Biochemical and biophysical research communications最新文献

Cells maintain proteostasis by sequestering misfolded proteins into deposition sites. Aggregation-prone endoplasmic reticulum (ER) proteins form membrane-bound nuclear compartments that are cleared during cell division, yet the mechanisms underlying their spatial organization remain unclear. Here, using transcriptomic and proteomic analyses, we identified the ER-localized Hsp70 chaperone BiP as a key player. Genetic depletion or chemical inhibition of BiP prevented nuclear aggregate formation, while manipulating BiP regulators perturbed the aggregate formation. BiP-driven aggregation precedes the inner nuclear membrane synthesis that encapsulated the aggregates. Under proteostatic stress, nuclear aggregates localized adjacent to ER-derived aggregates. Our findings demonstrate that BiP is essential for organizing ER-derived aggregates in the nucleus, which further regulate nuclear proteostasis through spatial interactions with nuclear aggregates.

Radioresistance is a key challenge in colorectal cancer (CRC) therapy. Through integrative analysis of TCGA datasets and RNA-seq of irradiated CRC cells, we identified PLS3-AS1 as a radiation-inducible lncRNA upregulated in recurrent tumors and post-irradiation. Functional assays revealed that PLS3-AS1 promotes CRC cell proliferation, survival, and radioresistance in vitro and in vivo. Mechanistically, PLS3-AS1 enhances NF-κB signaling by directly binding to p65 and IκBα, disrupting their interaction and facilitating p65 nuclear translocation. Moreover, PLS3-AS1 expression is itself induced by NF-κB activation, forming a positive feedback loop. Inhibition of NF-κB with BAY 11-7082 suppressed PLS3-AS1 expression and reversed its pro-tumorigenic effects. These findings identify PLS3-AS1 as a critical mediator of NF-κB-driven radioresistance in CRC and a potential therapeutic target to improve radiotherapy efficacy.

RALY, a heterogeneous nuclear ribonucleoprotein, binds to nascent RNA and participates in multiple aspects of RNA metabolism, including transport, splicing, transcription, and translation. Recent studies have revealed that RALY is overexpressed in various cancers, such as breast, uterine, and liver cancers. This overexpression has been associated with poor patient survival and uncontrolled carcinoma cell proliferation. In this study, we demonstrate that RALY functions as a key regulator of cell proliferation, migration, and invasion in the hepatocellular carcinoma (HCC) cell lines Hep3B and HepG2. Mechanistically, RALY promotes epithelial-mesenchymal transition (EMT) through regulation of the transcription factor Snail. RALY directly binds to Snail mRNA, thereby enhancing its stability. In addition, RALY modulates the TGF-β signaling pathway to promote Snail transcription. Together, our findings establish a functional link between RALY and EMT and reveal a previously unrecognized role of RALY in cancer cell metastasis. Accumulating evidence, including the results presented here, suggests that RALY represents a potential therapeutic target for cancer treatment.

Additives like arginine, while known to enhance protein stability, were observed to improve the solubility of proteins in an equimolar mixture of arginine and glutamic acid. This study examines the conformational stability of a model protein, chymotrypsin inhibitor 2, in arginine, glutamic acid, and their mixture over a temperature range of 300-440 K using Replica Exchange Molecular Dynamics. The study demonstrates that arginine stabilized CI2 through hydrogen bonds and electrostatic interactions with polar and charged residues, thereby restricting conformational fluctuations and reducing solvent penetration. Glutamic acid enhanced its stability by promoting preferential hydration and maintaining a hydration shell that minimized hydrophobic exposure during thermal fluctuations. The arginine-glutamic acid mixture exhibited intermediate stabilizing behavior, where hydrogen bonding interactions between the oppositely charged amino acids reduced arginine self-association, resulting in a balanced hydrophilic-hydrophobic environment around the protein. Free energy landscape analysis showed no significant unfolding transitions across temperatures, supporting a kinetically stable ensemble. Radial distribution and cluster analyses indicated that arginine formed compact clusters near the protein surface, whereas glutamic acid remained dispersed, allowing higher water mobility. Water diffusion and dipole-dipole correlation studies revealed faster hydration dynamics in glutamic acid and slower, more confined motion in arginine, with intermediate behavior in the mixed solution.

Objectives: Triple-negative breast cancer (TNBC) is the most aggressive subtype of breast cancer. Due to its radiotherapy resistance, high metastasis rate and recurrence rate, the prognosis is poor. So far, effective therapeutic drugs for it remain elusive. Cetylpyridinium chloride (CPC) has initially demonstrated anti-tumor properties in various tumors. Therefore, the aim of this study is to explore the intervention effect of CPC on macrophages in the tumor microenvironment (TME) and its therapeutic effect on TNBC.

Methods: The effects of CPC on macrophage polarization and expression changes were evaluated by WB and RT-qPCR. Multiple TNBC cell lines and non-cancer cells were exposed to CPC under different conditions (concentration and duration), and cell survival and proliferation were assessed by CCK8 and colony formation. The effects of CPC on apoptosis, migration and invasion of TNBC cells were further evaluated. The mitochondrial status was assessed by Mito-Tracker Red CMXRos staining, mitochondrial protein expression detection and ATP content detection to explain the cause of apoptosis. The effect of CPC on TNBC growth was further confirmed in animal models.

Results: Our results demonstrated that CPC (2 μM) inhibited the M2 polarization of macrophages induced by interleukin-4 (IL-4) and tumor secretions, and reduced their characteristic secretions that promote tumor growth. Meanwhile, CPC significantly suppressed the proliferation of multiple TNBC cell lines, induced mitochondrial damage, significantly decreased cellular ATP levels, and ultimately led to tumor cell apoptosis. Moreover, TNBC cells are more sensitive to CPC than non-tumor cells. Additionally, CPC significantly inhibited the migration and invasion of TNBC cells. Finally, we confirmed the growth inhibitory effect of CPC on TNBC in vivo.

Conclusion: CPC has dual effects: it inhibits the M2 polarization of tumor-associated macrophages (TAMs) and directly suppresses the proliferation and invasion of TNBC cells, ultimately effectively inhibiting the growth of TNBC in vivo. Considering that CPC has been administered in humans as a safe drug-disinfectant for decades, our study here provides a molecular basis for the application of CPC as a potential option to clinical treatments of TNBC.

Background: Proline supplementation promotes proliferation and invasion in cancers under acute nutrient deprivation. However, its molecular basis in hepatocellular carcinoma is not fully understood yet. This study investigates how proline metabolism promotes hepatocellular carcinoma (HCC) cell proliferation and survival during nutrient starvation.

Methods: Proline metabolism-related protein expression in HCC versus paracarcinoma tissues was analyzed using The Cancer Genome Atlas database. HCC cell viability was quantified via Cell Counting Kit 8 assays, and colony formation capacity was evaluated. Intracellular ROS levels were measured by flow cytometry. Autophagic flux was assessed by GFP/mCherry fluorescence ratio and autophagy-related proteins by immunoblotting. Lipid droplet deposition was visualized using Hoechst 33342 and BODIPY 493/503 staining.

Results: Proline metabolism-related proteins were significantly upregulated in HCC tissues compared to paracarcinoma controls. Under acute nutrient stress, exogenous proline reduced cellular ROS levels, attenuated lipid droplet accumulation, and suppressed excessive autophagy in HCC cells. Proline rescued nutrient deprivation-induced tumor growth inhibition, which could be reversed by H₂O₂ and rapamycin.

Conclusion: Proline metabolism sustains HCC cell survival during nutrient restriction by reducing ROS accumulation, thereby inhibiting lipid droplet formation and autophagy.

Acute kidney injury (AKI) survivors are at risk of progression to chronic kidney disease (CKD), but stage-specific roles of RIPK3 and optimal intervention timing remain unclear. Using a bilateral renal ischemia-reperfusion injury model in Ripk3+/+, Ripk3+/-, and Ripk3-/- mice together with time-window pharmacological inhibition, we profiled RIPK3 activity across 28 days and examined upstream regulation. RIPK3 and phosphorylated RIPK3 increased from 6 h after injury and remained elevated through day 28. Unexpectedly, Ripk3 ± mice showed the mildest injury and fibrosis, whereas Ripk3-/- mice were not superior to wild-type controls, indicating a U-shaped dose-response. Intervention timing was critical: the RIPK3 inhibitor GSK872 initiated on day 7 improved renal function, histology, and 90-day survival more than treatment starting on day 0 or day 14, coinciding with a shift from peak repair-factor expression to sustained inflammatory cytokine elevation. Mechanistically, TGF-β1 induced RIPK3 transcription via Smad3, while RIPK3-associated necroptotic injury promoted HMGB1 release that enhanced TGF-β1/Smad3 signaling, forming a positive feedback loop; subacute SB431542 treatment reduced fibrosis. These findings support stage- and dose-aware RIPK3 modulation as a strategy to limit AKI-to-CKD progression.

T-cell receptors (TCRs) that target tumor antigens are crucial for antitumor immunity; however, tumor-specific TCRs often exhibit low affinity for their cognate antigens, making the identification of functional TCRs challenging due to the limited sensitivity of current detection methods. In this study, we established a high-sensitivity TCR screening platform by generating Jurkat cell reporter clones with dual knockout (DKO) of endogenous Fas and TCRα via CRISPR-Cas9 system. In a viral antigen model system, these DKO Jurkat cells exhibited approximately 100-fold greater sensitivity to antigen stimulation compared with parental Jurkat cells. Notably, our DKO Jurkat-based platform enabled the identification of tumor-specific CD8⁺ T cells from a lung cancer patient that could not be detected using parental Jurkat cells. Moreover, the identified tumor-specific T-cell clone exhibited a unique phenotype characterized by robust cytotoxic T lymphocyte (CTL) activity and natural killer-like properties. Together, these findings demonstrate that dual deletion of Fas and TCRα in Jurkat cells enables highly sensitive functional TCR screening. Integration of this platform with single-cell analysis facilitates the discovery of previously uncharacterized tumor-reactive TCRs and provides a powerful tool for advancing TCR-based cancer immunotherapy.