Duckweed: Beyond an Efficient Plant Model System.

Duckweed (Lemnaceae) rises as a crucial model system due to its unique characteristics and wide-ranging utility. The significance of physiological research and phytoremediation highlights the intricate potential of duckweed in the current era of plant biology. Special attention to duckweed has been brought due to its distinctive features of nutrient uptake, ion transport dynamics, detoxification, intricate signaling, and stress tolerance. In addition, duckweed can alleviate environmental pollutants and enhance sustainability by participating in bioremediation processes and wastewater treatment. Furthermore, insights into the genomic complexity of Lemnaceae species and the flourishing field of transgenic development highlight the opportunities for genetic manipulation and biotechnological innovations. Novel methods for the germplasm conservation of duckweed can be adopted to preserve genetic diversity for future research endeavors and breeding programs. This review centers around prospects in duckweed research promoting interdisciplinary collaborations and technological advancements to drive its full potential as a model organism.

Comparative analysis, diversification, and functional validation of plant nucleotide-binding site domain genes.

Nucleotide-binding site (NBS) domain genes are one of the superfamily of resistance genes involved in plant responses to pathogens. The current study identified 12,820 NBS-domain-containing genes across 34 species covering from mosses to monocots and dicots. These identified genes are classified into 168 classes with several novel domain architecture patterns encompassing significant diversity among plant species. Several classical (NBS, NBS-LRR, TIR-NBS, TIR-NBS-LRR, etc.) and species-specific structural patterns (TIR-NBS-TIR-Cupin_1-Cupin_1, TIR-NBS-Prenyltransf, Sugar_tr-NBS etc.) were discovered. We observed 603 orthogroups (OGs) with some core (most common orthogroups; OG0, OG1, OG2, etc.) and unique (highly specific to species; OG80, OG82, etc.) OGs with tandem duplications. The expression profiling presented the putative upregulation of OG2, OG6, and OG15 in different tissues under various biotic and abiotic stresses in susceptible and tolerant plants to cotton leaf curl disease (CLCuD). The genetic variation between susceptible (Coker 312) and tolerant (Mac7) Gossypium hirsutum accessions identified several unique variants in NBS genes of Mac7 (6583 variants) and Coker312 (5173 variants). The protein-ligand and proteins-protein interaction showed a strong interaction of some putative NBS proteins with ADP/ATP and different core proteins of the cotton leaf curl disease virus. The silencing of GaNBS (OG2) in resistant cotton through virus-induced gene silencing (VIGS) demonstrated its putative role in virus tittering. The presented study will be further helpful in understanding the plant adaptation mechanism.

Molecular heterogeneity of quiescent melanocyte stem cells revealed by single-cell RNA-sequencing.

Melanocyte stem cells (McSCs) of the hair follicle are a rare cell population within the skin and are notably underrepresented in whole-skin, single-cell RNA sequencing (scRNA-seq) datasets. Using a cell enrichment strategy to isolate KIT+/CD45- cells from the telogen skin of adult female C57BL/6J mice, we evaluated the transcriptional landscape of quiescent McSCs (qMcSCs) at high resolution. Through this evaluation, we confirmed existing molecular signatures for qMcCS subpopulations (e.g., Kit+, Cd34+/-, Plp1+, Cd274+/-, Thy1+, Cdh3+/-) and identified novel qMcSC subpopulations, including two that differentially regulate their immune privilege status. Within qMcSC subpopulations, we also predicted melanocyte differentiation potential, neural crest potential, and quiescence depth. Taken together, the results demonstrate that the qMcSC population is heterogeneous and future studies focused on investigating changes in qMcSCs should consider changes in subpopulation composition.

Integrative systems biology framework discovers common gene regulatory signatures in multiple mechanistically distinct inflammatory skin diseases.

More than 20% of the population across the world is affected by non-communicable inflammatory skin diseases including psoriasis, atopic dermatitis, hidradenitis suppurativa, rosacea, etc. Many of these chronic diseases are painful and debilitating with limited effective therapeutic interventions. However, recent advances in psoriasis treatment have improved the effectiveness and provide better management of the disease. This study aims to identify common regulatory pathways and master regulators that regulate molecular pathogenesis. We designed an integrative systems biology framework to identify the significant regulators across several inflammatory skin diseases. With conventional transcriptome analysis, we identified 55 shared genes, which are enriched in several immune-associated pathways in eight inflammatory skin diseases. Next, we exploited the gene co-expression-, and protein-protein interaction-based networks to identify shared genes and protein components in different diseases with relevant functional implications. Additionally, the network analytics unravels 55 high-value proteins as significant regulators in molecular pathogenesis. We believe that these significant regulators should be explored with critical experimental approaches to identify the putative drug targets for more effective treatments. As an example, we identified IKZF1 as a shared significant master regulator in three inflammatory skin diseases, which can serve as a putative drug target with known disease-derived molecules for developing efficacious combinatorial treatments for hidradenitis suppurativa, atopic dermatitis, and rosacea. The proposed framework is very modular, which can indicate a significant path of molecular mechanism-based drug development from complex transcriptomics data and other multi-omics data.

Integrated Systems Biology Pipeline to Compare Co-Expression Networks in Plants and Elucidate Differential Regulators.

To identify sets of genes that exhibit similar expression characteristics, co-expression networks were constructed from transcriptome datasets that were obtained from plant samples at various stages of growth and development or treated with diverse biotic, abiotic, and other environmental stresses. In addition, co-expression network analysis can provide deeper insights into gene regulation when combined with transcriptomics. The coordination and integration of all these complex networks to deduce gene regulation are major challenges for plant biologists. Python and R have emerged as major tools for managing complex scientific data over the past decade. In this study, we describe a reproducible protocol POTFUL (pant co-expression transcription factor regulators), implemented in Python 3, for integrating co-expression and transcription factor target protein networks to infer gene regulation.

The ORFans' tale: new insights in plant biology.

Orphan genes (OGs) are protein-coding genes without a significant sequence similarity in closely related species. Despite their functional importance, very little is known about the underlying molecular mechanisms by which OGs participate in diverse biological processes. Here, we discuss the evolutionary mechanisms of OGs' emergence with relevance to species-specific adaptations. We also provide a mechanistic view of the involvement of OGs in multiple processes, including growth, development, reproduction, and carbon-metabolism-mediated immunity. We highlight the interconnection between OGs and the sucrose nonfermenting 1 (SNF1)-related protein kinases (SnRKs)-target of rapamycin (TOR) signaling axis for phytohormone signaling, nutrient metabolism, and stress responses. Finally, we propose a high-throughput pipeline for OGs' interspecies and intraspecies gene transfer through a transgenic approach for future biotechnological advances.

The Lost and Found: Unraveling the Functions of Orphan Genes.

Orphan Genes (OGs) are a mysterious class of genes that have recently gained significant attention. Despite lacking a clear evolutionary history, they are found in nearly all living organisms, from bacteria to humans, and they play important roles in diverse biological processes. The discovery of OGs was first made through comparative genomics followed by the identification of unique genes across different species. OGs tend to be more prevalent in species with larger genomes, such as plants and animals, and their evolutionary origins remain unclear but potentially arise from gene duplication, horizontal gene transfer (HGT), or de novo origination. Although their precise function is not well understood, OGs have been implicated in crucial biological processes such as development, metabolism, and stress responses. To better understand their significance, researchers are using a variety of approaches, including transcriptomics, functional genomics, and molecular biology. This review offers a comprehensive overview of the current knowledge of OGs in all domains of life, highlighting the possible role of dark transcriptomics in their evolution. More research is needed to fully comprehend the role of OGs in biology and their impact on various biological processes.

Ranking Plant Network Nodes Based on Their Centrality Measures.

Biological networks are often large and complex, making it difficult to accurately identify the most important nodes. Node prioritization algorithms are used to identify the most influential nodes in a biological network by considering their relationships with other nodes. These algorithms can help us understand the functioning of the network and the role of individual nodes. We developed CentralityCosDist, an algorithm that ranks nodes based on a combination of centrality measures and seed nodes. We applied this and four other algorithms to protein-protein interactions and co-expression patterns in Arabidopsis thaliana using pathogen effector targets as seed nodes. The accuracy of the algorithms was evaluated through functional enrichment analysis of the top 10 nodes identified by each algorithm. Most enriched terms were similar across algorithms, except for DIAMOnD. CentralityCosDist identified more plant-pathogen interactions and related functions and pathways compared to the other algorithms.

Network Biology Analyses and Dynamic Modeling of Gene Regulatory Networks under Drought Stress Reveal Major Transcriptional Regulators in Arabidopsis.

Drought is one of the most serious abiotic stressors in the environment, restricting agricultural production by reducing plant growth, development, and productivity. To investigate such a complex and multifaceted stressor and its effects on plants, a systems biology-based approach is necessitated, entailing the generation of co-expression networks, identification of high-priority transcription factors (TFs), dynamic mathematical modeling, and computational simulations. Here, we studied a high-resolution drought transcriptome of Arabidopsis. We identified distinct temporal transcriptional signatures and demonstrated the involvement of specific biological pathways. Generation of a large-scale co-expression network followed by network centrality analyses identified 117 TFs that possess critical properties of hubs, bottlenecks, and high clustering coefficient nodes. Dynamic transcriptional regulatory modeling of integrated TF targets and transcriptome datasets uncovered major transcriptional events during the course of drought stress. Mathematical transcriptional simulations allowed us to ascertain the activation status of major TFs, as well as the transcriptional intensity and amplitude of their target genes. Finally, we validated our predictions by providing experimental evidence of gene expression under drought stress for a set of four TFs and their major target genes using qRT-PCR. Taken together, we provided a systems-level perspective on the dynamic transcriptional regulation during drought stress in Arabidopsis and uncovered numerous novel TFs that could potentially be used in future genetic crop engineering programs.

Molecular heterogeneity of quiescent melanocyte stem cells revealed by single-cell RNA-sequencing.

Melanocyte stem cells (McSCs) of the hair follicle are a rare cell population within the skin and are notably underrepresented in whole-skin, single-cell RNA sequencing (scRNA-seq) datasets. Using a cell enrichment strategy to isolate KIT+/CD45-cells from the telogen skin of adult female C57BL/6J mice, we evaluated the transcriptional landscape of quiescent McSCs (qMcSCs) at high resolution. Through this evaluation, we confirmed existing molecular signatures for qMcCS subpopulations (e.g., Kit+, Cd34+/- , Plp1+, Cd274+/-, Thy1+, Cdh3+/- ) and identified novel qMcSC subpopulations, including two that differentially regulate their immune privilege status. Within qMcSC subpopulations, we also predicted melanocyte differentiation potential, neural crest potential, and quiescence depth. Taken together, the results demonstrate that the qMcSC population is heterogenous and future studies focused on investigating changes in qMcSCs should consider changes in subpopulation composition. Significance: Single cell transcriptomics has revolutionized our ability to interrogate the dynamic nature of tissues. Here we provide a high-resolution map of the melanocyte stem cell population during quiescence. This map provides one of few examples highlighting broad heterogeneity in stem cells during the quiescent cell state. The map also unifies previous observations using other cell, molecular and functional analyses to define the unique features of the quiescent melanocyte stem cell population. This data provides a valuable resource to individuals interested in further evaluating aspects of cellular quiescence in stem cells broadly or melanocyte stem cells specifically.

A modified standard American diet induces physiological parameters associated with metabolic syndrome in C57BL/6J mice.

Investigations into the causative role that western dietary patterns have on obesity and disease pathogenesis have speculated that quality and quantity of dietary fats and/or carbohydrates have a predictive role in the development of these disorders. Standard reference diets such as the AIN-93 rodent diet have historically been used to promote animal health and reduce variation of results across experiments, rather than model modern human dietary habits or nutrition-related pathologies. In rodents high-fat diets (HFDs) became a classic tool to investigate diet-induced obesity (DIO). These murine diets often relied on a single fat source with the most DIO consistent HFDs containing levels of fat up to 45-60% (kcal), higher than the reported human intake of 33-35% (kcal). More recently, researchers are formulating experimental animal (pre-clinical) diets that reflect mean human macro- and micronutrient consumption levels described by the National Health and Nutrition Examination Survey (NHANES). These diets attempt to integrate relevant ingredient sources and levels of nutrients; however, they most often fail to include high-fructose corn syrup (HFCS) as a source of dietary carbohydrate. We have formulated a modified Standard American Diet (mSAD) that incorporates relevant levels and sources of nutrient classes, including dietary HFCS, to assess the basal physiologies associated with mSAD consumption. Mice proffered the mSAD for 15 weeks displayed a phenotype consistent with metabolic syndrome, exhibiting increased adiposity, fasting hyperglycemia with impaired glucose and insulin tolerance. Metabolic alterations were evidenced at the tissue level as crown-like structures (CLS) in adipose tissue and fatty acid deposition in the liver, and targeted 16S rRNA metagenomics revealed microbial compositional shifts between dietary groups. This study suggests diet quality significantly affects metabolic homeostasis, emphasizing the importance of developing relevant pre-clinical diets to investigate chronic diseases highly impacted by western dietary consumption patterns.

A pipeline of integrating transcriptome and interactome to elucidate central nodes in host-pathogens interactions.

Investigating the complexity of host-pathogen interactions is challenging. Here, we outline a pipeline to identify important proteins and signaling molecules in human-viral interactomes. Firstly, we curate a comprehensive human interactome. Subsequently, we infer viral targets and transcriptome-specific human interactomes (VTTSHI) for papillomavirus and herpes viruses by integrating viral targets and transcriptome data. Finally, we reveal the common and shared nodes and pathways in viral pathogenesis following network topology and pathway enrichment analyses. For complete details on the use and execution of this protocol, please refer to Kumar et al. (2020).

Sparking a sulfur war between plants and pathogens.

The biochemical versatility of sulfur (S) lends itself to myriad roles in plant-pathogen interactions. This review evaluates the current understanding of mechanisms by which pathogens acquire S from their plant hosts and highlights new evidence that plants can limit S availability during the immune responses. We discuss the discovery of host disease-susceptibility genes related to S that can be genetically manipulated to create new crop resistance. Finally, we summarize future research challenges and propose a research agenda that leverages systems biology approaches for a holistic understanding of this important element's diverse roles in plant disease resistance and susceptibility.

A genome-wide comparative evolutionary analysis of zinc finger-BED transcription factor genes in land plants.

Zinc finger (Zf)-BED proteins are a novel superfamily of transcription factors that controls numerous activities in plants including growth, development, and cellular responses to biotic and abiotic stresses. Despite their important roles in gene regulation, little is known about the specific functions of Zf-BEDs in land plants. The current study identified a total of 750 Zf-BED-encoding genes in 35 land plant species including mosses, bryophytes, lycophytes, gymnosperms, and angiosperms. The gene family size was somewhat proportional to genome size. All identified genes were categorized into 22 classes based on their specific domain architectures. Of these, class I (Zf-BED_DUF-domain_Dimer_Tnp_hAT) was the most common in the majority of the land plants. However, some classes were family-specific, while the others were species-specific, demonstrating diversity at different classification levels. In addition, several novel functional domains were also predicated including WRKY and nucleotide-binding site (NBS). Comparative genomics, transcriptomics, and proteomics provided insights into the evolutionary history, duplication, divergence, gene gain and loss, species relationship, expression profiling, and structural diversity of Zf-BEDs in land plants. The comprehensive study of Zf-BEDs in Gossypium sp., (cotton) also demonstrated a clear footprint of polyploidization. Overall, this comprehensive evolutionary study of Zf-BEDs in land plants highlighted significant diversity among plant species.

Combined inhibition of BET bromodomain and mTORC1/2 provides therapeutic advantage for rhabdomyosarcoma by switching cell death mechanism.

Aberrant activation of multiple complex signaling pathways underlies the pathogenesis of rhabdomyosarcoma (RMS), which remains a cause of mortality in approximately 30% of children with RMS. Bromodomain and extraterminal (BET) domain chromatin remodeling regulates several of these pathways. Here, we targeted bromodomain 4 (BRD4) in combination with another molecular metabolic tumor driver, the Akt/mTOR signaling pathway, to provide a highly effective treatment for this neoplasm. We demonstrated that a nexus of these two molecular pathways underlies RMS pathogenesis. Our data show that the combined inhibition of the BET bromodomain and mTORC1/2 signaling abrogates aggressive RMS growth. Thus, the bromodomain inhibitor RVX-208 significantly augmented the therapeutic effects of the dual mTORC1/2 inhibitors, OSI-027 and PP242, both in vitro and in a human xenograft murine model. Drug-treated residual tumors showed a decrease in the activation of underlying signaling mechanisms characterized by a reduction in the expression of p-AKT, p-mTOR, p-p70S6K, cyclin D1, and proliferation. Our ChIP-seq data demonstrated that RVX-208 effectively blocked BRD4 occupancy on its target promoters. ChIP-qPCR assays further confirmed that RVX-208 treatment resulted in a significant decrease in H3K27ac and H4K8ac signals at their target loci. While single RVX-208 treatment induces apoptosis and a single mTORC1/2 inhibitor induces macropinocytosis, their combined treatment led to necroptosis-mediated cell death. These data suggest that combined treatment with drugs targeting BRD4 and mTORC1/2 may be an effective therapeutic intervention for drug-resistant RMS.

Genome wide study of cysteine rich receptor like proteins in Gossypium sp.

Cysteine-rich receptor-like-kinases (CRKs), a transmembrane subfamily of receptor-like kinase, play crucial roles in plant adaptation. As such cotton is the major source of fiber for the textile industry, but environmental stresses are limiting its growth and production. Here, we have performed a deep computational analysis of CRKs in five Gossypium species, including G. arboreum (60 genes), G. raimondii (74 genes), G. herbaceum (65 genes), G. hirsutum (118 genes), and G. barbadense (120 genes). All identified CRKs were classified into 11 major classes and 43 subclasses with the finding of several novel CRK-associated domains including ALMT, FUSC_2, Cript, FYVE, and Pkinase. Of these, DUF26_DUF26_Pkinase_Tyr was common and had elevated expression under different biotic and abiotic stresses. Moreover, the 35 land plants comparison identified several new CRKs domain-architectures. Likewise, several SNPs and InDels were observed in CLCuD resistant G. hirsutum. The miRNA target side prediction and their expression profiling in different tissues predicted miR172 as a major CRK regulating miR. The expression profiling of CRKs identified multiple clusters with co-expression under certain stress conditions. The expression analysis under CLCuD highlighted the role of GhCRK057, GhCRK059, GhCRK058, and GhCRK081 in resistant accession. Overall, these results provided primary data for future potential functional analysis as well as a reference study for other agronomically important crops.

Ex Vivo Culture Models of Hidradenitis Suppurativa for Defining Molecular Pathogenesis and Treatment Efficacy of Novel Drugs.

Hidradenitis suppurativa (HS) is a complex and debilitating inflammatory skin disease for which no effective treatment is available currently. This is partly because of the lack of adequate human or animal models for defining the pathobiology of the disease. Here, we describe the development of air-liquid (A-L) interface, liquid-submersion (L-S), and bioreactor (Bio) ex vivo skin culture models. All three ex vivo platforms were effective for culturing skin samples for up to 14 days. Tissue architecture and integrity remained intact for at least 3 days for healthy skin and 14 days for HS skin. Up to day 3, no significant differences were observed in % early apoptotic cells among all three platforms. However, late apoptotic/necrotic cell death was increased in HS skin at day 3 in A-L and Bio culture. These cultures efficiently support the growth of various cells populations, including keratinocytes and immune cells. Profiling inflammatory gene signatures in HS skin from these ex vivo cultures showed dynamic changes in expression at day 3 and day 14. All three culture platforms were necessary to represent the inflammatory gene status of HS skin at day 0, suggesting that not all gene clusters were identically altered in each culture method. Similarly, cytokine/chemokine profiling of the supernatants from vehicle- and drug-treated ex vivo HS cultures again showed a better prediction of drug efficacy against HS. Overall, development of these three culture systems collectively provides a powerful tool to uncover the pathobiology of HS progression and screen various drugs against HS.

A TIReless battle: TIR domains in plant-pathogen interactions.

Toll/interleukin-1 receptor (TIR) domain-containing proteins are conserved across kingdoms, and their mechanistic understanding holds promise for basic plant biology and agriculture. Here, we discuss the novel enzymatic TIR domain functions of nucleotide-binding leucine-rich repeat receptors (NLRs) in cell death, and posit how TIR domain-containing effectors mechanistically subvert host immune systems.

Advances in molecular pathogenesis of hidradenitis suppurativa: Dysregulated keratins and ECM signaling.

Hidradenitis suppurativa (HS) is characterized by deep-seated, highly inflamed, and painful lumps/abscesses, fistulae, and sinus tracts that grow extensively deep in the dermis and are highly immunogenic in nature. In about one-third of the HS patients there is strong evidence for the role of γ-secretase mutations along with dysregulated Notch signaling. However, the contribution of dysregulated Notch signaling in HS pathogenesis in relation to hair follicle alterations and hyper-activation of the immune system remains undefined. A genome-wide association study (GWAS), proteomic data and functional investigations of identified sequence variants in HS pathology are not fully revealing. The disease initiation or progression may involve bacterial infection besides intrinsic functional defects in keratinocytes, which may be key to further exacerbate immune cell infiltration and cytokine production in and around the lesional tissue. The absence of a suitable animal model that could fully recapitulate the pathogenesis of HS is a major impediment for proper understanding the underlying mechanisms and development of effective treatments. The presence of extracellular matrix (ECM) degradation products along with dysregulation in keratinocytes and, dermal fibroblasts ultimately affect immune regulation and are various components of HS pathogenesis. Bacterial infection further exacerbates the complexity of the disease progression. While anti-TNFα therapy shows partial efficacy, treatment to cure HS is absent. Multiple clinical trials targeting various cytokines, complement C5a and ECM products are in progress. This review provides state-of-the-art information on these aspects with a focus on dysregulated keratinocyte and immune cells; and role of ECM, and Keratin functions in this regard.