Molecules and Cells最新文献

Exposure to blue light can lead to retinal degeneration, causing adverse effects on eye health. Although the loss of retinal cells due to blue light exposure has been observed, the precise molecular mechanisms underlying this process remain poorly understood. In this study, we investigate the role of alpha-crystallin A (CRYAA) in neuro-retinal degeneration and their regulation by blue light. We observed significant apoptotic cell death in both the retina of rats and the cultured neuro-retinal cells. The expressions of Cryaa mRNA and protein were significantly downregulated in the retina exposed to blue light. We identified that miR-325-3p reduces Cryaa mRNA and protein by binding to its 3′-untranslated region. Upregulation of miR-325-3p destabilized Cryaa mRNA and suppresses CRYAA, whereas downregulation of miR-325-3p increased both expressions. Blue light-induced neuro-retinal cell death was alleviated by CRYAA overexpression. These results highlight the critical role of Cryaa mRNA and miR-325-3p molecular axis in blue light-induced retinal degeneration. Consequently, targeting CRYAA and miR-325-3p presents a potential strategy for protecting against blue light-induced retinal degeneration.

There has been growing emphasis on environmental pollutants, including heavy metals, pesticides, and nanoplastics, owing to the escalating significance of environmental pollution as a major global issue. Various toxicities induced by these compounds have been consistently reported, and many cell lines and animal models have been used in toxicity studies. Zebrafish are one of the most widely used animal models for verifying the toxic effects of environmental pollutants, owing to their many advantages. In this study, we provide brief guidelines for zebrafish maintenance and mating methods, toxicant treatments, survival measurements, and morphological abnormalities.

FB_MR5 is a nucleotide-binding domain and leucine-rich repeat protein identified from wild apple species Malus × robusta 5 conferring disease resistance to bacterial fire blight. FB_MR5 (hereafter MrMR5) recognizes the cysteine protease effector EaAvrRpt2 secreted from the causal agent of bacterial fire blight, Erwinia amylovora. We previously reported that MrMR5 is activated by the C-terminal cleavage product (ACP3) of Malus domestica RIN4 (MdRIN4) produced by EaAvrRpt2-directed proteolysis. We show that MbMR5 from a wild apple species Malus baccata shares 99.4% amino acid sequence identity with MrMR5. Surprisingly, transient expression of MbMR5 in Nicotiana benthamiana showed autoactivity in contrast to MrMR5. Domain swap and mutational analyses revealed that 1 amino acid polymorphism in the MbMR5 CC domain is critical in enhancing autoactivity. We further demonstrated that MrMR5 carrying 7 amino acid polymorphisms present in MbMR5 is not activated by MdRIN4 ACP3 but recognizes AvrRpt2 without MdRIN4 in N. benthamiana. Our findings indicate that naturally occurring polymorphisms of MR5 natural variants can confer its cell death-inducing activity and the effector recognition mechanism likely due to altered compatibility with RIN4.

Metabolic networks are fundamental to cellular processes, driving energy production, biosynthesis, redox regulation, and cellular signaling. Recent advancements in metabolic research tools have provided unprecedented insights into cellular metabolism. Among these tools, the extracellular flux analyzer stands out for its real-time measurement of key metabolic parameters: glycolysis, mitochondrial respiration, and fatty acid oxidation, leading to its widespread use. This review provides a comprehensive summary of the basic principles and workflow of the extracellular flux assay (the Seahorse assay) and its diverse applications. We highlight the assay's versatility across various biological models, including cancer cells, immunocytes, Caenorhabditis elegans, tissues, isolated mitochondria, and three-dimensional structures such as organoids, and summarize key considerations for using extracellular flux assay in these models. Additionally, we discuss the limitations of the Seahorse assay and propose future directions for its development. This review aims to enhance the understanding of extracellular flux assay and its significance in biological studies.

Plant growth must be regulated throughout the plant life cycle. The myeloblastosis (MYB) transcription factor (TF) family is one of the largest TF families and is involved in metabolism, lignin biosynthesis, and developmental processes. Here, we showed that OsMYB14, a rice R2R3-MYB TF, was expressed in leaves and roots, especially in rice culm and panicles, and that it localized to the nucleus. Overexpression of OsMYB14 (OsMYB14-ox) in rice resulted in a 30% reduction in plant height compared to that of the wild type (WT), while the height of the osmyb14-knockout (osmyb14-ko) mutant generated using the CRISPR/Cas9 system was not significantly different. Microscopic observations of the first internode revealed that the cell size did not differ significantly among the lines. RNA sequencing analysis revealed that genes associated with plant development, regulation, lipid metabolism, carbohydrate metabolism, and gibberellin (GA) and auxin metabolic processes were downregulated in the OsMYB14-ox line. Hormone quantitation revealed that inactive GA19 accumulated in OsMYB14-ox but not in the WT or knockout plants, suggesting that GA20 generation was repressed. Indole-3-acetic acid (IAA) and IAA-aspartate accumulated in OsMYB14-ox and osmyb14-ko, respectively. Indeed, real-time PCR analysis revealed that the expression of OsGA20ox1, encoding GA20 oxidase 1, and OsGH3-2, encoding IAA-amido synthetase, was downregulated in OsMYB14-ox and upregulated in osmyb14-ko. A protein-binding microarray revealed the presence of a consensus DNA-binding sequence, the ACCTACC-like motif, in the promoters of the OsGA20ox1 and GA20ox2 genes. These results suggest that OsMYB14 may act as a negative regulator of biological processes affecting plant height in rice by regulating GA biosynthesis and auxin metabolism.

Reverse genetics offers precise functional insights into genes through the targeted manipulation of gene expression followed by phenotypic assessment. While these approaches have proven effective in model organisms such as Saccharomyces cerevisiae, large-scale genetic manipulations in human cells were historically unfeasible due to methodological limitations. However, recent advancements in functional genomics, particularly clustered regularly interspaced short palindromic repeats (CRISPR)-based screening technologies and next-generation sequencing platforms, have enabled pooled screening technologies that allow massively parallel, unbiased assessments of biological phenomena in human cells. This review provides a comprehensive overview of cutting-edge functional genomic screening technologies applicable to human cells, ranging from short hairpin RNA screens to modern CRISPR screens. Additionally, we explore the integration of CRISPR platforms with single-cell approaches to monitor gene expression, chromatin accessibility, epigenetic regulation, and chromatin architecture following genetic perturbations at the omics level. By offering an in-depth understanding of these genomic screening methods, this review aims to provide insights into more targeted and effective strategies for genomic research and personalized medicine.

Metabolic aberrations, notably deviations in glutamine metabolism, are crucial in the oncogenic process, offering vital resources for the unlimited proliferation and enhanced survival capabilities of cancer cells. The dependency of malignant cells on glutamine metabolism has led to the proposition of targeted therapeutic strategies. However, the capability of cancer cells to initiate adaptive responses undermines the efficacy of these therapeutic interventions. This review meticulously examines the multifaceted adaptive mechanisms that cancer cells deploy to sustain survival and growth following the disruption of glutamine metabolism. Emphasis is placed on the roles of transcription factors, alterations in metabolic pathways, the mechanistic target of rapamycin complex 1 signaling axis, autophagy, macropinocytosis, nucleotide biosynthesis, and the scavenging of ROS. Thus, the delineation and subsequent targeting of these adaptive responses in the context of therapies aimed at glutamine metabolism offer a promising avenue for circumventing drug resistance in cancer treatment.