Lactiplantibacillus plantarum K8 lysates regulate hypoxia-induced gene expression.

Hypoxic responses have been implicated in critical pathologies, including inflammation, immunity, and tumorigenesis. Recently, efforts to identify effective natural remedies and health supplements are increasing. Previous studies have reported that the cell lysates and the cell wall-bound lipoteichoic acids of Lactiplantibacillus plantarum K8 (K8) exert anti-inflammatory and immunomodulative effects. However, the effect of K8 on cellular hypoxic responses remains unknown. In this study, we found that K8 lysates had a potent suppressive effect on gene expression under hypoxia. K8 lysates markedly downregulated hypoxia-induced HIF1α accumulation in the human bone marrow and lung cancer cell lines, SH-SY5Y and H460. Consequently, the transcription of known HIF1α target genes, such as p21, GLUT1, and ALDOC, was notably suppressed in the K8 lysate supplement and purified lipoteichoic acids of K8, upon hypoxic induction. Intriguingly, K8 lysates decreased the expression of PHD2 and VHL proteins, which are responsible for HIF1α destabilization under normoxic conditions, suggesting that K8 may regulate HIF1α stability in a non-canonical pathway. Overall, our results suggest that K8 lysates desensitize the cells to hypoxic stresses and suppress HIF1α-mediated hypoxic gene activation.

HSPA9 reduction exacerbates symptoms and cell death in DSS-Induced inflammatory colitis.

Inflammatory bowel disease (IBD) is a chronic inflammatory condition that is influenced by various factors, including environmental factors, immune responses, and genetic elements. Among the factors that influence IBD progression, macrophages play a significant role in generating inflammatory mediators, and an increase in the number of activated macrophages contributes to cellular damage, thereby exacerbating the overall inflammatory conditions. HSPA9, a member of the heat shock protein 70 family, plays a crucial role in regulating mitochondrial processes and responding to oxidative stress. HSPA9 deficiency disrupts mitochondrial dynamics, increasing mitochondrial fission and the production of reactive oxygen species. Based on the known functions of HSPA9, we considered the possibility that HSPA9 reduction may contribute to the exacerbation of colitis and investigated its relevance. In a dextran sodium sulfate-induced colitis mouse model, the downregulated HSPA9 exacerbates colitis symptoms, including increased immune cell infiltration, elevated proinflammatory cytokines, decreased tight junctions, and altered macrophage polarization. Moreover, along with the increased mitochondrial fission, we found that the reduction in HSPA9 significantly affected the superoxide dismutase 1 levels and contributed to cellular death. These findings enhance our understanding of the intricate mechanisms underlying colitis and contribute to the development of novel therapeutic approaches for this challenging condition.

Regulation of inflammatory response by LINC00346 via miR-25-3p-mediated modulation of the PTEN/PI3K/AKT/NF-κB pathway.

Long intergenic non-coding RNA 346 (LINC00346) has been reported to be involved in the development of atherosclerosis and specific cancers by affecting signaling pathways. However, its function in inflammation has not been thoroughly studied. Therefore, its expression pattern and function were determined in the human macrophage-like cell line THP-1. Lipopolysaccharide (LPS) treatment induced the expression of LINC00346. LPS-induced NF-κB activation and proinflammatory cytokine expression were suppressed or enhanced by the overexpression or knockdown of LINC00346, respectively. Analyses using dual luciferase assay and decoy RNAs that could block RNA-RNA interactions indicated that LINC00346 improves phosphatase and tensin homolog (PTEN) expression by sponging miR-25-3p. Subsequently, PTEN suppresses phosphoinositide-3 kinase (PI3K)-mediated conversion of phosphatidylinositol-4,5-bisphosphate (PIP2) into phosphatidylinositol-3,4,5-trisphosphate (PIP3) as well as consequent activation of protein kinase B (AKT) and NF-κB. Interestingly, database analysis revealed that the expression levels of LINC00346 and PTEN were simultaneously decreased in breast cancer tissues. Further analyses conducted using a breast cancer cell line, MDA-MB-231, confirmed the functional relationship among LINC00346, miR-25-3p, and PTEN in LPS-induced activation of NF-κB. These results indicate that miR-25-3p-sponging activity of LINC00346 affects the balance between PTEN and PI3K as well as the downstream activation of AKT/NF-κB pathway in inflammatory conditions.

Inhibition of VHL by VH298 Accelerates Pexophagy by Activation of HIF-1α in HeLa Cells.

Autophagy is a pivotal biological process responsible for maintaining the homeostasis of intracellular organelles. Yet the molecular intricacies of peroxisomal autophagy (pexophagy) remain largely elusive. From a ubiquitin-related chemical library for screening, we identified several inhibitors of the Von Hippel-Lindau (VHL) E3 ligase, including VH298, thereby serving as potent inducers of pexophagy. In this study, we observed that VH298 stimulates peroxisomal degradation by ATG5 dependently and escalates the ubiquitination of the peroxisomal membrane protein ABCD3. Interestingly, the ablation of NBR1 is similar to the curtailed peroxisomal degradation in VH298-treated cells. We also found that the pexophagy induced by VH298 is impeded upon the suppression of gene expression by the translation inhibitor cycloheximide. Beyond VHL inhibition, we discovered that roxadustat, a direct inhibitor of HIF-α prolyl hydroxylase, is also a potent inducer of pexophagy. Furthermore, we found that VH298-mediated pexophagy is blocked by silencing HIF-1α. In conclusion, our findings suggest that VH298 promotes pexophagy by modulating VHL-mediated HIF-α transcriptional activity.

Early life exposure to perfluorooctanesulfonate (PFOS) impacts vital biological processes in Xenopus laevis: Integrated morphometric and transcriptomic analyses.

Perfluorooctanesulfonate (PFOS) is a ubiquitous environmental pollutant associated with increasing health concerns and environmental hazards. Toxicological analyses of PFOS exposure are hampered by large interspecies variations and limited studies on the mechanistic details of PFOS-induced toxicity. We investigated the effects of PFOS exposure on Xenopus laevis embryos based on the reported developmental effects in zebrafish. X. laevis was selected to further our understanding of interspecies variation in response to PFOS, and we built upon previous studies by including transcriptomics and an assessment of ciliogenic effects. Midblastula-stage X. laevis embryos were exposed to PFOS using the frog embryo teratogenesis assay Xenopus (FETAX). Results showed teratogenic effects of PFOS in a time- and dose-dependent manner. The morphological abnormalities of skeleton deformities, a small head, and a miscoiled gut were associated with changes in gene expression evidenced by whole-mount in situ hybridization and transcriptomics. The transcriptomic profile of PFOS-exposed embryos indicated the perturbation in the expression of genes associated with cell death, and downregulation in adenosine triphosphate (ATP) biosynthesis. Moreover, we observed the effects of PFOS exposure on cilia development as a reduction in the number of multiciliated cells and changes in the directionality and velocity of the cilia-driven flow. Collectively, these data broaden the molecular understanding of PFOS-induced developmental effects, whereby ciliary dysfunction and disrupted ATP synthesis are implicated as the probable modes of action of embryotoxicity. Furthermore, our findings present a new challenge to understand the links between PFOS-induced developmental toxicity and vital biological processes.

Enhanced primary ciliogenesis via mitochondrial oxidative stress activates AKT to prevent neurotoxicity in HSPA9/mortalin-depleted SH-SY5Y cells.

The primary cilium, an antenna-like structure on the cell surface, acts as a mechanical and chemical sensory organelle. Primary cilia play critical roles in sensing the extracellular environment to coordinate various developmental and homeostatic signaling pathways. Here, we showed that the depletion of heat shock protein family A member 9 (HSPA9)/mortalin stimulates primary ciliogenesis in SH-SY5Y cells. The downregulation of HSPA9 enhances mitochondrial stress by increasing mitochondrial fragmentation and mitochondrial reactive oxygen species (mtROS) generation. Notably, the inhibition of either mtROS production or mitochondrial fission significantly suppressed the increase in primary ciliogenesis in HSPA9-depleted cells. In addition, enhanced primary ciliogenesis contributed to cell survival by activating AKT in SH-SY5Y cells. The abrogation of ciliogenesis through the depletion of IFT88 potentiated neurotoxicity in HSPA9-knockdown cells. Furthermore, both caspase-3 activation and cell death were increased by MK-2206, an AKT inhibitor, in HSPA9-depleted cells. Taken together, our results suggest that enhanced primary ciliogenesis plays an important role in preventing neurotoxicity caused by the loss of HSPA9 in SH-SY5Y cells.

PEX13 prevents pexophagy by regulating ubiquitinated PEX5 and peroxisomal ROS.

Peroxisomes are rapidly degraded during amino acid and oxygen deprivation by a type of selective autophagy called pexophagy. However, how damaged peroxisomes are detected and removed from the cell is poorly understood. Recent studies suggest that the peroxisomal matrix protein import machinery may serve double duty as a quality control machinery, where they are directly involved in activating pexophagy. Here, we explored whether any matrix import factors are required to prevent pexophagy, such that their loss designates peroxisomes for degradation. Using gene editing and quantitative fluorescence microscopy on culture cells and a zebrafish model system, we found that PEX13, a component of the peroxisomal matrix import system, is required to prevent the degradation of otherwise healthy peroxisomes. The loss of PEX13 caused an accumulation of ubiquitinated PEX5 on peroxisomes and an increase in peroxisome-dependent reactive oxygen species that coalesce to induce pexophagy. We also found that PEX13 protein level is downregulated to aid in the induction of pexophagy during amino acid starvation. Together, our study points to PEX13 as a novel pexophagy regulator that is modulated to maintain peroxisome homeostasis.Abbreviations: AAA ATPases: ATPases associated with diverse cellular activities; ABCD3: ATP binding cassette subfamily D member; 3ACOX1: acyl-CoA oxidase; 1ACTA1: actin alpha 1, skeletal muscle; ACTB: actin beta; ATG5: autophagy related 5; ATG7: autophagy related 7; ATG12: autophagy related 12; ATG16L1: autophagy related 16 like 1; CAT: catalase; CQ: chloroquine; Dpf: days post fertilization: FBS: fetal bovine serum; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; GFP: green fluorescent protein; H2O2: hydrogen peroxide; HA - human influenza hemagglutinin; HBSS: Hanks' Balanced Salt Solution; HCQ; hydroxychloroquine; KANL: lysine alanine asparagine leucine; KO: knockout; MAP1LC3B: microtubule associated protein 1 light chain 3 beta; MEF: mouse embryonic fibroblast; MTOR: mechanistic target of rapamycin kinase; MTORC1: mechanistic target of rapamycin kinase complex 1; MTORC2: mechanistic target of rapamycin kinase complex 2; MYC: MYC proto-oncogene, bHLH transcription factor; MZ: maternal and zygotic; NAC: N-acetyl cysteine; NBR1 - NBR1 autophagy cargo receptor; PBD: peroxisome biogenesis disorder; PBS: phosphate-buffered saline; PEX: peroxisomal biogenesis factor; PTS1: peroxisome targeting sequence 1; RFP: red fluorescent protein; ROS: reactive oxygen speciess; iRNA: short interfering RNA; SKL: serine lysine leucine; SLC25A17/PMP34: solute carrier family 25 member 17; Ub: ubiquitin; USP30: ubiquitin specific peptidase 30.

Zinc enhances autophagic flux and lysosomal function through transcription factor EB activation and V-ATPase assembly.

The stimulation of autophagy or lysosomes has been considered therapeutic for neurodegenerative disorders because the accumulation of misfolded proteins is commonly observed in the brains of individuals with these diseases. Although zinc is known to play critical roles in the functions of lysosomes and autophagy, the mechanism behind this regulatory relationship remains unclear. Therefore, in this study, we examined which mechanism is involved in zinc-mediated activation of autophagy and lysosome. Exposure to zinc at a sub-lethal concentration activated autophagy in a concentration-dependent manner in mRFP-GFP-LC3-expressing H4 glioma cells. Zinc also rescued the blocking of autophagic flux arrested by pharmaceutical de-acidification. Co-treatment with zinc attenuated the chloroquine (CQ)-induced increase in the number and size of mRFP-GFP-LC3 puncta in H4 cells and accumulation of p62 by CQ or ammonium chloride in both H4 and mouse cerebrocortical cultures. Zinc rapidly induced the expression of cathepsin B (CTSB) and cathepsin D (CTSD), representative lysosomal proteases in neurons, which appeared likely to be mediated by transcription factor EB (TFEB). We observed the translocation of TFEB from neurite to nucleus and the dephosphorylation of TFEB by zinc. The addition of cycloheximide, a chemical inhibitor of protein synthesis, inhibited the activity of CTSB and CTSD at 8 h after zinc exposure but not at 1 h, indicating that only late lysosomal activation was dependent on the synthesis of CTSB and CTSD proteins. At the very early time point, the activation of cathepsins was mediated by an increased assembly of V-ATPase on lysosomes and resultant lysosomal acidification. Finally, considering that P301L mutation in tau protein causes frontotemporal dementia through aggressive tau accumulation, we investigated whether zinc reduces the accumulation of protein aggregates in SK-N-BE(2)-C neuroblastoma cells expressing wild-type tau or mutant P301L-tau. Zinc markedly attenuated the levels of phosphorylated tau and total tau as well as p62 in both wild-type and mutant tau-overexpressing cells. We also observed that zinc was more effective than rapamycin at inducing TFEB-dependent CTSB and CTSD expression and V-ATPase-dependent lysosomal acidification and CTSB/CTSD activation. These results suggest that the regulation of zinc homeostasis could be a new approach for developing treatments for neurodegenerative diseases, including Alzheimer's and Parkinson's.

Carnitine Protects against MPP+-Induced Neurotoxicity and Inflammation by Promoting Primary Ciliogenesis in SH-SY5Y Cells.

Primary cilia help to maintain cellular homeostasis by sensing conditions in the extracellular environment, including growth factors, nutrients, and hormones that are involved in various signaling pathways. Recently, we have shown that enhanced primary ciliogenesis in dopamine neurons promotes neuronal survival in a Parkinson's disease model. Moreover, we performed fecal metabolite screening in order to identify several candidates for improving primary ciliogenesis, including L-carnitine and acetyl-L-carnitine. However, the role of carnitine in primary ciliogenesis has remained unclear. In addition, the relationship between primary cilia and neurodegenerative diseases has remained unclear. In this study, we have evaluated the effects of carnitine on primary ciliogenesis in 1-methyl-4-phenylpyridinium ion (MPP+)-treated cells. We found that both L-carnitine and acetyl-L-carnitine promoted primary ciliogenesis in SH-SY5Y cells. In addition, the enhancement of ciliogenesis by carnitine suppressed MPP+-induced mitochondrial reactive oxygen species overproduction and mitochondrial fragmentation in SH-SY5Y cells. Moreover, carnitine inhibited the production of pro-inflammatory cytokines in MPP+-treated SH-SY5Y cells. Taken together, our findings suggest that enhanced ciliogenesis regulates MPP+-induced neurotoxicity and inflammation.

Post-Translational Modifications of ATG4B in the Regulation of Autophagy.

Autophagy plays a key role in eliminating and recycling cellular components in response to stress, including starvation. Dysregulation of autophagy is observed in various diseases, including neurodegenerative diseases, cancer, and diabetes. Autophagy is tightly regulated by autophagy-related (ATG) proteins. Autophagy-related 4 (ATG4) is the sole cysteine protease, and four homologs (ATG4A-D) have been identified in mammals. These proteins have two domains: catalytic and short fingers. ATG4 facilitates autophagy by promoting autophagosome maturation through reversible lipidation and delipidation of seven autophagy-related 8 (ATG8) homologs, including microtubule-associated protein 1-light chain 3 (LC3) and GABA type A receptor-associated protein (GABARAP). Each ATG4 homolog shows a preference for a specific ATG8 homolog. Post-translational modifications of ATG4, including phosphorylation/dephosphorylation, O-GlcNAcylation, oxidation, S-nitrosylation, ubiquitination, and proteolytic cleavage, regulate its activity and ATG8 processing, thus modulating its autophagic activity. We reviewed recent advances in our understanding of the effect of post-translational modification on the regulation, activity, and function of ATG4, the main protease that controls autophagy.

Protective Effect of GIP against Monosodium Glutamate-Induced Ferroptosis in Mouse Hippocampal HT-22 Cells through the MAPK Signaling Pathway.

The effect of glucose-dependent insulinotropic polypeptide (GIP) on cells under oxidative stress induced by glutamate, a neurotransmitter, and the underlying molecular mechanisms were assessed in the present study. We found that in the pre-treatment of HT-22 cells with glutamate in a dose-dependent manner, intracellular ROS were excessively generated, and additional cell damage occurred in the form of lipid peroxidation. The neurotoxicity caused by excessive glutamate was found to be ferroptosis and not apoptosis. Other factors (GPx-4, Nrf2, Nox1 and Hspb1) involved in ferroptosis were also identified. In other words, it was confirmed that GIP increased the activity of sub-signalling molecules in the process of suppressing ferroptosis as an antioxidant and maintained a stable cell cycle even under glutamate-induced neurotoxicity. At the same time, in HT-22 cells exposed to ferroptosis as a result of excessive glutamate accumulation, GIP sustained cell viability by activating the mitogen-activated protein kinase (MAPK) signalling pathway. These results suggest that the overexpression of the GIP gene increases cell viability by regulating mechanisms related to cytotoxicity and reactive oxygen species production in hippocampal neuronal cell lines.

Nalfurafine Hydrochloride, a κ-Opioid Receptor Agonist, Induces Melanophagy via PKA Inhibition in B16F1 Cells.

Selective autophagy controls cellular homeostasis by degrading unnecessary or damaged cellular components. Melanosomes are specialized organelles that regulate the biogenesis, storage, and transport of melanin in melanocytes. However, the mechanisms underlying melanosomal autophagy, known as the melanophagy pathway, are poorly understood. To better understand the mechanism of melanophagy, we screened an endocrine-hormone chemical library and identified nalfurafine hydrochlorides, a κ-opioid receptor agonist, as a potent inducer of melanophagy. Treatment with nalfurafine hydrochloride increased autophagy and reduced melanin content in alpha-melanocyte-stimulating hormone (α-MSH)-treated cells. Furthermore, inhibition of autophagy blocked melanosomal degradation and reversed the nalfurafine hydrochloride-induced decrease in melanin content in α-MSH-treated cells. Consistently, treatment with other κ-opioid receptor agonists, such as MCOPPB or mianserin, inhibited excessive melanin production but induced autophagy in B16F1 cells. Furthermore, nalfurafine hydrochloride inhibited protein kinase A (PKA) activation, which was notably restored by forskolin, a PKA activator. Additionally, forskolin treatment further suppressed melanosomal degradation as well as the anti-pigmentation activity of nalfurafine hydrochloride in α-MSH-treated cells. Collectively, our data suggest that stimulation of κ-opioid receptors induces melanophagy by inhibiting PKA activation in α-MSH-treated B16F1 cells.

BRCA1-BARD1 regulates transcription through modulating topoisomerase IIß.

RNA polymerase II (Pol II)-dependent transcription in stimulus-inducible genes requires topoisomerase IIß (TOP2B)-mediated DNA strand break and the activation of DNA damage response signalling in humans. Here, we report a novel function of the breast cancer 1 (BRCA1)-BRCA1-associated ring domain 1 (BARD1) complex in this process. We found that BRCA1 is phosphorylated at S1524 by the kinases ataxia-telangiectasia mutated and ATR during gene activation, and that this event is important for productive transcription. Our biochemical and genomic analyses showed that the BRCA1-BARD1 complex interacts with TOP2B in the EGR1 transcription start site and in a large number of protein-coding genes. Intriguingly, the BRCA1-BARD1 complex ubiquitinates TOP2B, which stabilizes TOP2B binding to DNA while BRCA1 phosphorylation at S1524 controls the TOP2B ubiquitination by the complex. Together, these findings suggest the novel function of the BRCA1-BARD1 complex in the regulation of TOP2B and Pol II-mediated gene expression.

Primary Ciliogenesis by 2-Isopropylmalic Acid Prevents PM2.5-Induced Inflammatory Response and MMP-1 Activation in Human Dermal Fibroblasts and a 3-D-Skin Model.

Particulate matters (PMs) increase oxidative stress and inflammatory response in different tissues. PMs disrupt the formation of primary cilia in various skin cells, including keratinocytes and melanocytes. In this study, we found that 2-isopropylmalic acid (2-IPMA) promoted primary ciliogenesis and restored the PM2.5-induced dysgenesis of primary cilia in dermal fibroblasts. Moreover, 2-IPMA inhibited the generation of excessive reactive oxygen species and the activation of stress kinase in PM2.5-treated dermal fibroblasts. Further, 2-IPMA inhibited the production of pro-inflammatory cytokines, including IL-6 and TNF-α, which were upregulated by PM2.5. However, the inhibition of primary ciliogenesis by IFT88 depletion reversed the downregulated cytokines by 2-IPMA. Moreover, we found that PM2.5 treatment increased the MMP-1 expression in dermal fibroblasts and a human 3-D-skin model. The reduced MMP-1 expression by 2-IPMA was further reversed by IFT88 depletion in PM2.5-treated dermal fibroblasts. These findings suggest that 2-IPMA ameliorates PM2.5-induced inflammation by promoting primary ciliogenesis in dermal fibroblasts.

Physiological Functions of Thiol Peroxidases (Gpx1 and Prdx2) during Xenopus laevis Embryonic Development.

Glutathione peroxidase 1 (Gpx1) and peroxiredoxin 2 (Prdx2) belong to the thiol peroxidase family of antioxidants, and have been studied for their antioxidant functions and roles in cancers. However, the physiological significance of Gpx1 and Prdx2 during vertebrate embryogenesis are lacking. Currently, we investigated the functional roles of Gpx1 and Prdx2 during vertebrate embryogenesis using Xenopus laevis as a vertebrate model. Our investigations revealed the zygotic nature of gpx1 having its localization in the eye region of developing embryos, whereas prdx2 exhibited a maternal nature and were localized in embryonic ventral blood islands. Furthermore, the gpx1-morphants exhibited malformed eyes with incompletely detached lenses. However, the depletion of prdx2 has not established its involvement with embryogenesis. A molecular analysis of gpx1-depleted embryos revealed the perturbed expression of a cryba1-lens-specific marker and also exhibited reactive oxygen species (ROS) accumulation in the eye regions of gpx1-morphants. Additionally, transcriptomics analysis of gpx1-knockout embryos demonstrated the involvement of Wnt, cadherin, and integrin signaling pathways in the development of malformed eyes. Conclusively, our findings indicate the association of gpx1 with a complex network of embryonic developmental pathways and ROS responses, but detailed investigation is a prerequisite in order to pinpoint the mechanistic details of these interactions.

Clinically Applicable Serum Biomarkers Among 14 Candidates Associated With Recurrence of Stage II and III Colorectal Cancer.

BACKGROUND/AIM: We evaluated the predictive value of candidate serum biomarkers for recurrence in stage II and III colorectal cancer (CRC) after curative surgery. PATIENTS AND METHODS: A total of 33 and 120 patients with CRC with or without recurrence at 5 years after curative surgery were included in the training set and the validation set, respectively. Possible serum biomarkers were examined for associations with CRC recurrence using receiver operating characteristics (ROC) curve analysis. RESULTS: In the training set, the expression levels of the 14 biomarkers were compared according to recurrence. Among them, five biomarkers that had significantly different expression levels were validated in 60 patients with recurrence at 5 years after curative surgery and 60 patients without. Multivariate analysis showed that natural log-transformed values of carcinoembryonic antigen (CEA), cyclin-dependent kinase regulatory subunit 2 (CKS2), 2'-5'-oligoadenylate synthetase 2 (OAS2), and autophagy-related gene 5 (ATG5) in preoperative serum were significantly related to recurrence. ROC analysis showed that these biomarkers were able to discriminate patients with recurrence from those without (area under the curve=0.828, 95% confidence interval=0.755-0.990). CONCLUSION: Preoperative serum levels of CEA, CKS2, OAS2 and ATG5 were independent risk factors for recurrence. A combination of serum CEA, CKS2, OAS2 and ATG5 predicted tumor recurrence well in patients with stage II and III CRC.

ZNF507 affects TGF-ß signaling via TGFBR1 and MAP3K8 activation in the progression of prostate cancer to an aggressive state.

BACKGROUND: The progression of prostate cancer (PC) to the highly aggressive metastatic castration-resistant prostate cancer (mCRPC) or neuroendocrine prostate cancer (NEPC) is a fatal condition and the underlying molecular mechanisms are poorly understood. Here, we identified the novel transcriptional factor ZNF507 as a key mediator in the progression of PC to an aggressive state. METHODS: We analyzed ZNF507 expression in the data from various human PC database and high-grade PC patient samples. By establishment of ZNF507 knockdown and overexpression human PC cell lines, we assessed in vitro PC phenotype changes including cell proliferation, survival, migration and invasion. By performing microarray with ZNF507 knockdown PC cells, we profiled the gene clusters affected by ZNF507 knockdown. Moreover, ZNF507 regulated key signal was evaluated by dual-luciferase reporter and chromatin immunoprecipitation (ChIP) assays. Finally, we performed xenograft and in vivo metastasis assay to confirm the effect of ZNF507 knockdown in PC cells. RESULTS: We found that ZNF507 expression was increased, particularly in the highly graded PC. ZNF507 was also found to be associated with metastatic PC of a high grade. Loss- or gain-of-function-based analysis revealed that ZNF507 promotes the growth, survival, proliferation, and metastatic properties of PC (e.g., epithelial-mesenchymal transition) by upregulating TGF-ß signaling. Profiling of gene clusters affected by ZNF507 knockdown revealed that ZNF507 positively regulated the transcription of TGFBR1, MAP3K8, and FURIN, which in turn promoted the progression of PC to highly metastatic and aggressive state. CONCLUSIONS: Our findings suggest that ZNF507 is a novel key regulator of TGF-ß signaling in the progression of malignant PC and could be a promising target for studying the development of advanced metastatic PCs.

Tumor immune microenvironment of primary colorectal adenocarcinomas metastasizing to the liver or lungs.

BACKGROUND: Because of the heterogeneity of metastatic colorectal cancer (mCRC), a genome-wide analysis was performed to characterize the tumor immune microenvironment (TIME). METHODS: RNA-seq analysis of 62 primary CRCs without and 63 with systemic metastasis (SM- and SM+ groups) was conducted, and the data were used in a training set after adjustment by propensity score matching. Samples were further subdivided into those with hepatic metastasis (CHM subgroup), pulmonary metastasis (CPM subgroup), or concurrent CHM and CPM (concurrent group). Validation was done by quantitative reverse-transcription polymerase chain reaction using another 40 primary CRC samples. RESULTS: Compared with the CHM or CPM subgroups, the concurrent group showed upregulated in inflammatory or immune processes, cytokine secretion, and myeloid leukocyte migration. Nine candidate genes were selected: SM-specific IDO1, JAM3, and PDE2A; CHM- or CPM-specific BIRC7; CPM-specific HISI1H2BK, and both SM-specific and CHM- or CPM-specific EPHB6, LPL, THBD, and PPBP. In a validation set of primary CRCs, JAM3 and IDO1 (p = 0.044 and p = 0.036, respectively) were confirmed to show significant upregulation and downregulation, respectively, in the SM+ group, whereas HIST1H2BK (p = 0.017) was significantly upregulated in the CPM subgroup. CONCLUSIONS: Our findings indicate that a host-suppressive TIME is established in the primary tumor of mCRC and identify immune-related site-specific markers of mCRC.

Triamterene induces autophagic degradation of lysosome by exacerbating lysosomal integrity.

The maintenance of lysosomal integrity is essential for lysosome function and cell fate. Damaged lysosomes are degraded by lysosomal autophagy, lysophagy. The mechanism underlying lysophagy remains largely unknown; this study aimed to contribute to the understanding of this topic. A cell-based screening system was used to identify novel lysophagy modulators. Triamterene (6-phenylpteridine-2,4,7-triamine) was identified as one of the most potent lysophagy inducers from the screening process. We found that triamterene causes lysosomal rupture without affecting other cellular organelles and increases autophagy flux in HepG2 cells. Damaged lysosomes in triamterene-treated cells were removed by autophagy-mediated pathway, which was inhibited by depletion of the autophagy regulator, ATG5 or SQSTM1. In addition, treatment of triamterene decreased the integrity of lysosome and cell viability, which were rescued by removing the triamterene treatment in HepG2 cells. Hence, our data suggest that triamterene is a novel lysophagy inducer through the disruption of lysosomal integrity.