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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. 

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. 

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 inGossypiumsp., (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.

 

The unfolded protein response (UPR) is an adaptive eukaryotic reaction that controls the protein folding capacities of the endoplasmic reticulum (ER). The most ancient and well-conserved component of the UPR is Inositol-Requiring Enzyme 1 (IRE1). Arabidopsis IRE1a (AtIRE1) is a transmembrane sensor of ER stress equipped with dual protein kinase and ribonuclease (RNase) activities, encoded by its C-terminal domain. In response to both physiological stresses and pathological perturbations, AtIRE1a directly cleaves bZIP60 ( basic leucine zipper 60 ) mRNA. Here, we developed a quantitative in vitro cleavage assay that combines recombinant AtIRE1a protein that is expressed in Nicotiana benthamiana and total RNA isolated from Arabidopsis leaves. Wild-type AtIRE1a as well as its variants containing point mutations in the kinase or RNase domains that modify its cleavage activity were employed to demonstrate their contributions to cleavage activity levels. We show that, when exposed to total RNA in vitro , the AtIRE1a protein cleaves bZIP60 mRNA. Depletion of the bZIP60 transcript in the reaction mixture can be precisely quantified by a qRT-PCR-mediated assay. This method facilitates the functional studies of novel plant IRE1 variants by allowing to quickly and precisely assess the effects of protein mutations on the substrate mRNA cleavage activity before advancing to more laborious, stable transgenic approaches in planta . Moreover, this method is readily adaptable to other plant IRE1 paralogs and orthologs, and can also be employed to test additional novel mRNA substrates of plant IRE1, such as transcripts undergoing degradation through the process of regulated IRE1-dependent decay (RIDD). Finally, this method can also be modified and expanded to functional testing of IRE1 interactors and inhibitors, as well as for studies on the molecular evolution of IRE1 and its substrates, providing additional insights into the mechanistic underpinnings of IRE1-mediated ER stress homeostasis in plant tissues. 

Abstract The environmental effects shape genetic changes in the individuals within plant populations, which in turn contribute to the enhanced genetic diversity of the population as a whole. Thus, individuals within the same species can acquire and accumulate genetic differences in their genomes depending on their local environment and evolutionary history. IRE1 is a universal endoplasmic reticulum (ER) stress sensor that activates an evolutionarily conserved signalling cascade in response to biotic and abiotic stresses. Here, we selected nine different Arabidopsis accessions along with the reference ecotype Columbia-0, based on their geographical origins and differential endogenous IRE1 expression under steady-state conditions to investigate the natural variation of ER stress responses. We cloned and analysed selected upstream regulatory regions of IRE1a and IRE1b , which revealed differential levels of their inducibility. We also subjected these accessions to an array of biotic and abiotic stresses including heat, ER stress-inducing chemical tunicamycin, phytohormone salicylic acid, and pathogen infection. We measured IRE1-mediated splicing of its evolutionarily conserved downstream client as well as transcript accumulation of ER-resident chaperones and co-chaperones. Collectively, our results illustrate the expression polymorphism of a major plant stress receptor and its relationship with molecular and physiological ER stress sensitivity.