Search

The presence of cancer stem cells (CSC), which possess the ability of self-renewal and cancer initiation, is correlated with poor prognosis and drug resistance of breast cancer patients. But the molecular regulatory networks for maintenance of CSC function still remain unclear. Here, we identified that an estrogen-inducible gene FXYD3, whose expression is significantly upregulated in ER+ breast CSCs, is a critical player for regulating ER+ breast CSC function. FXYD3 amplification is crucial in mediating tamoxifen resistance in ER+ breast cancer cells. Interestingly, we also find that stem cell–related transcription factor SOX9 directly promotes FXYD3 expression, and FXYD3 is indispensable for SOX9 nucleus localization, thus forming a positive regulatory feedback loop for FXYD3 amplification and function. In terms of mechanism, FXYD3 interacts with Src and ERα to form an activated complex and triggers Src to transduce nongenomic estrogen signaling for facilitating ER+ breast CSCs. Collectively, these results establish a critical role for SOX9/FXYD3/Src axis in boosting nongenomic estrogen signaling and SOX9 nucleus entry, which is required for maintenance of ER+ breast CSCs and endocrine resistance. Targeting FXYD3-mediated pathway might be a promising therapeutic strategy for hormone therapy–refractory ER+ breast cancer.Implications:SOX9/FXYD3/Src axis is critical for promoting CSC function and tamoxifen resistance in ER+ breast cancer.

S1. FXYD3 is required for the maintenance of the ER+ cancer stem cells. S2. A) Immunohistochemistry of FXYD3 in a tissue microarray (TMA) of human breast cancers showed representative examples of strong (score 4), moderate (score 3), weak (score 2), and negative (score 1) staining. Scale bar, 100 μm (upper), 20 μm (lower). B) Bioinformatics analysis of the public dataset (GEO: GSE75688) for the expression of FXYD3 based on the indicated stratifications (unpaired t test, *p

Gastric cancer (GC) patients with metastasis had limited treatment options and dismal outcome. We have previously reported the aberrant expression of Zic family member 1 (Zic1) in GC. However, the functional roles and underlying mechanism of Zic1 in GC metastasis remain unknown. Here, we demonstrate that lower expression of Zic1 was correlated with more lymph node metastasis and poor outcome of GC patients. Ectopic expression of Zic1 suppressed both lung metastasis and peritoneal tumor dissemination of GC in mice. The metastatic suppressing ability of Zic1 was mediated by regulating the process of cell invasion, adhesion and epithelial-mesenchymal transition (EMT). Mechanistically, Zic1 could downregulate Wnt targets including c-Myc and Cyclin D1 by inhibiting LEF transcriptional activity in GC cells. Notably, Zic1 was inversely related to the expression of Cyclin D1 in GC tissues tested. In addition, Zic1 could physically interact with β-catenin/transcription factor 4 (TCF4) and disrupt their complex formation, while not affecting β-catenin nuclear localization. Collectively, our study indicated that Zic1 suppressed GC metastasis through attenuating Wnt/β-catenin signaling and the EMT process. Our work may provide novel therapeutic strategies for the metastasis of GC.

Resident and inflammatory mononuclear phagocytes (MPhs) with functional plasticity in the intestine are critically involved in the pathology of inflammatory bowel diseases (IBDs), the mechanism of which remains incompletely understood. In the present study, we found that increased expression of the E3 ligase F-box and WD repeat domain–containing 7 (FBXW) in the inflamed intestine was significantly correlated with IBD severity in both human diseases and in mouse models. Myeloid Fbxw7 deficiency protected mice from colitis induced by dextran sodium sulfate (DSS) or 2,6,4-trinitrobenzene sulfonic acid (TNBS). Fbxw7 deficiency resulted in decreased production of the chemokines CCL2 and CCL7 by colonic CX3CR1(hi) resident macrophages and reduced the accumulation of CX3CR1(int) proinflammatory MPhs in colitis-affected colon tissue. Mice that received adeno-associated virus–shFbxw7 (AAV-shFbxw7) showed significantly improved survival rates and alleviation of colitis. Mechanism screening demonstrated that FBXW7 suppressed H3K27me3 modification and promoted Ccl2 and Ccl7 expression via degradation of the histone-lysine N-methyltransferase enhancer of zeste homolog 2 (EZH2) in macrophages. Taken together, our results indicate that FBXW7 degrades EZH2 and increases Ccl2 and Ccl7 in CX3CR1(hi) macrophages, thereby promoting the recruitment of CX3CR1(int) proinflammatory MPhs into local colon tissues with colitis. Targeting FBXW7 might represent a potential therapeutic approach for the treatment of intestinal inflammation.

The classic mode of STING activation is through binding the cyclic dinucleotide 2′3′-cyclic GMP–AMP (cGAMP), produced by the DNA sensor cyclic GMP–AMP synthase (cGAS), which is important for the innate immune response to microbial infection and autoimmune disease. Modes of STING activation that are independent of cGAS are much less well understood. Here, through a spatiotemporally resolved proximity labelling screen followed by quantitative proteomics, we identify the lysosomal membrane protein Niemann–Pick type C1 (NPC1) as a cofactor in the trafficking of STING. NPC1 interacts with STING and recruits it to the lysosome for degradation in both human and mouse cells. Notably, we find that knockout of Npc1 ‘primes’ STING signalling by physically linking or ‘tethering’ STING to SREBP2 trafficking. Loss of NPC1 protein also ‘boosts’ STING signalling by blocking lysosomal degradation. Both priming and boosting of STING signalling are required for severe neurological disease in the Npc1−/− mouse. Genetic deletion of Sting1 (the gene that encodes STING) or Irf3, but not that of Cgas, significantly reduced the activation of microglia and relieved the loss of Purkinje neurons in the cerebellum of Npc1−/− mice, leading to improved motor function. Our study identifies a cGAS- and cGAMP-independent mode of STING activation that affects neuropathology and provides a therapeutic target for the treatment of Niemann–Pick disease type C. The lysosomal protein Niemann–Pick type C1 (NPC1) is identified as a cofactor in the trafficking of STING, and loss of NPC1 is shown to both ‘prime’ and ‘boost’ STING signalling leading to neurological disease.

The presence of cancer stem cells (CSC), which possess the ability of self-renewal and cancer initiation, is correlated with poor prognosis and drug resistance of breast cancer patients. But the molecular regulatory networks for maintenance of CSC function still remain unclear. Here, we identified that an estrogen-inducible gene FXYD3, whose expression is significantly upregulated in ER+ breast CSCs, is a critical player for regulating ER+ breast CSC function. FXYD3 amplification is crucial in mediating tamoxifen resistance in ER+ breast cancer cells. Interestingly, we also find that stem cell–related transcription factor SOX9 directly promotes FXYD3 expression, and FXYD3 is indispensable for SOX9 nucleus localization, thus forming a positive regulatory feedback loop for FXYD3 amplification and function. In terms of mechanism, FXYD3 interacts with Src and ERα to form an activated complex and triggers Src to transduce nongenomic estrogen signaling for facilitating ER+ breast CSCs. Collectively, these results establish a critical role for SOX9/FXYD3/Src axis in boosting nongenomic estrogen signaling and SOX9 nucleus entry, which is required for maintenance of ER+ breast CSCs and endocrine resistance. Targeting FXYD3-mediated pathway might be a promising therapeutic strategy for hormone therapy–refractory ER+ breast cancer. Implications: SOX9/FXYD3/Src axis is critical for promoting CSC function and tamoxifen resistance in ER+ breast cancer.

STING (stimulator of interferon genes) is an important innate immune protein, but its homeostatic regulation at the resting state is unknown. Here, we identified TOLLIP as a stabilizer of STING through direct interaction to prevent its degradation. Tollip deficiency results in reduced STING protein in nonhematopoietic cells and tissues, and renders STING protein unstable in immune cells, leading to severely dampened STING signaling capacity. The competing degradation mechanism of resting-state STING requires IRE1α and lysosomes. TOLLIP mediates clearance of Huntington's disease-linked polyQ protein aggregates. Ectopically expressed polyQ proteins in vitro or endogenous polyQ proteins in Huntington's disease mouse striatum sequester TOLLIP away from STING, leading to reduced STING protein and dampened immune signaling. Tollip-/- also ameliorates STING-mediated autoimmune disease in Trex1-/- mice. Together, our findings reveal that resting-state STING protein level is strictly regulated by a constant tug-of-war between 'stabilizer' TOLLIP and 'degrader' IRE1α-lysosome that together maintain tissue immune homeostasis.

STING is an endoplasmic reticulum (ER)-associated transmembrane protein that is essential for the host innate immune response to pathogens. STING is activated by the second messenger cyclic dinucleotides which is produced from the invading pathogen or by the host DNA sensor cGAS. Activation of STING triggers its trafficking from the ER, through ERGIC/Golgi, to vesicles, during which time STING activates and quickly terminates downstream interferon signaling. Despite the importance of STING signaling in infection, autoimmune disease and cancer immunotherapy, the dynamics of STING trafficking and kinetics of signaling are poorly understood. Here, we performed a time-resolved proteomic screen to capture protein interaction networks of STING during its trafficking process. We also established a synchronized live-cell microscopy technique that allows us to define changes in protein trafficking kinetics. We have identified several co-factor candidates that affect various steps of the STING trafficking and signaling. Collectively, our findings revealed the previously unanticipated the regulation of STING trafficking as well as its downstream signaling by the host factors in the secretory pathway.

Malignant gliomas are the most common primary brain tumors, and novel ways of treating gliomas are urgently needed. Ursolic acid (UA), a pentacyclic triterpenoid, has been reported to exhibit promising antitumor activity. Here, we evaluated the effects of UA on U87MG cells and explored the underlying molecular mechanisms. The results demonstrated that both G1-phase arrest and autophagy were induced by UA in U87MG cells. Evidence of UA-induced autophagy included the formation of acidic vesicular organelles, increase of autophagolysosomes and LC3-II accumulation. UA was also found to induce ER stress and an increase in intracellular calcium accompanied by ROS production. The increase in free cytosolic calcium induced by UA activated the CaMKK-AMPK-mTOR kinase signaling cascade, which ultimately triggered autophagy. Western blot analysis showed that UA promoted the phosphorylation of PERK and eIF2α; this was followed by the upregulation of the downstream protein CHOP, implying the involvement of the ER stress-mediated PERK/eIF2α/CHOP pathway in glioma cells. Meanwhile, UA activated IRE1α and subsequently increased the levels of phosphorylated JNK and Bcl-2, resulting in the dissociation of Beclin1 from Bcl-2. Furthermore, TUDCA and the silencing of either PERK or IRE1α partially blocked the UA-induced accumulation of LC3-II, suggesting that ER stress precedes the process of autophagy. Additionally, NAC attenuated the UA-induced elevation in cytosolic calcium, ER stress markers and autophagy-related proteins, indicating that UA triggered ER stress and autophagy via a ROS-dependent pathway. Collectively, our findings revealed a novel cellular mechanism triggered by UA and provide a molecular basis for developing UA into a drug candidate.