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  1. Fulltext Perigone Lobe Transcriptome Analysis Provides Insights into Rafflesia cantleyi Flower Development
    Lee XW, Mat-Isa MN, Mohd-Elias NA, Aizat-Juhari MA, Goh HH, Dear PH, et al.
    PLoS One, 2016;11(12):e0167958.
    PMID: 27977777 DOI: 10.1371/journal.pone.0167958
    Rafflesia is a biologically enigmatic species that is very rare in occurrence and possesses an extraordinary morphology. This parasitic plant produces a gigantic flower up to one metre in diameter with no leaves, stem or roots. However, little is known about the floral biology of this species especially at the molecular level. In an effort to address this issue, we have generated and characterised the transcriptome of the Rafflesia cantleyi flower, and performed a comparison with the transcriptome of its floral bud to predict genes that are expressed and regulated during flower development. Approximately 40 million sequencing reads were generated and assembled de novo into 18,053 transcripts with an average length of 641 bp. Of these, more than 79% of the transcripts had significant matches to annotated sequences in the public protein database. A total of 11,756 and 7,891 transcripts were assigned to Gene Ontology categories and clusters of orthologous groups respectively. In addition, 6,019 transcripts could be mapped to 129 pathways in Kyoto Encyclopaedia of Genes and Genomes Pathway database. Digital abundance analysis identified 52 transcripts with very high expression in the flower transcriptome of R. cantleyi. Subsequently, analysis of differential expression between developing flower and the floral bud revealed a set of 105 transcripts with potential role in flower development. Our work presents a deep transcriptome resource analysis for the developing flower of R. cantleyi. Genes potentially involved in the growth and development of the R. cantleyi flower were identified and provide insights into biological processes that occur during flower development.
  2. Fulltext Transcriptome landscape of Rafflesia cantleyi floral buds reveals insights into the roles of transcription factors and phytohormones in flower development
    Amini S, Rosli K, Abu-Bakar MF, Alias H, Mat-Isa MN, Juhari MA, et al.
    PLoS One, 2019;14(12):e0226338.
    PMID: 31851702 DOI: 10.1371/journal.pone.0226338
    Rafflesia possesses unique biological features and known primarily for producing the world's largest and existing as a single flower. However, to date, little is known about key regulators participating in Rafflesia flower development. In order to further understand the molecular mechanism that regulates Rafflesia cantleyi flower development, RNA-seq data from three developmental stages of floral bud, representing the floral organ primordia initiation, floral organ differentiation, and floral bud outgrowth, were analysed. A total of 89,890 transcripts were assembled of which up to 35% could be annotated based on homology search. Advanced transcriptome analysis using K-mean clustering on the differentially expressed genes (DEGs) was able to identify 12 expression clusters that reflect major trends and key transitional states, which correlate to specific developmental stages. Through this, comparative gene expression analysis of different floral bud stages identified various transcription factors related to flower development. The members of WRKY, NAC, bHLH, and MYB families are the most represented among the DEGs, suggesting their important function in flower development. Furthermore, pathway enrichment analysis also revealed DEGs that are involved in various phytohormone signal transduction events such as auxin and auxin transport, cytokinin and gibberellin biosynthesis. Results of this study imply that transcription factors and phytohormone signalling pathways play major role in Rafflesia floral bud development. This study provides an invaluable resource for molecular studies of the flower development process in Rafflesia and other plant species.
  3. Fulltext Transcriptome analysis of Rafflesia cantleyi flower stages reveals insights into the regulation of senescence
    Mohd-Elias NA, Rosli K, Alias H, Juhari MA, Abu-Bakar MF, Md-Isa N, et al.
    Sci Rep, 2021 Dec 08;11(1):23661.
    PMID: 34880337 DOI: 10.1038/s41598-021-03028-x
    Rafflesia is a unique plant species existing as a single flower and produces the largest flower in the world. While Rafflesia buds take up to 21 months to develop, its flowers bloom and wither within about a week. In this study, transcriptome analysis was carried out to shed light on the molecular mechanism of senescence in Rafflesia. A total of 53.3 million high quality reads were obtained from two Rafflesia cantleyi flower developmental stages and assembled to generate 64,152 unigenes. Analysis of this dataset showed that 5,166 unigenes were differentially expressed, in which 1,073 unigenes were identified as genes involved in flower senescence. Results revealed that as the flowers progress to senescence, more genes related to flower senescence were significantly over-represented compared to those related to plant growth and development. Senescence of the R. cantleyi flower activates senescence-associated genes in the transcription activity (members of the transcription factor families MYB, bHLH, NAC, and WRKY), nutrient remobilization (autophagy-related protein and transporter genes), and redox regulation (CATALASE). Most of the senescence-related genes were found to be differentially regulated, perhaps for the fine-tuning of various responses in the senescing R. cantleyi flower. Additionally, pathway analysis showed the activation of genes such as ETHYLENE RECEPTOR, ETHYLENE-INSENSITIVE 2, ETHYLENE-INSENSITIVE 3, and ETHYLENE-RESPONSIVE TRANSCRIPTION FACTOR, indicating the possible involvement of the ethylene hormone response pathway in the regulation of R. cantleyi senescence. Our results provide a model of the molecular mechanism underlying R. cantleyi flower senescence, and contribute essential information towards further understanding the biology of the Rafflesiaceae family.
  4. Fulltext Comparative metabolomics analysis reveals alkaloid repertoires in young and mature Mitragyna speciosa (Korth.) Havil. Leaves
    Veeramohan R, Zamani AI, Azizan KA, Goh HH, Aizat WM, Razak MFA, et al.
    PLoS One, 2023;18(3):e0283147.
    PMID: 36943850 DOI: 10.1371/journal.pone.0283147
    The fresh leaves of Mitragyna speciosa (Korth.) Havil. have been traditionally consumed for centuries in Southeast Asia for its healing properties. Although the alkaloids of M. speciosa have been studied since the 1920s, comparative and systematic studies of metabolite composition based on different leaf maturity levels are still lacking. This study assessed the secondary metabolite composition in two different leaf stages (young and mature) of M. speciosa, using an untargeted liquid chromatography-electrospray ionisation-time-of-flight-mass spectrometry (LC-ESI-TOF-MS) metabolite profiling. The results revealed 86 putatively annotated metabolite features (RT:m/z value) comprising 63 alkaloids, 10 flavonoids, 6 terpenoids, 3 phenylpropanoids, and 1 of each carboxylic acid, glucoside, phenol, and phenolic aldehyde. The alkaloid features were further categorised into 14 subclasses, i.e., the most abundant class of secondary metabolites identified. As per previous reports, indole alkaloids are the most abundant alkaloid subclass in M. speciosa. The result of multivariate analysis (MVA) using principal component analysis (PCA) showed a clear separation of 92.8% between the young and mature leaf samples, indicating a high variance in metabolite levels between them. Akuammidine, alstonine, tryptamine, and yohimbine were tentatively identified among the many new alkaloids reported in this study, depicting the diverse biological activities of M. speciosa. Besides delving into the knowledge of metabolite distribution in different leaf stages, these findings have extended the current alkaloid repository of M. speciosa for a better understanding of its pharmaceutical potential.
  5. Fulltext RNA-seq analysis for secondary metabolite pathway gene discovery in Polygonum minus
    Loke KK, Rahnamaie-Tajadod R, Yeoh CC, Goh HH, Mohamed-Hussein ZA, Mohd Noor N, et al.
    Genom Data, 2016 Mar;7:12-3.
    PMID: 26981350 DOI: 10.1016/j.gdata.2015.11.003
    Polygonum minus plant is rich in secondary metabolites, especially terpenoids and flavonoids. Present study generates transcriptome resource for P. minus to decipher its secondary metabolite biosynthesis pathways. Raw reads and the transcriptome assembly project have been deposited at GenBank under the accessions SRX313492 (root) and SRX669305 (leaf) respectively.
  6. Recent advancement of engineering microbial hosts for the biotechnological production of flavonoids
    Shah FLA, Ramzi AB, Baharum SN, Noor NM, Goh HH, Leow TC, et al.
    Mol Biol Rep, 2019 Dec;46(6):6647-6659.
    PMID: 31535322 DOI: 10.1007/s11033-019-05066-1
    Flavonoids are polyphenols that are important organic chemicals in plants. The health benefits of flavonoids that result in high commercial values make them attractive targets for large-scale production through bioengineering. Strategies such as engineering a flavonoid biosynthetic pathway in microbial hosts provide an alternative way to produce these beneficial compounds. Escherichia coli, Saccharomyces cerevisiae and Streptomyces sp. are among the expression systems used to produce recombinant products, as well as for the production of flavonoid compounds through various bioengineering approaches including clustered regularly interspaced short palindromic repeats (CRISPR)-based genome engineering and genetically encoded biosensors to detect flavonoid biosynthesis. In this study, we review the recent advances in engineering model microbial hosts as being the factory to produce targeted flavonoid compounds.
  7. Fulltext RNA-seq data from different developmental stages of Rafflesia cantleyi floral buds
    Amini S, Alias H, Aizat-Juhari MA, Mat-Isa MN, Adam JH, Goh HH, et al.
    Genom Data, 2017 Dec;14:5-6.
    PMID: 28761813 DOI: 10.1016/j.gdata.2017.07.008
    Rafflesia cantleyi, known as one of the world's largest flowers, is a specialised holoparasite due to dramatic morphological modifications. It possesses highly reduced vegetative structure and only appears as a flower for sexual reproduction. Moreover, it has an unusual life cycle in that its floral bud development takes up to nine months. In order to fully understand the highly modified floral organ structure and long life cycle of R. cantleyi, we used Illumina sequencing technology (HiSeq) for sequence generation followed by de novo assembly of sequence reads. We obtained the RNA-seq data from three different stages of floral bud, representing the early, mid and advanced developmental stages. These data are available via BioProject accession number PRJNA378435. More than 10.3 Gb raw sequence data were generated, corresponding to 102,203,042 raw reads. Following removal of low-quality reads and trimming of adapter sequences, a total of 91,638,836 reads were obtained. De novo assembly of these sequences using Trinity resulted in 89,690 unique transcripts with an N50 of 1653 bp. The obtained transcriptomic data will be useful for further study to understand the molecular interactions that result in R. cantleyi floral development.
  8. Fulltext Metabolomics data of Mitragyna speciosa leaf using LC-ESI-TOF-MS
    Veeramohan R, Azizan KA, Aizat WM, Goh HH, Mansor SM, Yusof NSM, et al.
    Data Brief, 2018 Jun;18:1212-1216.
    PMID: 29900296 DOI: 10.1016/j.dib.2018.04.001
    Mitragyna speciosa is a psychoactive plant known as "ketum" in Malaysia and "kratom" in Thailand. This plant is distinctly known to produce two important alkaloids, namely mitragynine (MG) and 7-hydroxymitragynine (7-OH-MG) that can bind to opioid receptors [1]. MG was reported to exhibit antidepressant properties in animal studies [2]. These compounds were also proposed to have the potential to replace opioid analgesics with much lower risks of side effects [3]. To date, there are only over 40 metabolites identified in M. speciosa [4,5]. To obtain a more complete profile of secondary metabolites in ketum, we performed metabolomics study using mature leaves of the green M. speciosa variety. The leaf samples were extracted using methanol prior to liquid chromatography-electrospray ionization-time of flight-mass spectrometry (LC-ESI-TOF-MS) analysis. This data can be useful to for the identification of unknown metabolites that are associated with alkaloid biosynthesis pathway in M. speciosa.
  9. Harnessing landfill gas (LFG) for electricity: A strategy to mitigate greenhouse gas (GHG) emissions in Jakarta (Indonesia)
    Kurniawan TA, Liang X, Singh D, Othman MHD, Goh HH, Gikas P, et al.
    J Environ Manage, 2022 Jan 01;301:113882.
    PMID: 34638040 DOI: 10.1016/j.jenvman.2021.113882
    Due to its increasing demands for fossil fuels, Indonesia needs an alternative energy to diversify its energy supply. Landfill gas (LFG), which key component is methane (CH4), has become one of the most attractive options to sustain its continued economic development. This exploratory study seeks to demonstrate the added value of landfilled municipal solid waste (MSW) in generating sustainable energy, resulting from CH4 emissions in the Bantargebang landfill (Jakarta). The power generation capacity of a waste-to-energy (WTE) plant based on a mathematical modeling was investigated. This article critically evaluated the production of electricity and potential income from its sale in the market. The project's environmental impact assessment and its socio-economic and environmental benefits in terms of quantitative and qualitative aspects were discussed. It was found that the emitted CH4 from the landfill could be reduced by 25,000 Mt annually, while its electricity generation could reach one million kW ⋅h annually, savings on equivalent electricity charge worth US$ 112 million/year (based on US' 8/kW ⋅ h). An equivalent CO2 mitigation of 3.4 × 106 Mt/year was obtained. The income from its power sale were US$ 1.2 ×106 in the 1st year and 7.7 ×107US$ in the 15th year, respectively, based on the projected CH4 and power generation. The modeling study on the Bantargebang landfill using the LFG extraction data indicated that the LFG production ranged from 0.05 to 0.40 m3 per kg of the landfilled MSW. The LFG could generate electricity as low as US' 8 per kW ⋅ h. With respect to the implications of this study, the revenue not only defrays the cost of landfill's operations and maintenance (O&M), but also provides an incentive and means to further improve its design and operations. Overall, this work not only leads to a diversification of primary energy, but also improves environmental protection and the living standard of the people surrounding the plant.
  10. Overview of Repressive miRNA Regulation by Short Tandem Target Mimic (STTM): Applications and Impact on Plant Biology
    Othman SMIS, Mustaffa AF, Che-Othman MH, Samad AFA, Goh HH, Zainal Z, et al.
    Plants (Basel), 2023 Feb 03;12(3).
    PMID: 36771753 DOI: 10.3390/plants12030669
    The application of miRNA mimic technology for silencing mature miRNA began in 2007. This technique originated from the discovery of the INDUCED BY PHOSPHATE STARVATION 1 (IPS1) gene, which was found to be a competitive mimic that prevents the cleavage of the targeted mRNA by miRNA inhibition at the post-transcriptional level. To date, various studies have been conducted to understand the molecular mimic mechanism and to improve the efficiency of this technology. As a result, several mimic tools have been developed: target mimicry (TM), short tandem target mimic (STTM), and molecular sponges (SPs). STTM is the most-developed tool due to its stability and effectiveness in decoying miRNA. This review discusses the application of STTM technology on the loss-of-function studies of miRNA and members from diverse plant species. A modified STTM approach for studying the function of miRNA with spatial-temporal expression under the control of specific promoters is further explored. STTM technology will enhance our understanding of the miRNA activity in plant-tissue-specific development and stress responses for applications in improving plant traits via miRNA regulation.
  11. Source, occurrence, distribution, fate, and implications of microplastic pollutants in freshwater on environment: A critical review and way forward
    Kurniawan TA, Haider A, Ahmad HM, Mohyuddin A, Umer Aslam HM, Nadeem S, et al.
    Chemosphere, 2023 Jun;325:138367.
    PMID: 36907482 DOI: 10.1016/j.chemosphere.2023.138367
    The generation of microplastics (MPs) has increased recently and become an emerging issue globally. Due to their long-term durability and capability of traveling between different habitats in air, water, and soil, MPs presence in freshwater ecosystem threatens the environment with respect to its quality, biotic life, and sustainability. Although many previous works have been undertaken on the MPs pollution in the marine system recently, none of the study has covered the scope of MPs pollution in the freshwater. To consolidate scattered knowledge in the literature body into one place, this work identifies the sources, fate, occurrence, transport pathways, and distribution of MPs pollution in the aquatic system with respect to their impacts on biotic life, degradation, and detection techniques. This article also discusses the environmental implications of MPs pollution in the freshwater ecosystems. Certain techniques for identifying MPs and their limitations in applications are presented. Through a literature survey of over 276 published articles (2000-2023), this study presents an overview of solutions to the MP pollution, while identifying research gaps in the body of knowledge for further work. It is conclusive from this review that the MPs exist in the freshwater due to an improper littering of plastic waste and its degradation into smaller particles. Approximately 15-51 trillion MP particles have accumulated in the oceans with their weight ranging between 93,000 and 236,000 metric ton (Mt), while about 19-23 Mt of plastic waste was released into rivers in 2016, which was projected to increase up to 53 Mt by 2030. A subsequent degradation of MPs in the aquatic environment results in the generation of NPs with size ranging from 1 to 1000 nm. It is expected that this work facilitates stakeholders to understand the multi-aspects of MPs pollution in the freshwater and recommends policy actions to implement sustainable solutions to this environmental problem.
  12. Decarbonization in waste recycling industry using digitalization to promote net-zero emissions and its implications on sustainability
    Kurniawan TA, Othman MHD, Liang X, Goh HH, Gikas P, Kusworo TD, et al.
    J Environ Manage, 2023 Jul 15;338:117765.
    PMID: 36965421 DOI: 10.1016/j.jenvman.2023.117765
    Digitalization and sustainability have been considered as critical elements in tackling a growing problem of solid waste in the framework of circular economy (CE). Although digitalization can enhance time-efficiency and/or cost-efficiency, their end-results do not always lead to sustainability. So far, the literatures still lack of a holistic view in understanding the development trends and key roles of digitalization in waste recycling industry to benefit stakeholders and to protect the environment. To bridge this knowledge gap, this work systematically investigates how leveraging digitalization in waste recycling industry could address these research questions: (1) What are the key problems of solid waste recycling? (2) How the trends of digitalization in waste management could benefit a CE? (3) How digitalization could strengthen waste recycling industry in a post-pandemic era? While digitalization boosts material flows in a CE, it is evident that utilizing digital solutions to strengthen waste recycling business could reinforce a resource-efficient, low-carbon, and a CE. In the Industry 4.0 era, digitalization can add 15% (about USD 15.7 trillion) to global economy by 2030. As digitalization grows, making the waste sector shift to a CE could save between 30% and 35% of municipalities' waste management budget. With digitalization, a cost reduction of 3.6% and a revenue increase of 4.1% are projected annually. This would contribute to USD 493 billion in an increasing revenue yearly in the next decade. As digitalization enables tasks to be completed shortly with less manpower, this could save USD 421 billion annually for the next decade. With respect to environmental impacts, digitalization in the waste sector could reduce global CO2 emissions by 15% by 2030 through technological solutions. Overall, this work suggests that digitalization in the waste sector contributes net-zero emission to a digital economy, while transitioning to a sustainable world as its social impacts.
  13. Leveraging food waste for electricity: A low-carbon approach in energy sector for mitigating climate change and achieving net zero emission in Hong Kong (China)
    Kurniawan TA, Liang X, Goh HH, Dzarfan Othman MH, Anouzla A, Al-Hazmi HE, et al.
    J Environ Manage, 2024 Feb;351:119879.
    PMID: 38157574 DOI: 10.1016/j.jenvman.2023.119879
    In recent years, food waste has been a global concern that contributes to climate change. To deal with the rising impacts of climate change, in Hong Kong, food waste is converted into electricity in the framework of low-carbon approach. This work provides an overview of the conversion of food waste into electricity to achieve carbon neutrality. The production of methane and electricity from waste-to-energy (WTE) conversion are determined. Potential income from its sale and environmental benefits are also assessed quantitatively and qualitatively. It was found that the electricity generation from the food waste could reach 4.33 × 109 kWh annually, avoiding equivalent electricity charge worth USD 3.46 × 109 annually (based on US' 8/kWh). An equivalent CO2 mitigation of 9.9 × 108 kg annually was attained. The revenue from its electricity sale in market was USD 1.44×109 in the 1st year and USD 4.24 ×109 in the 15th year, respectively, according to the projected CH4 and electricity generation. The modelling study indicated that the electricity production is 0.8 kWh/kg of landfilled waste. The food waste could produce electricity as low as US' 8 per kW ∙ h. In spite of its promising results, there are techno-economic bottlenecks in commercial scale production and its application at comparable costs to conventional fossil fuels. Issues such as high GHG emissions and high production costs have been determined to be resolved later. Overall, this work not only leads to GHG avoidance, but also diversifies energy supply in providing power for homes in the future.
  14. Harnessing the potential of non-coding RNA: An insight into its mechanism and interaction in plant biotic stress
    Othman SMIS, Mustaffa AF, Mohd Zahid NII, Che-Othman MH, Samad AFA, Goh HH, et al.
    Plant Physiol Biochem, 2024 Feb;207:108387.
    PMID: 38266565 DOI: 10.1016/j.plaphy.2024.108387
    Plants have developed diverse physical and chemical defence mechanisms to ensure their continued growth and well-being in challenging environments. Plants also have evolved intricate molecular mechanisms to regulate their responses to biotic stress. Non-coding RNA (ncRNA) plays a crucial role in this process that affects the expression or suppression of target transcripts. While there have been numerous reviews on the role of molecules in plant biotic stress, few of them specifically focus on how plant ncRNAs enhance resistance through various mechanisms against different pathogens. In this context, we explored the role of ncRNA in exhibiting responses to biotic stress endogenously as well as cross-kingdom regulation of transcript expression. Furthermore, we address the interplay between ncRNAs, which can act as suppressors, precursors, or regulators of other ncRNAs. We also delve into the regulation of ncRNAs in response to attacks from different organisms, such as bacteria, viruses, fungi, nematodes, oomycetes, and insects. Interestingly, we observed that diverse microorganisms interact with distinct ncRNAs. This intricacy leads us to conclude that each ncRNA serves a specific function in response to individual biotic stimuli. This deeper understanding of the molecular mechanisms involving ncRNAs in response to biotic stresses enhances our knowledge and provides valuable insights for future research in the field of ncRNA, ultimately leading to improvements in plant traits.
  15. Fulltext Transcriptome analysis of Polygonum minus reveals candidate genes involved in important secondary metabolic pathways of phenylpropanoids and flavonoids
    Loke KK, Rahnamaie-Tajadod R, Yeoh CC, Goh HH, Mohamed-Hussein ZA, Zainal Z, et al.
    PeerJ, 2017;5:e2938.
    PMID: 28265493 DOI: 10.7717/peerj.2938
    BACKGROUND: Polygonum minus is an herbal plant in the Polygonaceae family which is rich in ethnomedicinal plants. The chemical composition and characteristic pungent fragrance of Polygonum minus have been extensively studied due to its culinary and medicinal properties. There are only a few transcriptome sequences available for species from this important family of medicinal plants. The limited genetic information from the public expressed sequences tag (EST) library hinders further study on molecular mechanisms underlying secondary metabolite production.

    METHODS: In this study, we performed a hybrid assembly of 454 and Illumina sequencing reads from Polygonum minus root and leaf tissues, respectively, to generate a combined transcriptome library as a reference.

    RESULTS: A total of 34.37 million filtered and normalized reads were assembled into 188,735 transcripts with a total length of 136.67 Mbp. We performed a similarity search against all the publicly available genome sequences and found similarity matches for 163,200 (86.5%) of Polygonum minus transcripts, largely from Arabidopsis thaliana (58.9%). Transcript abundance in the leaf and root tissues were estimated and validated through RT-qPCR of seven selected transcripts involved in the biosynthesis of phenylpropanoids and flavonoids. All the transcripts were annotated against KEGG pathways to profile transcripts related to the biosynthesis of secondary metabolites.

    DISCUSSION: This comprehensive transcriptome profile will serve as a useful sequence resource for molecular genetics and evolutionary research on secondary metabolite biosynthesis in Polygonaceae family. Transcriptome assembly of Polygonum minus can be accessed at http://prims.researchfrontier.org/index.php/dataset/transcriptome.

  16. Fulltext DNA-seq analysis of Garcinia mangostana
    Abu Bakar S, Sampathrajan S, Loke KK, Goh HH, Mohd Noor N
    Genom Data, 2016 Mar;7:62-3.
    PMID: 26981362 DOI: 10.1016/j.gdata.2015.11.018
    Mangosteen (Garcinia mangostana Linn.) is a tropical tree mainly found in South East Asia and considered as "the queen of fruits". The asexually produced fruit is dark purple or reddish in color, with white flesh which is slightly acidic with sweet flavor and a pleasant aroma. The purple pericarp tissue is rich in xanthones which are useful for medical purposes. We performed the first genome sequencing of this commercially important fruit tree to study its genome composition and attempted draft genome assembly. Raw reads of the DNA sequencing project have been deposited to SRA database with the accession number SRX1426419.
  17. Fulltext RNA-seq analysis for plant carnivory gene discovery in Nepenthes × ventrata
    Wan Zakaria WN, Loke KK, Goh HH, Mohd Noor N
    Genom Data, 2016 Mar;7:18-9.
    PMID: 26981352 DOI: 10.1016/j.gdata.2015.11.007
    Carnivorous plants have the ability to capture and digest insects for nutrients, which allows them to survive in land deprived of nitrogenous nutrients. Nepenthes spp. are one of the carnivorous plants, which uniquely produce pitcher from the tip of an elongated leaf. This study provides the first transcriptome resource from pitcher of a Nepenthes ventricosa × Nepenthes alata hybrid, Nepenthes × ventrata to understand carnivory mechanism in Nepenthes spp., as well as in other carnivorous species. Raw reads and the transcriptome assembly project have been deposited to SRA database with the accession numbers SRX1389337 (day 0 control), SRX1389392 (day 3 longevity), and SRX1389395 (day 3 chitin-treated).
  18. Transcriptome-wide effects of expansin gene manipulation in etiolated Arabidopsis seedling
    Ilias IA, Negishi K, Yasue K, Jomura N, Morohashi K, Baharum SN, et al.
    J Plant Res, 2019 Mar;132(2):159-172.
    PMID: 30341720 DOI: 10.1007/s10265-018-1067-0
    Expansin is a non-enzymatic protein which plays a pivotal role in cell wall loosening by inducing stress relaxation and extension in the plant cell wall. Previous studies on Arabidopsis, Petunia × hybrida, and tomato demonstrated that the suppression of expansin gene expression reduced plant growth but expansin overexpression does not necessarily promotes growth. In this study, both expansin gene suppression and overexpression in dark-grown transgenic Arabidopsis seedlings resulted in reduced hypocotyl length at late growth stages with a more pronounced effect for the overexpression. This defect in hypocotyl elongation raises questions about the molecular effect of expansin gene manipulation. RNA-seq analysis of the transcriptomic changes between day 3 and day 5 seedlings for both transgenic lines found numerous differentially expressed genes (DEGs) including transcription factors and hormone-related genes involved in different aspects of cell wall development. These DEGs imply that the observed hypocotyl growth retardation is a consequence of the concerted effect of regulatory factors and multiple cell-wall related genes, which are important for cell wall remodelling during rapid hypocotyl elongation. This is further supported by co-expression analysis through network-centric approach of differential network cluster analysis. This first transcriptome-wide study of expansin manipulation explains why the effect of expansin overexpression is greater than suppression and provides insights into the dynamic nature of molecular regulation during etiolation.
  19. Protein replenishment in pitcher fluids of Nepenthes × ventrata revealed by quantitative proteomics (SWATH-MS) informed by transcriptomics
    Wan Zakaria WNA, Aizat WM, Goh HH, Mohd Noor N
    J Plant Res, 2019 Sep;132(5):681-694.
    PMID: 31422552 DOI: 10.1007/s10265-019-01130-w
    Carnivorous plants capture and digest insects for nutrients, allowing them to survive in soil deprived of nitrogenous nutrients. Plants from the genus Nepenthes produce unique pitchers containing secretory glands, which secrete enzymes into the digestive fluid. We performed RNA-seq analysis on the pitcher tissues and liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis on the pitcher fluids of Nepenthes × ventrata to study protein expression in this carnivory organ during early days of pitcher opening. This transcriptome provides a sequence database for pitcher fluid protein identification. A total of 32 proteins of diverse functions were successfully identified in which 19 proteins can be quantified based on label-free quantitative proteomics (SWATH-MS) analysis while 16 proteins were not reported previously. Our findings show that certain proteins in the pitcher fluid were continuously secreted or replenished after pitcher opening, even without any prey or chitin induction. We also discovered a new aspartic proteinase, Nep6, secreted into pitcher fluid. This is the first SWATH-MS analysis of protein expression in Nepenthes pitcher fluid using a species-specific reference transcriptome. Taken together, our study using a gel-free shotgun proteomics informed by transcriptomics (PIT) approach showed the dynamics of endogenous protein secretion in the digestive organ of N. × ventrata and provides insights on protein regulation during early pitcher opening prior to prey capture.
  20. Fulltext Transcriptome-wide shift from photosynthesis and energy metabolism upon endogenous fluid protein depletion in young Nepenthes ampullaria pitchers
    Goh HH, Baharin A, Mohd Salleh F', Ravee R, Wan Zakaria WNA, Mohd Noor N
    Sci Rep, 2020 04 20;10(1):6575.
    PMID: 32313042 DOI: 10.1038/s41598-020-63696-z
    Carnivorous pitcher plants produce specialised pitcher organs containing secretory glands, which secrete acidic fluids with hydrolytic enzymes for prey digestion and nutrient absorption. The content of pitcher fluids has been the focus of many fluid protein profiling studies. These studies suggest an evolutionary convergence of a conserved group of similar enzymes in diverse families of pitcher plants. A recent study showed that endogenous proteins were replenished in the pitcher fluid, which indicates a feedback mechanism in protein secretion. This poses an interesting question on the physiological effect of plant protein loss. However, there is no study to date that describes the pitcher response to endogenous protein depletion. To address this gap of knowledge, we previously performed a comparative RNA-sequencing experiment of newly opened pitchers (D0) against pitchers after 3 days of opening (D3C) and pitchers with filtered endogenous proteins (>10 kDa) upon pitcher opening (D3L). Nepenthes ampullaria was chosen as a model study species due to their abundance and unique feeding behaviour on leaf litters. The analysis of unigenes with top 1% abundance found protein translation and stress response to be overrepresented in D0, compared to cell wall related, transport, and signalling for D3L. Differentially expressed gene (DEG) analysis identified DEGs with functional enrichment in protein regulation, secondary metabolism, intracellular trafficking, secretion, and vesicular transport. The transcriptomic landscape of the pitcher dramatically shifted towards intracellular transport and defence response at the expense of energy metabolism and photosynthesis upon endogenous protein depletion. This is supported by secretome, transportome, and transcription factor analysis with RT-qPCR validation based on independent samples. This study provides the first glimpse into the molecular responses of pitchers to protein loss with implications to future cost/benefit analysis of carnivorous pitcher plant energetics and resource allocation for adaptation in stochastic environments.
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