability in gene identification and/or characterization. It can also contribute immensely in the existing mulberry improvement programs.

Comprehensive expression analysis of rice Armadillo gene family during abiotic stress and development.

Genes in the Armadillo (ARM)-repeat superfamily encode proteins with a range of developmental and physiological processes in unicellular and multicellular eukaryotes. These 42 amino acid, long tandem repeat-containing proteins have been abundantly recognized in many plant species. Previous studies have confirmed that Armadillo proteins constitute a multigene family in Arabidopsis. In this study, we performed a computational analysis in the rice genome (Oryza sativa L. subsp. japonica), and identified 158 genes of Armadillo superfamily. Phylogenetic study classified them into several arbitrary groups based on a varying number of non-conserved ARM repeats and accessory domain(s) associated with them. An in-depth analysis of gene expression through microarray and Q-PCR revealed a number of ARM proteins expressing differentially in abiotic stresses and developmental conditions, suggesting a potential roles of this superfamily in development and stress signalling. Comparative phylogenetic analysis between Arabidopsis and rice Armadillo genes revealed a high degree of evolutionary conservation between the orthologues in two plant species. The non-synonymous and synonymous substitutions per site ratios (Ka/Ks) of duplicated gene pairs indicate a purifying selection. This genome-wide identification and expression analysis provides a basis for further functional analysis of Armadillo genes under abiotic stress and reproductive developmental condition in the plant lineage.

Genome-wide expressional and functional analysis of calcium transport elements during abiotic stress and development in rice.

Ca²âº homeostasis is required to maintain a delicate balance of cytosolic Ca²âº during normal and adverse growth conditions. Various Ca²âº transporters actively participate to maintain this delicate balance especially during abiotic stresses and developmental events in plants. In this study, we present a genome-wide account, detailing expression profiles, subcellular localization and functional analysis of rice Ca²âº transport elements. Exhaustive in silico data mining and analysis resulted in the identification of 81 Ca²âº transport element genes, which belong to various groups such as Ca²âº-ATPases (pumps), exchangers, channels, glutamate receptor homologs and annexins. Phylogenetic analysis revealed that different Ca²âº transporters are evolutionarily conserved across different plant species. Comprehensive expression analysis by gene chip microarray and quantitative RT-PCR revealed that a substantial proportion of Ca²âº transporter genes were expressed differentially under abiotic stresses (salt, cold and drought) and reproductive developmental stages (panicle and seed) in rice. These findings suggest a possible role of rice Ca²âº transporters in abiotic stress and development triggered signaling pathways. Subcellular localization of Ca²âº transporters from different groups in Nicotiana benthamiana revealed their variable localization to different compartments, which could be their possible sites of action. Complementation of Ca²âº transport activity of K616 yeast mutant by Ca²âº-ATPase OsACA7 and involvement in salt tolerance verified its functional behavior. This study will encourage detailed characterization of potential candidate Ca²âº transporters for their functional role in planta.

Heterosis: emerging ideas about hybrid vigour.

Perceived by Charles Darwin in many vegetable plants and rediscovered by George H Shull and Edward M East in maize, heterosis or hybrid vigour is one of the most widely utilized phenomena, not only in agriculture but also in animal breeding. Although, numerous studies have been carried out to understand its genetic and/or molecular basis in the past 100 years, our knowledge of the underlying molecular processes that results in hybrid vigour can best be defined as superficial. Even after century long deliberations, there is no consensus on the relative/individual contribution of the genetic/epigenetic factors in the manifestation of heterosis. However, with the recent advancements in functional genomics, transcriptomics, proteomics, and metabolomics-related technologies, the riddle of heterosis is being reinvestigated by adopting systems-level approaches to understand the underlying molecular mechanisms. A number of intriguing hypotheses are converging towards the idea of a cumulative positive effect of the differential expression of a variety of genes, on one or several yield-affecting metabolic pathways or overall energy-use efficiency, as the underlying mechanism for the manifestation of heterosis. Presented here is a brief account of clues gathered from various investigative approaches targeted towards better scientific understanding of this process.

Comprehensive expression analysis of rice phospholipase D gene family during abiotic stresses and development.

Phospholipase D is one of the crucial enzymes involved in lipid mediated signaling, triggered during various developmental and physiological processes. Different members of PLD gene family have been known to be induced under different abiotic stresses and during developmental processes in various plant species. In this report, we are presenting a detailed microarray based expression analysis and expression profiles of entire set of PLD genes in rice genome, under three abiotic stresses (salt, cold and drought) and different developmental stages (3-vegetative stages and 11-reproductive stages). Seven and nine PLD genes were identified, which were expressed differentially under abiotic stresses and during reproductive developmental stages, respectively. PLD genes, which were expressed significantly under abiotic stresses exhibited an overlapping expression pattern and were also differentially expressed during developmental stages. Moreover, expression pattern for a set of stress induced genes was validated by real time PCR and it supported the microarray expression data. These findings emphasize the role of PLDs in abiotic stress signaling and development in rice. In addition, expression profiling for duplicated PLD genes revealed a functional divergence between the duplicated genes and signify the role of gene duplication in the evolution of this gene family in rice. This expressional study will provide an important platform in future for the functional characterization of PLDs in crop plants.

Rice phospholipase A superfamily: organization, phylogenetic and expression analysis during abiotic stresses and development.

BACKGROUND: Phospholipase A (PLA) is an important group of enzymes responsible for phospholipid hydrolysis in lipid signaling. PLAs have been implicated in abiotic stress signaling and developmental events in various plants species. Genome-wide analysis of PLA superfamily has been carried out in dicot plant Arabidopsis. A comprehensive genome-wide analysis of PLAs has not been presented yet in crop plant rice. METHODOLOGY/PRINCIPAL FINDINGS: A comprehensive bioinformatics analysis identified a total of 31 PLA encoding genes in the rice genome, which are divided into three classes; phospholipase A(1) (PLA(1)), patatin like phospholipases (pPLA) and low molecular weight secretory phospholipase A(2) (sPLA(2)) based on their sequences and phylogeny. A subset of 10 rice PLAs exhibited chromosomal duplication, emphasizing the role of duplication in the expansion of this gene family in rice. Microarray expression profiling revealed a number of PLA members expressing differentially and significantly under abiotic stresses and reproductive development. Comparative expression analysis with Arabidopsis PLAs revealed a high degree of functional conservation between the orthologs in two plant species, which also indicated the vital role of PLAs in stress signaling and plant development across different plant species. Moreover, sub-cellular localization of a few candidates suggests their differential localization and functional role in the lipid signaling. CONCLUSION/SIGNIFICANCE: The comprehensive analysis and expression profiling would provide a critical platform for the functional characterization of the candidate PLA genes in crop plants.