Search Results (106)

Background/Objectives: Truncating mutations in PALB2, a critical component of the BRCA1-PALB2-BRCA2 homologous recombination repair complex, are associated with increased risk and aggressiveness of breast cancer. The consequences of PALB2 truncation on the expression, localization, and functional dynamics of the scaffold protein IQGAP1 were investigated in this study based on two cases of truncated PALB2 human breast invasive ductal carcinoma (IDC), specifically, c.1240C>T (p.Arg414*) and c.2257C>T (p.Arg753*). Methods: Using confocal microscopy, we examined co-expression patterns of IQGAP1 with PALB2, PCNA, CK7, and β-tubulin in tumor tissues from both control cancer and PALB2-mutated cases. Results: In PALB2-truncated tumors, IQGAP1 exhibited enhanced peripheral and plasma membrane localization with elevated co-localization levels compared to controls, suggesting altered cytoskeletal organization. PALB2 truncation increased nuclear and cytoplasmic N-terminal PALB2 immunoreactivity, indicating the presence of truncated isoforms disrupting the homologous recombination repair system. Co-expression analyses with PCNA revealed an inverse expression pattern between IQGAP1 and proliferation markers, suggesting S-phase cell cycle-dependent heterogeneity. Furthermore, the loss of IQGAP1 dominance over CK7 and β-tubulin in mutant tumors, along with persistent intercellular spacing, implied a loss of cell–cell cohesion and the acquisition of invasive traits. Conclusions: These data support a model where PALB2 truncation triggers a reorganization of IQGAP1 that disrupts its canonical structural functions and facilitates tumor progression via enhanced motility and impaired cell–cell interaction. IQGAP1 thus serves as both a functional effector and potential biomarker in PALB2-mutated IDC, opening novel paths for diagnosis and targeted therapeutic intervention. Full article

SUMOylation plays a crucial role in regulating gene expression by promoting interactions between transcription factors and corepressors. Daxx, a multifunctional scaffold protein, specifically recognizes and binds SUMOylated transcription factors through its SUMO-interacting motifs (SIMs), acting as a transcriptional corepressor. In this review, we systematically elucidate the structural basis of the interaction between Daxx and SUMO, revealing the synergistic mechanism by which Daxx SIM phosphorylation and SUMO acetylation dynamically regulate Daxx function. In promyelocytic leukemia nuclear bodies (PML NBs), phosphorylation of Daxx’s SIM enhances its binding to SUMOylated PML, leading to the sequestration and inactivation of Daxx within PML NBs. Conversely, SUMO acetylation disrupts the electrostatic interactions between SUMO and SIMs, prompting the release of Daxx from PML NBs and its translocation to the nucleoplasm, where it inhibits the activity of transcription factors such as ETS1, GR, and SMAD4. Daxx SIMs are common binding sites for the interaction between SUMOylated transcription factors and Daxx, and different SUMOylated transcription factors may compete to bind to Daxx, which cross-regulates cellular life activities. This mechanism highlights the dynamic regulation of Daxx subcellular localization and transcriptional repression by SUMO and PML NBs, providing valuable insights into understanding Daxx-mediated transcriptional repression. Full article

Background: Type 2 diabetes mellitus (T2DM) presents a significant global health challenge, with obesity being a major contributing risk factor alongside genetic and non-genetic elements. Current treatments focus on reducing hyperglycemia and preventing T2DM progression, often involving drug combinations for enhanced efficacy. This study introduces two novel nitrogen-containing heterocyclic scaffolds: neocryptolepine–thiazolidinedione (NC-TZD) 8 and acridine–thiazolidinedione (AC-TZD) 11. Methods: These compounds were synthesized and characterized using various spectroscopic techniques. Their antihyperglycemic and antihyperlipidemic effects were assessed in an obesity-induced zebrafish model. Hyperglycemia was induced by immersing zebrafish in 100 mM glucose monohydrate for two weeks. Fish were then divided into groups receiving either 20 mg or 80 mg of the drugs per kg of body weight, alongside negative and positive control groups. Results: Both doses of hybrids 8 and 11 effectively restored glucose, triglyceride, insulin, and nuclear factor kappa beta (nfκβ) mRNA levels to normal. However, only the lower doses restored peroxisomal acyl-CoA oxidase (acox1) mRNA levels, with higher doses proving less effective. A molecular modeling study supported the antidiabetic potential of hybrids 8 and 11, suggesting interactions with target proteins PPAR-α and acox1. In silico ADMET analysis revealed promising oral bioavailability and drug likeness for both compounds. Conclusions: The findings indicate that both hybrids exhibit significant antihyperglycemic and antihypertriglyceridemic effects, particularly at lower doses. These results highlight the promising therapeutic potential of these novel oral bioavailable compounds in managing T2DM. Further research is warranted to elucidate their mechanisms of action. Full article

Begonia semperflorens (B. semperflorens) is a popular ornamental plant widely used in landscapes such as plazas and flower beds, and it is also commonly grown as a potted plant indoors. It is known for its adaptability to high temperatures, drought, and shade. Under heat-tolerant conditions, heat shock transcription factors (HSFs) are key transcriptional regulatory proteins that play crucial roles in cellular processes. Despite extensive studies on the HSF family in various species, there has been no specific analysis targeting B. semperflorens. In this study, we identified 37 members of the BsHSF gene family in B. semperflorens based on its genome scaffold, which are unevenly distributed across the genome. Phylogenetic analysis reveals that these 37 members can be divided into three subfamilies. Analysis of their physicochemical properties shows significant diversity among these proteins. Except for the BsHSFB7 protein located in the cytoplasm, all other BsHSF proteins were found to be nuclear-localized. A comparison of the amino acid sequences indicates that all BsHSF proteins contain a conserved DNA-binding domain structure. Analysis of the promoter cis-acting elements also suggests that BsHSFs may be associated with heat stress and plant secondary metabolism. We further investigated the duplication events of BsHSF genes and their collinearity with genes from other Begonia species. Finally, through real-time quantitative PCR, we examined the expression patterns of the 37 BsHSFs in different plant tissues (roots, stems, leaves, and flowers) and their expression levels under heat stress treatment. The results show that, except for BsHSF29, all BsHSFs were expressed in various tissues, with varying expression levels across tissues. Except for BsHSF33 and BsHSF34, the expression levels of almost all BsHSF genes increased in response to heat treatment. In summary, these findings provide a better understanding of the role and regulatory mechanisms of HSFs in the heat stress response of B. semperflorens and lay the foundation for further exploration of the biological functions of BsHSFs in the stress responses of B. semperflorens. Full article

Rare diseases typically evade the application of the standard drug discovery and development pipelines due to their understudied molecular etiology and the small market size. Herein, we report a rare disease-directed workflow that rapidly studies the molecular features of the disorder, establishes a high-throughput screening (HTS) platform, and conducts an HTS of thousands of approved drugs to identify and validate repositioning drug candidates. This study examines the pediatric neurological disorder caused by de novo mutations in YWHAG, the gene encoding the scaffolding protein 14-3-3γ, and the workflow discovers nuclear relocalization and a severe drop in 14-3-3γ binding to its phosphorylated protein partners as the key molecular features of the pathogenic hotspot YWHAG mutations. We further established a robust in vitro HTS platform and screened ca. 3000 approved drugs to identify the repositioning drug candidates that restore the deficient 14-3-3γ-phosphotarget interactions. Our workflow can be applied to other 14-3-3-related disorders and upscaled for many other rare diseases. Full article

DNA is frequently damaged by genotoxic stresses such as ionizing radiation, reactive oxygen species, and nitrogen species. DNA damage is a key contributor to cancer initiation and progression, and thus the precise and timely repair of these harmful lesions is required. Recent studies revealed transcription as a source of genome instability, and transcription-coupled DNA damage has been a focus in cancer research. Impaired mRNA export is closely related to DNA damage through R-loop formation. The molecular machineries of transcription-coupled DNA damage have been extensively analyzed in Saccharomyces cerevisiae. However, the molecular basis of these phenomena in higher eukaryotes remains elusive. In this review, we focus on the relationship between deregulated mRNA export through the transcription-export-2 (TREX-2) complex and cancer development. Particularly, the expression of germinal center-associated nuclear protein (GANP), a molecular scaffold in the TREX-2 complex, is highly associated with tumorigenesis in mice and humans. Although the deregulated expression of other components in the TREX-2 complex might affect cancer development, we have directly demonstrated the significance of GANP in tumorigenesis using genetically modified mice. Additionally, we describe recent evidence for medical applications demonstrating that the downregulation of the other components may be a good candidate for a chemotherapeutic target in terms of reducing the side effects. Full article

The USH1G protein SANS is a small multifunctional scaffold protein. It is involved in several different cellular processes, such as intracellular transport, in the cytoplasm, or splicing of pre-mRNA, in the cell nucleus. Here, we aimed to gain insight into the regulation of the subcellular localization and the nuclear–cytoplasmic shuttling of SANS and its paralog ANKS4B, not yet reported in the nucleus. We identified karyopherins mediating the nuclear import and export by screening the nuclear interactome of SANS. Sequence analyses predicted in silico evolutionarily conserved nuclear localization sequences (NLSs) and nuclear export sequences (NESs) in SANS, but only NESs in ANKS4B, which are suitable for karyopherin binding. Quantifying the nuclear–cytoplasmic localization of wild-type SANS and NLS/NES mutants, we experimentally confirmed in silico predicted NLS and NES functioning in the nuclear–cytoplasmic shuttling in situ in cells. The comparison of SANS and its paralog ANKS4B revealed substantial differences in the interaction with the nuclear splicing protein PRPF31 and in their nuclear localization. Finally, our results on pathogenic USH1G/SANS mutants suggest that the loss of NLSs and NESs and thereby the ability to control nuclear–cytoplasmic shuttling is disease-relevant. Full article

Injectable, in situ-forming hydrogels, both biocompatible and biodegradable, have garnered significant attention in tissue engineering due to their potential for creating adaptable scaffolds. The adaptability of these hydrogels, made from natural proteins and polysaccharides, opens up a world of possibilities. In this study, sodium alginate was used to synthesize alginate di-aldehyde (ADA) through periodate oxidation, resulting in a lower molecular weight and reduced viscosity, with different degrees of oxidation (54% and 70%). The dual-crosslinking mechanism produced an injectable in situ hydrogel. Initially, physical crosslinking occurred between ADA and borax via borax complexation, followed by chemical crosslinking with gelatin through a Schiff’s base reaction, which takes place between the amino groups of gelatin and the aldehyde groups of ADA, without requiring an external crosslinking agent. The formation of Schiff’s base was confirmed by Fourier-transform infrared (FT-IR) spectroscopy. At the same time, the aldehyde groups in ADA were characterized using FT-IR, proton nuclear magnetic resonance (¹H NMR), and gel permeation chromatography (GPC), which determined its molecular weight. Furthermore, borax complexation was validated through boron-11 nuclear magnetic resonance (¹¹B NMR). The hydrogel formulation containing 70% ADA, polyethylene glycol (PEG), and 9% gelatin exhibited a decreased gelation time at physiological temperature, attributed to the increased gelatin content and higher degree of oxidation. Rheological analysis mirrored these findings, showing a correlation with gelation time. The swelling capacity was also enhanced due to the increased oxidation degree of PEG and the system’s elevated gelatin content and hydrophilicity. The hydrogel demonstrated an average pore size of 40–60 µm and a compressive strength of 376.80 kPa. The lower molecular weight and varied pH conditions influenced its degradation behavior. Notably, the hydrogel’s syringeability was deemed sufficient for practical applications, further enhancing its potential in tissue engineering. Given these properties, the 70% ADA/gelatin/PEG hydrogel is a promising candidate and a potential game-changer for injectable, self-crosslinking applications in tissue engineering. Its potential to revolutionize the field is inspiring and should motivate further exploration. Full article

Inflammatory breast cancer (IBC) is characterized by numerous tumor emboli within lymphatics. In a recent study, we observed tumor embolic budding both in vitro and in vivo within lymphovascular spaces and proposed this to account for the plethora of tumor emboli seen in IBC. These observations did not address, however, how lymphovascular invasion is initiated or the mechanisms involved. In the present study, using the well-characterized patient-derived xenograft (PDX), Mary-X, which exhibited florid lymphovascular invasion (LVI) in athymic mice (LVI) as defined by E-cadherin-positive tumor emboli within lymphatic channels distinguished by podoplanin and LYVE1 membrane and Prox1 nuclear immunoreactivities and spontaneous spheroidgenesis in vitro and human cases of IBC which showed similar LVI, we compared laser-captured microdissected emboli from Mary-X and from the cases of human IBC to non-embolic areas. Mary-X and IBC emboli expressed high levels of E-cadherin and no evidence of epithelial–mesenchymal transition (EMT). Mary-X spheroids expressed high levels of VEGF, especially VEGF-C, and stimulated both vascular and lymphatic endothelial haptotaxis. We then transplanted Mary-X serially into green, cyano, red, and nestin-green fluorescing protein (GFP-, CFP-, RFP-, and nestin-GFP) transgenic reporter mice in various combinations. Multicolor murine imaging studies indicated that reporter-labeled stroma initially encircled clumps of tumor cells and then served as a scaffold that supported nestin-GFP-labeled endothelial haptotaxis resulting in encircling lymphangiogenesis, confirmed by dual LYVE1 immunofluorescence. The present studies demonstrate a possible mechanism of a critical step of the tumor emboli formation of IBC. Full article

Conventional biochemical methods for studying cellular signaling cascades have relied on destructive cell disruption. In contrast, the live cell imaging of fluorescent-tagged transfected proteins offers a non-invasive approach to understanding signal transduction events. One strategy involves monitoring the phosphorylation-dependent shuttling of a fluorescent-labeled kinase between the nucleus and cytoplasm using nuclear localization, export signals, or both. In this paper, we introduce a simple method to visualize intracellular signal transduction in live cells by exploring the translocation properties of PKC from the cytoplasm to the membrane. We fused bait protein to PKC, allowing the bait (RFP-labeled) and target (GFP-labeled) proteins to co-translocate from the cytoplasm to the membrane. However, in non-interacting protein pairs, only the bait protein was translocated to the plasma membrane. To verify our approach, we examined the Raf–MEK–ERK signaling cascade (ERK pathway). We successfully visualized direct Raf1/MEK2 interaction and the KSR1-containing ternary complex (Raf1/MEK2/KSR1). However, the interaction between MEK and ERK was dependent on the presence of the KSR1 scaffold protein under our experimental conditions. Full article

The protein menin is encoded by the MEN1 gene and primarily serves as a nuclear scaffold protein, regulating gene expression through its interaction with and regulation of chromatin modifiers and transcription factors. While the scope of menin’s functions continues to expand, one area of growing investigation is the role of menin in cancer. Menin is increasingly recognized for its dual function as either a tumor suppressor or a tumor promoter in a highly tumor-dependent and context-specific manner. While menin serves as a suppressor of neuroendocrine tumor growth, as seen in the cancer risk syndrome multiple endocrine neoplasia type 1 (MEN1) syndrome caused by pathogenic germline variants in MEN1, recent data demonstrate that menin also suppresses cholangiocarcinoma, pancreatic ductal adenocarcinoma, gastric adenocarcinoma, lung adenocarcinoma, and melanoma. On the other hand, menin can also serve as a tumor promoter in leukemia, colorectal cancer, ovarian and endometrial cancers, Ewing sarcoma, and gliomas. Moreover, menin can either suppress or promote tumorigenesis in the breast and prostate depending on hormone receptor status and may also have mixed roles in hepatocellular carcinoma. Here, we review the rapidly expanding literature on the role and function of menin across a broad array of different cancer types, outlining tumor-specific differences in menin’s function and mechanism of action, as well as identifying its therapeutic potential and highlighting areas for future investigation. Full article

Solid tumors as well as leukemias and lymphomas show striking changes in nuclear structure including nuclear size and shape, the number and size of nucleoli, and chromatin texture. These alterations have been used in cancer diagnosis and might be related to the altered functional properties of cancer cells. The nuclear matrix (NM) represents the structural composition of the nucleus and consists of nuclear lamins and pore complexes, an internal ribonucleic protein network, and residual nucleoli. In the nuclear microenvironment, the NM is associated with multi-protein complexes, such as basal transcription factors, signaling proteins, histone-modifying factors, and chromatin remodeling machinery directly or indirectly through scaffolding proteins. Therefore, alterations in the composition of NM could result in altered DNA topology and changes in the interaction of various genes, which could then participate in a cascade of the cancer process. Using an androgen-sensitive prostate cancer cell line, LNCaP, and its androgen-independent derivative, LN96, conventional 2D-proteomic analysis of the NM proteins revealed that purine-rich element binding protein alpha (PURα) was detected in the NM proteins and differentially expressed between the cell lines. In this article, we will review the potential role of the molecule in prostate cancer. Full article

Chronic inflammation contributes to a number of diseases. Therefore, control of the inflammatory response is an important therapeutic goal. To identify novel anti-inflammatory compounds, we synthesized and screened a library of 80 pyrazolo[1,5-a]quinazoline compounds and related derivatives. Screening of these compounds for their ability to inhibit lipopolysaccharide (LPS)-induced nuclear factor κB (NF-κB) transcriptional activity in human THP-1Blue monocytic cells identified 13 compounds with anti-inflammatory activity (IC₅₀ < 50 µM) in a cell-based test system, with two of the most potent being compounds 13i (5-[(4-sulfamoylbenzyl)oxy]pyrazolo[1,5-a]quinazoline-3-carboxamide) and 16 (5-[(4-(methylsulfinyl)benzyloxy]pyrazolo[1,5-a]quinazoline-3-carboxamide). Pharmacophore mapping of potential targets predicted that 13i and 16 may be ligands for three mitogen-activated protein kinases (MAPKs), including extracellular signal-regulated kinase 2 (ERK2), p38α, and c-Jun N-terminal kinase 3 (JNK3). Indeed, molecular modeling supported that these compounds could effectively bind to ERK2, p38α, and JNK3, with the highest complementarity to JNK3. The key residues of JNK3 important for this binding were identified. Moreover, compounds 13i and 16 exhibited micromolar binding affinities for JNK1, JNK2, and JNK3. Thus, our results demonstrate the potential for developing lead anti-inflammatory drugs based on the pyrazolo[1,5-a]quinazoline and related scaffolds that are targeted toward MAPKs. Full article

Hepatocellular carcinoma (HCC) is a heterogeneous malignancy with complex carcinogenesis. Although there has been significant progress in the treatment of HCC over the past decades, drug resistance to chemotherapy remains a major obstacle in its successful management. In this study, we were able to reduce chemoresistance in cisplatin-resistant HepG2 cells by either silencing the expression of transglutaminase type 2 (TG2) using siRNA or by the pre-treatment of cells with the TG2 enzyme inhibitor cystamine. Further analysis revealed that, whereas the full-length TG2 isoform (TG2-L) was almost completely cytoplasmic in its distribution, the majority of the short TG2 isoform (TG2-S) was membrane-associated in both parental and chemoresistant HepG2 cells. Following the induction of cisplatin toxicity in non-chemoresistant parental cells, TG2-S, together with cisplatin, quickly relocated to the cytosolic fraction. Conversely, no cytosolic relocalisation of TG2-S or nuclear accumulation cisplatin was observed, following the identical treatment of chemoresistant cells, where TG2-S remained predominantly membrane-associated. This suggests that the deficient subcellular relocalisation of TG2-S from membranous structures into the cytoplasm may limit the apoptic response to cisplatin toxicity in chemoresistant cells. Structural analysis of TG2 revealed the presence of binding motifs for interaction of TG2-S with the membrane scaffold protein LC3/LC3 homologue that could contribute to a novel mechanism of chemotherapeutic resistance in HepG2 cells Full article

Neutrophils present the host’s first line of defense against bacterial infections. These immune effector cells are mobilized rapidly to destroy invading pathogens by (a) reactive oxygen species (ROS)-mediated oxidative bursts and (b) via phagocytosis. In addition, their antimicrobial service is capped via a distinct cell death mechanism, by the release of their own decondensed nuclear DNA, supplemented with a variety of embedded proteins and enzymes. The extracellular DNA meshwork ensnares the pathogenic bacteria and neutralizes them. Such neutrophil extracellular DNA traps (NETs) have the potential to trigger a hemostatic response to pathogenic infections. The web-like chromatin serves as a prothrombotic scaffold for platelet adhesion and activation. What is less obvious is that platelets can also be involved during the initial release of NETs, forming heterotypic interactions with neutrophils and facilitating their responses to pathogens. Together, the platelet and neutrophil responses can effectively localize an infection until it is cleared. However, not all microbial infections are easily cleared. Certain pathogenic organisms may trigger dysregulated platelet–neutrophil interactions, with a potential to subsequently propagate thromboinflammatory processes. These may also include the release of some NETs. Therefore, in order to make rational intervention easier, further elucidation of platelet, neutrophil, and pathogen interactions is still needed. Full article