Search Results (707)

As with other non-coding RNAs (ncRNAs), the aberrant expression of long non-coding RNAs (lncRNAs) can be associated with different forms of cancers, including breast cancer (BC). Various lncRNAs may either promote or suppress cell proliferation, metastasis, and other related cancer signaling pathways by interacting with other cellular machinery, thus affecting the expression of BC-related genes. However, lncRNAs are characterized by features that are unlike protein-coding genes, which pose unique challenges when it comes to their study and utility. They are highly diverse and may display contradictory functions depending on factors like the BC subtype, isoform diversity, epigenetic regulation, subcellular localization, interactions with various molecular partners, and the tumor microenvironment (TME), which contributes to the intratumoral heterogeneity and phenotypic plasticity. While lncRNAs have potential clinical utility, their functional heterogeneity coupled with a current paucity of knowledge of their functions present challenges for clinical translation. Strategies to address this heterogeneity include improving classification systems, employing CRISPR/Cas tools for functional studies, utilizing single-cell and spatial sequencing technologies, and prioritizing robust targets for therapeutic development. A comprehensive understanding of the lncRNA functional heterogeneity and context-dependent behavior is crucial for advancing BC research and precision medicine. This review discusses the sources of lncRNA heterogeneity, their implications in BC biology, and approaches to resolve knowledge gaps in order to harness lncRNAs for clinical applications. Full article

Small heat shock proteins play a pivotal role in maintaining protein homeostasis under abiotic stress conditions and are indispensable for plant viability. In the present study, a comprehensive characterization of this gene family in Allium sativum was conducted through genome-wide sequence identification, phylogenetic reconstruction, conserved motif analysis, promoter cis-element profiling, transcriptomic investigation, quantitative real-time PCR, subcellular localization, and yeast-based functional assays. A total of 114 small heat shock protein genes were identified across eight chromosomes and subsequently classified into ten phylogenetic subgroups. All encoded proteins conserved the α-crystallin domain, whereas their exon–intron architectures and promoter elements responsive to environmental stress or phytohormones exhibited considerable diversity. The predicted proteins range from 130 to 364 amino acids, with isoelectric points (pI) spanning 3.97 to 9.95 and GRAVY values from −1.131 to −0.014, indicating predominantly hydrophilic characteristics. Subcellular localization analysis revealed a broad distribution across the cytoplasm, chloroplasts, mitochondria, and other compartments, with the majority (74 proteins) localized in the cytoplasm. Synteny analysis uncovered two segmentally duplicated gene pairs (AsHSP20-80/31, and AsHSP20-81/32), both showing strong purifying selection (Ka/Ks = 0.0459 and 0.2545, respectively), suggesting functional conservation. Expression profiling demonstrated predominant transcript accumulation in bulbs and floral organs, with significant induction under heat, salinity, and jasmonic acid treatments. qRT–PCR validation further confirmed that several candidate genes, notably AsHSP20-94 and AsHSP20-79, were strongly and consistently upregulated across multiple stress conditions, underscoring their roles as core stress-responsive regulators. Subcellular localization experiments demonstrated that representative proteins are targeted to the cytoplasm, nucleus and chloroplasts. Furthermore, heterologous expression of AsHSP20-79 in yeast conferred marked thermotolerance. Collectively, these findings reveal extensive expansion and functional divergence of the small heat shock protein gene family in garlic and provide valuable candidate genes for improving stress resilience in this important crop species. Full article

IMF is a key determinant of meat quality, influencing tenderness, juiciness and flavor, yet the mechanisms underlying its formation remain poorly understood. Previous studies performed whole-genome resequencing and GWAS on pigs with divergent IMF content, identifying MCEE as a candidate gene associated with IMF deposition. Subsequently, gain- and loss-of-function approaches were employed to investigate the role of MCEE in porcine intramuscular preadipocytes. Here, we isolated primary preadipocytes and subjected them to adipogenic induction. The overexpression of MCEE enhanced the proliferation and adipogenic differentiation of porcine intramuscular preadipocytes, whereas its knockdown exerted the opposite effect. Transcriptomic analysis revealed that DEGs were primarily enriched in pathways related to oxidative phosphorylation, mitochondrial dysfunction-associated disorders and others. Subcellular localization prediction indicated mitochondrial targeting of MCEE, and its expression level influenced mitochondrial function, including reactive oxygen species levels, mitochondrial membrane potential and permeability transition pore opening. Collectively, MCEE regulates IMF deposition by modulating mitochondrial function, and these findings provide a potential molecular target for improving meat quality. Full article

Cathepsin B (CTSB), a lysosomal cysteine protease, plays pivotal roles in cellular homeostasis and pathology, including cancer progression. This study investigates the regulatory interplay between CTSB and Stefin A (STFA), an endogenous inhibitor of cysteine proteases, in renal and prostate cancer cells. Using plasmid-based overexpression and silencing systems, we demonstrated that overexpressing STFA significantly reduces CTSB activity and protein levels, while silencing STFA leads to elevated CTSB activity and expression in cancer cells but not in non-cancerous cells (embryonic kidney cells—Hek293T and endothelial cells—EA.hy926). Furthermore, STFA modulates the subcellular distribution of CTSB, with STFA overexpression reducing nuclear CTSB levels and silencing inducing cytoplasmic accumulation in cancer cells. Colocalization analysis confirms a direct interaction between STFA and CTSB, highlighting the spatial coordination necessary for effective protease inhibition. These findings underscore the critical role of the CTSB-STFA axis in maintaining proteolytic balance and suggest potential therapeutic strategies targeting this interaction in renal carcinoma and other cancers. Full article

Alzheimer’s disease (AD) is a progressive neurodegenerative disorder that poses an increasing burden on society. It is characterized by the presence of neurofibrillary tangles (NFTs) and amyloid-beta (Aβ) plaques. AD is a multifactorial disease, with one of the strongest genetic risk factors being the APOE4 allele. In this study, we investigated the impact of APOE4 on NF-κB signaling in induced pluripotent stem (iPS) cells. Our results indicate that APOE4 may influence the subcellular localization of the pluripotency marker OCT4, showing a predominantly nuclear localization in APOE4 cells, whereas it appears cytoplasmic in APOE3 cells. Additionally, NF-κB activation via its canonical subunits is blunted in APOE4 cells. Interestingly, APOE4 cells still exhibit increased transcription of key hyperinflammatory markers CCL2, CXCL10 and COX2, which are known NF-κB target genes, and exhibit a significantly higher rate of apoptosis compared to APOE3 cells—independent of TNF-α stimulation. Moreover, an elevated incidence of DNA double-strand breaks was observed in APOE4 cells. However, the precise molecular mechanisms by which APOE4 suppresses NF-κB activation while simultaneously promoting inflammation and apoptosis remain unclear. Further research is required to elucidate these underlying pathways. Full article

Background: The increasing prevalence of antibiotic-resistant Mycoplasma genitalium poses a significant challenge to global public health, necessitating the exploration of alternative therapeutic strategies, including vaccine development. Methods: In this study, we employed an immuno-informatics-based reverse vaccinology approach augmented with artificial intelligence-driven tools, to identify and characterize potential B-cell and T-cell epitopes from the hypothetical proteins (HPs) retrieved from the genome of the MG_G37T strain, a previously uncharacterized yet promising vaccine target. Using multiple softwares, a systematic pipeline was utilized to assess the sub-cellular localization, antigenicity, and allergenicity of the selected proteins. Results: Sub-cellular localization analysis identified the presence of several outer membrane and extracellular proteins in the genome of MG_G37T, indicating their surface association and accessibility to immune surveillance. Antigenicity and allergenicity prediction tools led to the identification of two top-scoring hypothetical proteins (fig|2097.71.peg.1 (UniProt ID: P22747) and fig|2097.70.peg.33 (UniProt ID: Q57081)) that demonstrated strong antigenic potential, non-allergenic properties, and suitability as vaccine candidates. Epitope mapping and structural modeling analyses further validated the immunogenic potential of these epitopes, highlighting their ability to interact with host immune components effectively. Comparative analyses with mouse allelic regions indicated the potential translational relevance of these predicted epitopes for preclinical studies. Conclusions: In particular, this study highlights the potential of these two hypothetical proteins as a promising vaccine candidate and provides a strong reason for experimental validation towards the design and development of effective vaccines to combat M. genitalium infections in the era of antimicrobial resistance. Full article

Rosa rugosa is an important ornamental and edible species that is valued for its floral colors and essential oils in the cosmetic and pharmaceutical industries. Carotenoids, beyond their health-promoting roles, function as accessory pigments that influence petal coloration, flower quality, and stress responses. However, their accumulation patterns and molecular biosynthesis in R. rugosa remain poorly understood. Here, UPLC-APCI-MS/MS analysis across three developmental stages (bud, semi-open, and full bloom) revealed stage-specific carotenoid accumulation, with phytoene and phytofluene markedly increasing at the semi-open stage. In total, 11 carotenoids were identified, comprising four carotenes and seven xanthophylls. Differential accumulation of metabolites (DAMs) analysis indicated shifts in compounds, including (E/Z)-phytoene, phytofluene, and β-carotene across stages. Genetic complementation assays in Escherichia coli and transient overexpression in rose petals confirmed that RrPSY1 functions as a phytoene synthase. qRT-PCR results showed its upregulation under salt treatment, suggesting a role in enhancing stress tolerance through carotenoid-mediated antioxidant protection. Furthermore, sub-cellular localization experiments confirmed plastid targeting of RrPSY1. Together, these findings clarify the role of RrPSY1 in carotenoid biosynthesis and provide a foundation for future studies on metabolic regulation and biosynthesis of carotenoids in R. rugosa. Full article

Nicotinamide adenine dinucleotide (NAD⁺) is an essential metabolite, and abnormal NAD⁺ metabolism has been linked to numerous human diseases. The nicotinamide mononucleotide adenylyl transferases (NMNATs) catalyze NAD⁺ production through both de novo and salvage pathways. NMNATs are multi-functional enzymes with NAD⁺ synthesis activity and chaperone activity. Interestingly, NMNATs are involved in neuroprotection, and whether these neuroprotective effects require NAD⁺ synthesis activity appears to vary depending on the context. Nevertheless, NMNATs can modulate cellular processes primarily through supporting NAD⁺ homeostasis. In this review, we discuss the roles of NMNATs in NAD⁺ homeostasis, their functional domains, and how their subcellular localizations influence the compartmentalized NAD⁺ pools. We present an integrative framework to help understand the diverse impacts of NMNATs in human diseases, with a focus on neurological disorders caused by different insults. To address knowledge gaps, we integrate the regulation of NMNATs in both human and model organisms. We also discuss the current understanding and limitations of NMNAT activators and inhibitors to help evaluate their translational significance as therapeutic targets for NAD⁺ modulation. Full article

Gastrodin is the main active ingredient of Gastrodia elata Blume, known for its prominent neuroprotective effects, especially in Alzheimer’s disease. Meanwhile, aging is a critical risk factor in age-related diseases, including AD. Now, the underlying mechanisms of gastrodin against aging-related genes in Alzheimer’s disease remain unclear. Our study aimed to identify and validate the molecular mechanisms of gastrodin on aging-related genes in Alzheimer’s disease. Firstly, we analyzed gene expression datasets from GEO in NCBI, and weighted gene co-expression network analysis (WGCNA) was used to identify the intersected genes with differentially expressed genes (DEGs) and aging genes. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) functional enrichment analyses and protein–protein interaction (PPI) network were performed to analyze and evaluate the intersected genes to screen key genes. Subsequently, we used machine learning techniques to screen hub genes and subcellular localization to confirm the reliability of the hub genes. Finally, molecular docking and molecular dynamics simulation were combined to explore the binding interactions between core targets and gastrodin. We identified 29 intersecting genes among DEGs of AD, key module genes of AD, and aging genes. Next, nine common hub genes were identified by four algorithms of the cytoHubba plug-in. Four hub genes, GFAP, NPY, SNAP25, and SST, were found as possibly hub aging-related genes in Alzheimer’s disease from machine algorithms by adopting the random forest, LASSO, SVM, and Boruta models. Among them, GFAP showed marked upregulation, while NPY, SNAP25, and SST exhibited significant downregulation (p < 0.05). Finally, molecular docking and molecular dynamics simulation exhibited that gastrodin has an excellent affinity in docking with SNAP25. This study demonstrates that SNAP25 can be considered a key aging-related gene in AD, and gastrodin could treat AD by targeting specific genes and signaling pathways. These findings provide critical insights for the clinical application of gastrodin. Full article

Neuronal chloride (Cl⁻) homeostasis is fundamental for brain function, with disruptions increasingly recognized as pathogenic across neurological disorders. This review synthesizes evidence from preclinical models and clinical studies, integrating electrophysiological measurements, molecular analyses, imaging with genetically encoded sensors like ClopHensor, and behavioral assays. Key findings demonstrate that Cl⁻ dysregulation follows distinct patterns: (1) in epilepsy, KCC2 downregulation converts GABAergic inhibition to excitation, promoting seizures; (2) in Alzheimer’s disease (AD) models, pre-symptomatic KCC2 loss in hippocampus is observed, with KCC2 restoration reversing aspects of cognitive decline; (3) in autism spectrum disorders (ASD), developmental delays in GABA polarity shifts feature due to altered NKCC1/KCC2 ratios; and (4) in Huntington’s disease (HD), striatal neuron-specific Cl⁻ imbalances are linked to motor dysfunction. Methodologically, advanced tools—including subcellular Cl⁻ imaging and high-throughput drug screening—have enabled precise dissection of these mechanisms. Therapeutic strategies targeting Cl⁻ transporters (NKCC1 inhibitors like bumetanide, KCC2 enhancers like CLP290) show preclinical promise but require improved central nervous system (CNS) delivery and selectivity. These findings establish Cl⁻ homeostasis as both a biomarker and therapeutic target, necessitating precision medicine approaches to address heterogeneity in neurological disorders. Full article

Intravital two-photon microscopy enables deep-tissue imaging of subcellular structures in live animals, but its original spatial resolution and image quality are limited by scattering, motion, and low signal-to-noise ratios. To address these challenges, we used a combination of tissue stabilization, denoising methods, and motion correction, together with resolution enhancement algorithms, including enhanced Super-Resolution Radial Fluctuations (eSRRF) and deconvolution, to acquire high-fidelity time-lapse images of internal organs. We applied this imaging pipeline to image genetically labeled mitochondria in vivo, in Dendra2 mice. Our results demonstrate that the eSRRF-combined method, compared to other evaluated algorithms, significantly shows improved spatial resolution and mitochondrial structure visualization, while each method exhibiting distinct strengths in terms of noise tolerance, edge preservation, and computational efficiency. These findings provide a practical framework for selecting enhancement strategies in intravital imaging studies targeting dynamic subcellular processes. Full article

The intracellular topography of RNA molecules, encompassing ribonucleotides with biochemical modifications, such as N6-methyladenosine (m6A), 5-methylcytosine (m5C), adenosine to inosine (A → I) editing, and isomerization of uridine to pseudouridine (Ψ), as well as of non-coding RNA molecules, is currently studied within the frame of the epigenome. Circulating RNA molecules in the intracellular space that have incorporated information by carrying specific modifications depend on the balanced activity and correct subcellular installation of their modifying enzymes, the “writers”, “readers” and “erasers”. Modifications are critical for RNA translocation from the nucleus to the cytoplasm, for stability and translation efficiency, and for other, still-uncovered functions. Moreover, trafficking of non-coding RNA molecules depends on membrane transporters capable of recognizing signal sequences and RNA recognition-binding proteins that can facilitate their transport to different intracellular locations, guiding the establishment of interconnection possibilities with different macromolecular networks. The potential of long non-coding RNAs to form multilayer molecular connections, as well as the differential topology of micro-RNAs in cell nuclei, compared to cytoplasm, has been recognized by several studies. The study of the intercellular compartmentalization of these molecules has recently become feasible thanks to technological progress; however, a wealth of information has not yet been produced that would lead to safe conclusions regarding non-coding RNA’s contributions to the early steps of pathogenesis and disease progression in hematological malignancies. Both, the bone marrow, as the main hematopoietic tissue, and the lymphoid tissues are composed of cells with highly reactive potential to signals affecting the epigenome and initiating cascade pathways in response. Independently or in combination with coexistent driver genetic mutations, especially mutations of enzymes involved in epigenomic surveillance, intracellular microenvironmental alterations within the cell nuclear, cytoplasmic, and mitochondrial compartments can lead to disorganization of hematopoietic stem cells’ epigenomes, promoting the generation of hematological malignancies. In this review, we discuss the various intracellular processes that, when disrupted, may result in the ectopic placement of RNA molecules, either inducing specific modifications or non-coding molecules or promoting hematological malignant phenotypes. The crosstalk between mitochondrial and nuclear genomes and the complex regulatory effects of mis-localized RNA molecules are highlighted. This research approach may constitute a field for new, more specifically targeted therapies in hematology based on RNA technology. Full article

Giardiasis is a globally prevalent waterborne zoonosis. Rapid enrichment and detection technologies for this disease are essential. Cyst outer wall proteins are ideal targets for the enrichment and detection of cysts in the environment, but there are few available targets with suboptimal effectiveness. In this study, Giardia duodenalis (G. duodenalis) cysts were purified, and outer wall proteins were biotinylated, followed by streptavidin magnetic bead purification and mass spectrometry. Sixty-three novel cyst wall proteins were identified, and their functions were annotated through Gene Ontology (GO) and KEGG analyses. The β-giardin and α-1 giardin were among the newly identified and predicted to be located on the outer wall of G. duodenalis cysts. For the characterization of these two targets, we applied sequence analysis, prokaryotic expression, preparation of polyclonal antibodies, and determination of subcellular localization. Finally, based on β-giardin immunomagnetic beads were prepared using the polyclonal antibodies and tested for their enrichment efficiency. Immunomagnetic beads targeting β-giardin achieved 65% cyst enrichment efficiency in fecal samples, comparable to conventional methods. Clinical evaluation across 163 multi-host fecal samples (ferrets, Siberian tigers, red-crowned cranes) demonstrated concordance with nested PCR, successfully enriching cysts from PCR-positive specimens. The immunomagnetic beads method targeting β-giardin demonstrated effective G. duodenalis cyst enrichment in multi-host fecal samples. These results provide a proteomic framework for the cyst wall proteins of G. duodenalis, expanding the detection targets for G. duodenalis cysts. It also establishes a theoretical foundation for subsequent research on the composition and function of G. duodenalis cysts. Full article

Terpenoids, which are essential pharmaceutical compounds, encounter significant production challenges due to their low yields in native plants and associated ecological concerns. This review summarizes recent advances in metabolic engineering strategies applied across three complementary platforms: native medicinal plants, microbial systems, and heterologous plant hosts. We present how the “Genomic Insights to Biotechnological Applications” paradigm, supported by multi-omics technologies such as genomics, transcriptomics, metabolomics, and related disciplines, contributes to advancing research in this field. These technologies enable the systematic identification of key biosynthetic genes and regulatory networks. CRISPR-based tools, enzyme engineering, and subcellular targeting are presented as pivotal transformative strategies in advancing metabolic engineering approaches. Strategic co-expression and optimization approaches have achieved substantial improvements in product yields, as demonstrated by a 25-fold increase in paclitaxel production and a 38% enhancement in artemisinin yield. Persistent challenges, such as metabolic flux balancing, cytotoxicity, and scale-up economics, are discussed in conjunction with emerging solutions, including machine learning and photoautotrophic chassis systems. We conclude by proposing a strategic roadmap for industrial translation that highlights the essential integration of systems biology and synthetic biology approaches to accelerate the transition of terpenoid biomanufacturing from discovery to commercial-scale application. Full article

Bacterial degradation is one important Microcystin (MC) removal method in the natural environment. The traditional MC-degrading pathway was proposed based on the functions of individual recombinant Mlr enzymes and the structures of the main MC-degrading products. However, the actual MC-degrading mechanism by Mlr enzymes in wild-type bacteria remains unclear. In this study, bioinformatic analysis, heterologous expression, and knockout mutation were performed to elaborate the MC-degrading mechanism by Mlr enzymes in Sphingopyxis sp. m6. The results showed that mlr gene cluster was initially acquired by horizontal gene transfer, followed by vertical inheritance within Alphaproteobacteria. Mlr enzymes exhibit distinct subcellular localizations and possess diverse conserved catalytic domains. The enzymatic cascade MlrA/MlrB/MlrC sequentially cleaves Microcystin-LR (MC-LR) via Adda-Arg, Ala-Leu, and Adda-Glu bonds, generating characteristic intermediates (linearized MC-LR, tetrapeptide, and Adda). Notably, recombinant MlrC demonstrated dual-targeting degrading capability (linearized MC-LR and tetrapeptide), while tetrapeptide specificity in endogenous processing of Sphingopyxis sp. m6. Marker-free knockout mutants of mlr genes were first constructed in MC-degrading bacteria, unveiling that mlrA was indispensable in initial MC cleavage, whereas mlrB/mlrC/mlrD displayed functional compensation through other enzymes with similar functions. This study promotes the mechanistic understanding of MC bacterial degradation and offers a theoretical basis for a bioremediation strategy targeting cyanotoxin pollution. Full article