Multidrug resistance (MDR) poses a significant obstacle to developing chemotherapeutic treatments. In previous studies using a traditional model of adriamycin resistance (ADR) with K562 cells, we demonstrated that N-acetylglucosaminyltransferase III (GnT-III) expression negatively regulates chemoresistance. Additionally, we observed that fucosylation levels were increased in the ADR cells.

The interaction of drugs with DNA is crucial for understanding their mechanism of action, particularly in the context of gene expression regulation. Erdafitinib (EDB), a pan-FGFR (fibroblast growth factor receptor) inhibitor approved by the FDA, is a potent anticancer agent used primarily in the treatment of urothelial carcinoma. In this study, the binding interaction between EDB and calf thymus DNA (ctDNA) was assessed using molecular docking, UV-absorption spectroscopy, fluorescence spectroscopy, and circular dichroism (CD) spectroscopy. The absorption spectra indicated a hypochromic effect when EDB was combined with ctDNA. The binding constant (K) of EDB-ctDNA complex was calculated as 7.84 × 10 M, corresponds to a free energy change (ΔG) value of approximately -5.06 kcal/mol, indicating a moderate binding affinity. Fluorometric analysis revealed a static binding mechanism in the ground state, with a bimolecular enhancement constant (K) of 7.56 × 10 M. Displacement experiments demonstrated that EDB preferentially binds to the minor groove of ctDNA, with a Ksv value of 5.14 × 10 M. Further, KI quenching and CD spectroscopy confirmed the minor groove binding mode, which was associated with a decrease in the T from 68.28 °C to 65.84 °C, reflecting a destabilizing effect on DNA helix. Molecular docking supported these findings, showing that EDB exhibits a strong affinity for the minor groove of ctDNA and hydrogen bonding and Vander Waal interactions are the major forces involved in the binding. These results suggest that EDB primarily binds to the minor groove of ctDNA, which may play a role in its anticancer activity.

Peptidyl prolyl cis/trans isomerases (PPIases), a ubiquitously distributed superfamily of enzymes, associated with signal transduction, trafficking, assembly, biofilm formation, stress tolerance, cell cycle regulation, gene expression and tissue regeneration, is a key regulator of metabolic disorders and microbial virulence. This review assumes an integrative approach, to provide a holistic overview of the structural and functional diversity of PPIases, examining their conformational dynamics, cellular distribution, and physiological significance. We explore their intricate involvement in cellular processes and virulence modulation in both eukaryotic and prokaryotic systems. Additionally, we evaluate the potential of these molecular chaperones as drug targets and vaccine candidates, emphasizing their relevance in therapeutic development. By synthesizing recent findings and providing a broader perspective on these proteins, this review aims to enhance our understanding of their multifaceted roles in biology and their potential applications in medicine.

The abiotic stress tolerance mechanism in plants is regulated by multiple physiological, biochemical, and molecular processes; hence, omics approaches to underpin these mechanisms are essential. It is clear that transcription factors (TFs) are one of the fundamental molecular switches that play a crucial role in modulating, regulating, and orchestrating plants in response to various climatic vagaries. Several reports are available now, focusing on understanding the roles of TFs, including those in Poaceae family in regulating different biological processes and stress responses. However, research on bamboo TFs' regulatory role in providing abiotic stress tolerance is limited. Hence the present review offers innovative insights into unraveling the molecular regulation of known family of TFs in different species of bamboo which have been identified as regulators of transcript abundance in numerous genes responsive to various abiotic stresses. Additionally, this review highlights recent discoveries concerning bamboo TFs, encompassing their classification, promoter analysis and functional dynamics in response to different abiotic stresses. Attempt has also been made to delve into the molecular interplay and cross-talk among these TFs during abiotic stresses, thus proposing potential strategies for enhancing the intricate regulatory networks involved in the adaptive responses of bamboo species.

The gating process of voltage-gated sodium (Na) channels is extraordinary intrinsic and involves numerous factors, such as voltage-sensing domain (VSD), the N-terminus and C-terminus, and the auxiliary subunits. To date, the gating mechanism of Na channel has not been clearly elucidated. Na1.9 has garnered significant attention due to its slow gating kinetics. Due to the challenges of Na1.9 heterologous expression, research on its gating mechanism is relatively limited. Whether there are any differences in the functions of the four VSDs in Na1.9 compared to those in other subtypes remains an open question. Here, we employed the established chimera method to transplant the S3b-S4 motif from the VSDIV of the toxin-sensitive donor channel (Na1.9) into the receptor channel (Na1.9/1.8 DIV S3b-S4 chimera). This modification imparted animal toxin sensitivity to the other three VSDs. Our results demonstrate that all four VSDs of Na1.9 are involved in channel opening, VSDIII and VSDIV are primarily involved in regulating fast inactivation, and VSDII also regulates the steady-state inactivation of channels. These findings provide a new insight into the gating mechanism of Na1.9.

In viviparous black rockfish (Sebastes schlegelii), the kidney of reproductive-phase males actively produces lipocalin-type prostaglandin D synthase homolog (LPGDSh) protein, which is presumably involved in inter-sexual communication when emitted in the urine. The present study was undertaken to discover whether androgens and their nuclear receptors (Ars) are engaged in regulation of renal LPGDSh protein synthesis in black rockfish. Quantitative real-time polymerase chain reaction, in conjunction with immunohistochemistry and highly sensitive enzyme-linked immunosorbent assay, revealed that intra-abdominal administration of a synthetic androgen, 17α-methyltestosterone (MT), to juvenile black rockfish induced their renal expression of LPGDSh transcript and protein. In situ hybridization visualized arα and arβ transcripts in the renal tubules of mature males during the copulation season, where they were co-localized with LPGDSh protein. Androgens, such as 11β-hydroxytestosterone, MT, dihydrotestosterone, 11-ketotestosterone (11KT), testosterone, and androstenedione transactivated a luciferase reporter vector containing four repeats of a consensus androgen response element (ARE) in the presence of black rockfish Ars (either Arα or Arβ), with differences in ligand-preference and dose-response profiles being observed between the two Ars. In the presence of 11KT, the Ars transactivated a reporter vector containing the proximal 5'-flanking region of an LPGDSh gene in luciferase reporter assays. The region between 2100 bp and 1110 bp upstream from the start codon of the LPGDSh gene, wherein many ARE-like motifs are densely distributed, was imperative for the androgenic transactivation response of the 5'-flanking region. Collectively, these observations verify that renal synthesis of LPGDSh protein is upregulated by androgens.

Microcrystal electron diffraction (MicroED) is an emerging method for the structure determination of proteins and peptides, enzyme-inhibitor complexes. Several structures of biomolecules, including lysozyme, proteinase K, adenosine receptor A2A, insulin, xylanase, thermolysin, DNA, and Granulovirus occlusion bodies, have been successfully determined through MicroED. As MicroED uses very small crystals for structure determination, therefore, it has several advantages over conventional X-ray diffraction methods. In this review article, we discussed the most recent developments in the field of MicroED and its applications for the structural determination of different types of peptides, proteins, enzymes, DNA, and enzyme-inhibitor-complexed structures.

Finger millet, a C plant with mesophyll and bundle sheath cells, has been cultivated at high altitudes in the Himalayas owing to its adaptability to stressful environments. Under environmental stresses such as high light and drought, finger millet mesophyll chloroplasts move toward the bundle sheath, a phenomenon known as aggregative arrangement.

This study aimed to compare and evaluate the growth inhibition effects of eight previously synthesized compounds, cis-3,4-diaryl-α-methylene-γ-butyrolactams (compounds 1-8), on two human renal carcinoma cell (RCC) lines: CRL-1932 (rapid growth) and HTB-44 (slow growth). MTT assays and flow cytometry were conducted, revealing that compounds 5 and 6 had the potential to induce cell death in the slow-growing RCC cells (HTB-44), while compound 8 demonstrated effectiveness in both RCC lines (HTB-44 and CRL-1932). Additionally, a non-transformed HEK293 cell line and a transgenic zebrafish with a green fluorescent kidney Tg(wt1b:egfp) were used to assess the toxicities of compounds 5, 6, and 8. The findings suggested that compound 8 was relatively non-toxic compared to the others. Western blot analysis indicated that compounds 5, 6, and 8 may interact with the P53/mTOR pathways. Based on these results, we concluded that compound 8 exhibits RCC growth inhibition properties and has lower toxicity, making it a candidate for further investigation in mammalian models.

Three-dimensional(3D) cell culture systems provide a larger space for cell proliferation, which is crucial for simulating cellular behavior and drug responses in the tumor microenvironment. In this study, we developed a novel 3D co-culture system for cell interactions, utilizing a commercialized bioreactor-microcarrier system. Mesenchymal stem cells (MSCs) were extracted via enzymatic digestion, and markers CD105 and CD31 were identified. Cell growth was observed using AO and immunofluorescence staining. No significant differences in Ki67 and GFAP expression were found between 2D and 3D cultures, though the 3D system offered more space for proliferation and reduced contact inhibition. Therefore, this 3D culture system may represent the tumor microenvironment more accurately than 2D cultures and will facilitate the investigation of the characteristics and functions of exosomes derived from this system. Exosomes are nanoscale vesicles that mediate intercellular communication by transferring molecules such as miRNAs between cells. Exosomes from 3D cultures were collected via ultra-high-speed centrifugation and characterized using nano-flow cytometry, transmission electron microscopy, and western blotting for markers CD9, Alix, and TSG101. PKH26 staining revealed peak exosome uptake by tumor cells at 24 h and complete metabolism by 72 h. Exosomes from 3D cultures inhibited GBM cell proliferation, migration, and invasion. Lastly, miRNA sequencing of exosomes was performed. This study emphasizes the importance of creating 3D co-culture systems to advance cancer research and offers a helpful tool for studying the complex cell interaction environment of GBM and other malignancies.

In vitro and ex vivo studies on drug metabolism and stability are vital for drug development and pre-clinical safety assessment. Traditional in vitro models, such as liver enzyme (S9) fractions and microsomes, often fail to account for individual variability. Personalized models, including 3D cell models and organoids, offer promising alternatives but may not fully replicate physiological processes, especially for Cytochrome P450 (CYP) families involved in extrahepatic metabolism. A major challenge in these studies is the low stability and expression of CYP enzymes. This study aimed to stabilize native CYP activity in vitro by developing an optimized buffer formulation. Initial experiments using recombinant CYP supersomes and liver microsomes identified 45 μM cysteine, 4 mM dithiothreitol (DTT), and 300 μM phosphocholine (PC) as the most effective stabilizers. The applicability of these stabilizers was subsequently confirmed in primary human brain tissue, where they enabled the successful determination of CYP2D6 activity. This highlights the stabilizing buffer's utility for enhancing CYP functionality in diverse tissue types, including the brain, which plays a critical role in cerebral detoxification and drug metabolism. These findings suggest that specific enzyme stabilization can enable comprehensive evaluations of CYP function in ex vivo tissue samples, advancing the development of organoid human tissue models and supporting drug metabolism research.

Postprandial hyperglycemia induces expression of inflammatory cytokines including tumor necrosis factor (TNF), which promotes the onset of type 2 diabetes and cardiovascular diseases. In this study, we investigated whether a transient high-glucose culture enhanced sustained expression of TNF, or whether the induction is associated with histone acetylation, and bromodomain protein containing protein 4 (BRD4), which binds acetylated histone, in human juvenile macrophage-like THP-1 cells.

Ladderanes are highly strained hydrocarbons consisting of two or more linearly concatenated cyclobutane rings. Strikingly, ladderane moieties are part of unique fatty acids and fatty alcohols that are exclusively found in the membrane lipids of anaerobic ammonium-oxidizing (anammox) bacteria. These bacteria express a distinctive gene cluster (cluster I) that has been suggested to be responsible for ladderane fatty acid (FA) biosynthesis in addition to a cluster likely involved in canonical FA biosynthesis (cluster III). In the anammox organism Kuenenia stuttgartiensis, cluster I encodes a unique acyl carrier protein (amxACP), whereas the ACP encoded by cluster III (KsACPII) was suggested to be involved in the production of canonical fatty acids. Here we present targeted isotope labeling studies using C-malonyl-ACPs to distinguish the roles of these ACPs. While in-vitroC incorporation into ladderane FAs was not observed, we show that KsACPII indeed functions in palmitate biosynthesis in the anammox organism Kuenenia stuttgartiensis. We present an experimental framework for continuing studies into fatty acid biosynthesis in anammox- and similar organisms.

Biomolecular condensates like U-bodies are specialized cellular structures formed through multivalent interactions among intrinsically disordered regions. U-bodies sequester small nuclear ribonucleoprotein complexes (snRNPs) in the cytoplasm, and their formation in mammalian cells depends on stress conditions. Because of their location adjacent to P-bodies, U-bodies have been considered potential sites for snRNP storage or turnover. SMN, a chaperone for snRNP biogenesis, forms condensates through its Tudor domain. In fly models, defects in SMN trigger innate immune responses similar to those observed with excess cytoplasmic snRNA during viral infection in mammalian cells. Additionally, spinal muscular atrophy (SMA), caused by SMN deficiency, is associated with inflammation. Therefore, SMN may help prevent innate immune aberrant activation due to defective snRNP biogenesis by forming U-bodies to sequester these molecules. Further studies on U-body functions may provide therapeutic insights for diseases related to RNA metabolism.

Metabolic dysfunction-associated steatotic liver disease (MASLD) covers a range of liver conditions marked by the buildup of fat, spanning from simple fatty liver to more advanced stages like metabolic dysfunction-associated steatohepatitis and cirrhosis.

Protein kinase C (PKC) signalling has been shown to be dysregulated in various cancers including acute lymphoblastic leukemia (ALL). We have previously determined that changes in the expression levels of SLC43A3-encoded equilibrative nucleobase transporter 1 (ENBT1) can significantly alter 6-mercaptopurine (6-MP) toxicity in ALL cells. 6-MP is a common drug used in ALL chemotherapy. Furthermore, it has been reported that activation of PKC by phorbol 12-myristate 13-acetate (PMA) impacts nucleobase uptake via an ENBT1-like transporter in Lilly Laboratories Culture-Porcine Kidney 1 (LLC-PK1) cells. We hypothesized that activation of PKC would also alter ENBT1-mediated uptake of nucleobases in leukemia cell models. Using MOLT-4, SUP-B15, and K562 cells, we incubated the cells with PMA or its inactive isoform 4α-PMA for 30 min and determined changes to ENBT1-mediated substrate uptake. All of the cell lines tested showed decreased ENBT1-mediated substrate uptake when exposed PMA, relative to that observed using 4α-PMA. Pre-incubation with the broad-spectrum PKC inhibitor, Gö6983, reversed the decrease caused by PMA. Finally, to determine the residue responsible for this PKC-mediated effect, we transiently transfected HEK293 cells (which do not express endogenous ENBT1) with wild-type SLC43A3 transcript or constructs mutated to modify the predicted PKC sites in ENBT1. We found that the mutation of threonine 231 to alanine prevents the decrease in ENBT1-mediated uptake following incubation with PMA, suggesting its involvement. This study shows that activation of PKC decreases ENBT1-mediated uptake, suggesting that aberrant activation of PKC in ALL could decrease ENBT1-mediated 6-MP uptake potentially leading to decreased therapeutic efficacy.

The diversity of molecular entities emerging from a single gene are recognized. Several studies have thus established the cellular role(s) of transcript variants and protein isoforms. A step ahead in challenging the central dogma towards expanding molecular diversity is the identification of fusion genes, chimeric transcripts and chimeric proteins that harbor sequences from more than one gene. The mechanisms for generation of chimeras largely follow similar patterns across all levels of gene regulation but also have interdependence and mutual exclusivity. Whole genome and RNA-seq technologies supported by development of computational algorithms and programs for processing datasets have increasingly enabled the identification of fusion genes and chimeric transcripts, while the discovery of chimeric proteins is as yet more subtle. Earlier thought to be associated with cellular transformation, the contribution of chimeric molecules to normal physiology is also realized and found to influence the expression of their parental genes and regulate cellular pathways. This review offers a collective and comprehensive overview of cellular chimeric entities encompassing the mechanisms involved in their generation, insights on their evolution, functions in gene regulation and their current and novel clinical applications.

Aberrant glycosylation has been implicated in promoting the progression and metastasis of pancreatic ductal adenocarcinoma (PDAC). However, the contribution of different glycosylation-related genes in PDAC remains to be clarified. In this study, we performed a differential analysis of RNA-Seq data from TCGA and GTEx and found GALNT5 as the most significant upregulated glycosylation-related gene in PDAC. Using publicly available single-cell sequencing data, we further revealed that GALNT5 is predominantly expressed in malignant ductal epithelial cells of PDAC. Correlation analysis indicated that GALNT5 is the essential member of the GALNT family associated with poor prognosis of PDAC. Overexpression of GALNT5 in PANC-1 or MIAPaCa-2 cells with low endogenous GALNT5 enhances migration and invasion. Conversely, knockdown of GALNT5 in AsPC-1 cells with high endogenous GALNT5 inhibits migration and invasion. Mechanistically, we discovered that GALNT5 activates the Erk signaling pathway in PDAC. Our findings suggest GALNT5 is a potential therapeutic target for PDAC.

Conformational switching in RNA binding proteins (RBPs) is crucial for regulation of RNA processing and transport. Dysregulation or mutations in RBPs and broad RNA processing abnormalities are related to many human diseases including neurodegenerative disorders. Here, we review the role of protein-RNA conformational switches in RBP-RNA complexes. RBP-RNA complexes exhibit wide range of conformational switching depending on the RNA molecule and its ability to induce conformational changes in its partner RBP. We categorize the conformational switches into three groups: rigid body, semi-flexible and full flexible. We also investigate conformational switches in large cellular assemblies including ribosome, spliceosome and RISC complexes. In addition, the role of intrinsic disorder in RBP-RNA conformational switches is discussed. We have also discussed the effect of different disease-causing mutations on conformational switching of proteins associated with neurodegenerative diseases. We believe that this study will enhance our understanding on the role of protein-RNA conformational switches. Furthermore, the availability of a large number of atomic structures of RBP-RNA complexes in near future would facilitate to create a complete repertoire of human RBP-RNA conformational switches.

Apolipoprotein E (apoE) polymorphism is associated with different pathologies such as atherosclerosis and Alzheimer's disease. Knowledge of the three-dimensional structure of apoE and isoform-specific structural differences are prerequisites for the rational design of small molecule structure modulators that correct the detrimental effects of pathological isoforms. In this study, cross-linking mass spectrometry (XL-MS) targeting Asp, Glu and Lys residues was used to explore the intramolecular interactions in the E2, E3 and E4 isoforms of apoE. The resulting quantitative XL-MS data combined with molecular modeling revealed isoform-specific characteristics of the N- and C-terminal domain interfaces as well as the isoform-dependent dynamic equilibrium of these interfaces. Finally, the data identified a network of salt bridges formed by R61-R112-E109 residues in the N-terminal helical bundle as a modulator of the interaction with the C-terminal domain making this network a potential drug target.