TMEM216 promotes primary ciliogenesis and Hedgehog signaling through the SUFU-GLI2/GLI3 axis.

Primary cilia are enriched in signaling receptors, and defects in their formation or function can induce conditions such as polycystic kidney disease, postaxial hexadactyly, and microphthalmia. Mammalian Hedgehog (Hh) signaling is important in the development of primary cilia, and TMEM216, a transmembrane protein that localizes to the base of cilia, is also implicated in ciliogenesis in zebrafish. Here, we found that Tmem216-deficient mice had impaired Hh signaling and displayed typical ciliopathic phenotypes. These phenomena were also observed in cells deficient in TMEM216. Furthermore, TMEM216 interacted with core Hh signaling proteins, including SUFU, a negative regulator of Hh, and GLI2/GLI3, transcription factors downstream of Hh. The competition between TMEM216 and SUFU for binding to GLI2/GLI3 inhibited the cleavage of GLI2/GLI3 into their repressor forms, which resulted in the nuclear accumulation of full-length GLI2 and the decreased nuclear localization of cleaved GLI3, ultimately leading to the activation of Hh signaling. Together, these data suggest that the TMEM216-SUFU-GLI2/GLI3 axis plays a role in TMEM216 deficiency-induced ciliopathies and Hh signaling abnormalities.

Crotonylated BEX2 interacts with NDP52 and enhances mitophagy to modulate chemotherapeutic agent-induced apoptosis in non-small-cell lung cancer cells.

Brain expressed X-linked gene 2 (BEX2) encoded protein was originally identified to promote transcription by interacting with several transcription factors in the DNA-binding complexes. Recently, BEX2 was found to be localized in cytosol and/or mitochondria and regulate apoptosis in cancer cells and tumor growth. However, the molecular mechanism underlying its roles in cancer cells remains unclear. Here, we report that crotonylated BEX2 plays an important role in inhibiting chemotherapeutic agent-induced apoptosis via enhancing mitophagy in human lung cancer cells. BEX2 promotes mitophagy by facilitating interaction between NDP52 and LC3B. Moreover, BEX2 crotonylation at K59 is critical in the BEX2-mediated mitophagy in lung cancer cells. The K59R mutation of BEX2 inhibits mitophagy by affecting the interaction of NDP52 and LC3B. BEX2 expression is elevated after anticancer drug treatment, and its overexpression inhibits chemotherapy-induced apoptosis. In addition, inhibition of BEX2-regulated mitophagy sensitizes tumor cells to apoptosis. Furthermore, BEX2 promotes tumor growth and inhibits apoptosis by regulating mitophagy in vivo. We also confirm that BEX2 is overexpressed in lung adenocarcinoma and is associated with poor prognosis in lymph node metastasis-free cancer. Therefore, combination treatment with pharmaceutical approaches targeting BEX2-induced mitophagy and anticancer drugs may represent a potential strategy for NSCLC therapy.

DIRAS3 enhances RNF19B-mediated RAC1 ubiquitination and degradation in non-small-cell lung cancer cells.

Distant metastasis remains the leading cause of high mortality in patients with non-small-cell lung cancer (NSCLC). DIRAS3 is a candidate tumor suppressor protein that is decreased in various tumors. However, the regulatory mechanism of DIRAS3 on metastasis of NSCLC remains unclear. Here, we found that DIRAS3 suppressed the migration of NSCLC cells. Besides, DIRAS3 stimulated the polyubiquitination of RAC1 and suppressed its protein expression. Furthermore, RNF19B, a member of the RBR E3 ubiquitin ligase family, was observed to be the E3 ligase involved in the DIRAS3-induced polyubiquitination of RAC1. DIRAS3 could promote the binding of RAC1 and RNF19B, thus enhancing the degradation of RAC1 by the ubiquitin-proteasome pathway. Finally, the DIRAS3-RNF19B-RAC1 axis was confirmed to be associated with the malignant progression of NSCLC. These findings may be beneficial for developing potential prognostic markers of NSCLC and may provide an effective treatment strategy.

Gasdermin E regulates the stability and activation of EGFR in human non-small cell lung cancer cells.

BACKGROUND: Lung cancer is the most lethal malignancy, with non-small cell lung cancer (NSCLC) being the most common type (~ 85%). Abnormal activation of epidermal growth factor receptor (EGFR) promotes the development of NSCLC. Chemoresistance to tyrosine kinase inhibitors, which is elicited by EGFR mutations, is a key challenge for NSCLC treatment. Therefore, more thorough understanding of EGFR expression and dynamics are needed. METHODS: Human non-small cell lung cancer cells and HEK293FT cells were used to investigate the molecular mechanism of gasdermin E (GSDME) regulating EGFR stability by Western blot analysis, immunoprecipitation and immunofluorescence. GSDME and EGFR siRNAs or overexpression plasmids were used to characterize the functional role of GSDME and EGFR in vitro. EdU incorporation, CCK-8 and colony formation assays were used to determine the proliferation ability of non-small cell lung cancer cells. RESULTS: GSDME depletion reduced the proliferation of non-small cell lung cancer cells in vitro. Importantly, both GSDME-full length (GSDME-FL) and GSDME-N fragment physically interacted with EGFR. GSDME interacted with cytoplasmic fragment of EGFR. GSDME knockdown inhibited EGFR dimerization and phosphorylation at tyrosine 1173 (EGFRY1173), which activated ERK1/2. GSDME knockdown also promoted phosphorylation of EGFR at tyrosine 1045 (EGFRY1045) and its degradation. CONCLUSION: These results indicate that GSDME-FL increases the stability of EGFR, while the GSDME N-terminal fragment induces EGFR degradation. The GSDME-EGFR interaction plays an important role in non-small cell lung cancer development, reveal a previously unrecognized link between GSDME and EGFR stability and offer new insight into cancer pathogenesis. Video abstract.

Circuit theory-based ecological security pattern could promote ecological protection in the Heihe River Basin of China.

Building ecological security patterns is essential to maintain regional ecological security and achieve sustainable development in the inland river basins with ecologically vulnerable environment. Numerous methods have been developed to build the ecological security pattern. However, to our knowledge, rare studies have quantified to what extent the derived pattern can improve ecological protection in the future. Taking Heihe River Basin (HRB), the second largest inland river basin in China, as the study area, we applied the circuit theory to build the ecological security pattern of HRB, and simulated how our built pattern contributed to ecological protection using the CLUMondo model. The results showed that the ecological security pattern of HRB contained 17 ecological sources, 35 key ecological corridors, and some ecological strategic points. The ecological sources were distributed in areas with better ecological conditions such as the Qilian Mountain Nature Reserve and Heihe National Wetland Park. The ecological corridors showed a pattern of "two horizontal and three vertical belts." Pinch points were mostly close to ecological sources or distributed on the corridors that played a key role in landscape connectivity, while barriers were mainly distributed on the corridors with large ecological resistance in the middle and lower reaches. The optimal ecological security pattern presented a "one screen, one belt, four districts and multiple centers" shape in HRB and could more effectively promote ecological protection compared to current development and protection scenarios. Our study provides a reliable decision-making guide for ecological protection and restoration of HRB, and can be extended to build ecological security patterns for broad-scale arid areas.

Cancer cell-targeted nanoprobe for multilayer imaging of diverse biomarkers and precise photodynamic therapy.

A novel multifunctional nanoprobe was designed for cancer cell targeted multilayer imaging of two cancer biomarkers. Based on the proposed method, in situ imaging of membrane MUC1 mucin and cytoplasmic microRNA miR-21 coupled with precise photodynamic therapy was achieved.

A seesaw ratiometric probe for dual-spectrum imaging and detection of telomerase activity in single living cells.

Herein, a seesaw ratiometric (SR) probe is designed which integrates fluorescence and surface enhanced Raman scattering (SERS) technology. Fluorescence imaging enables tracking of the spatiotemporal dynamic behaviour of telomerase. Meanwhile, SERS reverse ratiometric measurement can enable sensitive detection of telomerase activity in single living cells.

Telomere elongation-based DNA-Catalytic amplification strategy for sensitive SERS detection of telomerase activity.

Telomerase has been regarded as a biomarker for cancer diagnosis as well as the clinical treatment and the reliable detection of intracellular telomerase activity is of great significance. By developing a telomere elongation-based DNA-catalytic amplification strategy, a novel surface-enhanced Raman scattering (SERS) method is proposed for the assay of telomerase activity. In the presence of telomerase and nucleotide mixture dNTPs, the telomerase substrate (TS) primer extended and generated a long single-strand DNA (ssDNA) containing the telomere repeat units (TTAGGG)n, which could catalyze the entropy-driven circuit reaction (EDCR). One of the products of EDCR was ingeniously used as the catalyst of catalytic hairpin assembly (CHA) occured on magnetic beads (MBs). As a result, a large amount of ROX-labeled Raman probes could be anchored on the surface of MBs and used for SERS detection. Using this strategy, the assay can detect telomerase activity from cell extracts equivalent down to single HeLa cell.

Dual cycle amplification and dual signal enhancement assisted sensitive SERS assay of MicroRNA.

A sensitive surface-enhanced Raman scattering (SERS) approach has been developed for detection of microRNA (miRNA) based on target-triggered dual signal amplification including strand displancement amplification (SDA) and hybridization chain reaction (HCR). With the assistant of polymerase and nicking endonuclease (NEase), target miRNA combines with the single stranded template DNA to generate a great amount of trigger DNA which can induce HCR. Coupled the dual cycle amplification of SDA and HCR with the dual enhancement of gold nanoparticles (AuNPs), a low detection limit of 0.5 fM for miRNA is obtained using the proposed strategy. With high sensitivity, universality, rapid analysis, and high selectivity, this method has a great potential for detecting biomolecules with trace amounts in bioanalysis and clinical biomedicine.