IRG1 restrains M2 macrophage polarization and suppresses intrahepatic cholangiocarcinoma progression via the CCL18/STAT3 pathway.

Intrahepatic cholangiocarcinoma (ICC) is a highly malignant and aggressive cancer whose incidence and mortality continue to increase, whereas its prognosis remains dismal. Tumor-associated macrophages (TAMs) promote malignant progression and immune microenvironment remodeling through direct contact and secreted mediators. Targeting TAMs has emerged as a promising strategy for ICC treatment. Here, we revealed the potential regulatory function of immune responsive gene 1 (IRG1) in macrophage polarization. We found that IRG1 expression remained at a low level in M2 macrophages. IRG1 overexpression can restrain macrophages from polarizing to the M2 type, which results in inhibition of the proliferation, invasion, and migration of ICC, whereas IRG1 knockdown exerts the opposite effects. Mechanistically, IRG1 inhibited the tumor-promoting chemokine CCL18 and thus suppressed ICC progression by regulating STAT3 phosphorylation. The intervention of IRG1 expression in TAMs may serve as a potential therapeutic target for delaying ICC progression.

TMEM147 aggravates the progression of HCC by modulating cholesterol homeostasis, suppressing ferroptosis, and promoting the M2 polarization of tumor-associated macrophages.

BACKGROUND: The endoplasmic reticulum (ER) regulates critical processes, including lipid synthesis, which are affected by transmembrane proteins localized in the ER membrane. One such protein, transmembrane protein 147 (TMEM147), has recently been implicated for its role in hepatocellular carcinoma (HCC) tumorigenesis; however, the mechanisms remain unclear. We investigated the role of TMEM147 in HCC and the underlying mechanisms. METHODS: TMEM147 expression was examined in human HCC cells and adjacent non-tumorous tissues using quantitative reverse transcription-polymerase chain reaction, western blotting, and immunohistochemistry. In vitro and in vivo studies were conducted to investigate the impact of TMEM147 on the progression of HCC. Proteins interacting with TMEM147 were identified via RNA-seq, immunoprecipitation, and mass spectrometry analyses. Lipidomic analysis and enzyme-linked immunosorbent assay (ELISA) were employed to determine and analyze cholesterol and 27-hydroxycholesterol (27HC) contents. Extensive experimental techniques were used to study ferroptosis in HCC cells. The fatty acid content of macrophages affected by TMEM147 was quantified using ELISA. Macrophage phenotypes were determined using immunofluorescence assay and flow cytometric analysis. RESULTS: TMEM147 mRNA and protein levels were increased in HCC cells, and the increased TMEM147 expression was associated with a poor survival. TMEM147 promoted tumor cell proliferation and metastases in vitro and in vivo. The protein was found to interact with the key enzyme 7-dehydrocholesterol reductase (DHCR7), which affected cellular cholesterol homeostasis and increased the extracellular levels of 27HC in HCC cells. TMEM147 also promoted the expression of DHCR7 by enhancing the activity of signal transducer and activator of transcription 2. 27HC expression upregulated glutathione peroxidase 4 in HCC, leading to ferroptosis resistance and promotion of HCC proliferation. HCC cell-derived 27HC expression increased the lipid metabolism in macrophages and activated peroxisome proliferator-activated receptor-γ signaling, thereby activating M2 macrophage polarization and promoting HCC cell invasion and migration. CONCLUSIONS: Our results indicate that TMEM147 confers ferroptosis resistance and M2 macrophage polarization, which are primarily dependent on the upregulation of cellular cholesterol homeostasis and 27HC secretion, leading to cancer growth and metastasis. These findings suggest that the TMEM147/STAT2/DHCR7/27HC axis in the tumor microenvironment may serve as a promising therapeutic target for HCC.

Geochemical behaviors of uranium and thorium during weathering and pedogenesis of carbonate rock: constraint from their speciation.

During weathering and pedogenesis of carbonate rock with poor-uranium (U) and thorium (Th), U and Th present the characteristics of strong leaching (especially U) and significant residual enrichment, the cause of which is still unclear. In this paper, a weathering profile developed by dolomite in karst area of Guizhou province in southwest China was selected, which showed zonation characteristics of bedrock (Y), powdery rock (Yf), and soil layer (T1 to T12) from the bottom to up. Through the determination of the occurrence speciation of U and Th in Y and weathering profile, combined with mineralogical, geochemical characteristics, and element mass balance calculation, the constraints of U and Th speciation on the geochemical behavior of U and Th during the weathering of carbonate rock were revealed. The results proved that U and Th in Y preferentially existed in acid insoluble phase, for example, the contents of U and Th in Y were 0.90 mg·kg-1 and 0.28 mg·kg-1, respectively, while those in acid insoluble matter were 2.34 mg·kg-1 and 2.57 mg·kg-1, respectively, but because the mass percentage of acid insoluble matter was extremely low (0.95%), the mass percentages of U and Th in the acid soluble phase in the whole rock were absolutely superior (96% of U and 86% Th). The U and Th in the acid soluble phase of Y were mainly adsorbed on the crystal surface of carbonate minerals or existed in the cement, and the U and Th in the carbonate lattice only accounted for a small proportion. From Y to Yf with the initial dissolution, U and Th released from the surface of carbonate minerals and cements were in carbonate-rich alkaline environment, and these portions of U and Th were leached out, resulting in strong loss of U and Th in the Yf (the loss rates are 83% of U and 65% of Th, respectively). From the Yf to the overlying soil layer T1, the carbonate components were completely dissolved, and the U and Th released from the carbonate lattice showed different behaviors, where U was completely leached and Th tended to stay in the weathered residue. Thus, in the soil layer T1 formed by Y or Yf , the residual U was the inheritance of the U in the acid insoluble phase of Y; For Th, it not only inherited the Th of acid insoluble phase of Y, but also superimposed the Th from carbonate lattice in Y. On the other hand, during the evolution process from Y to Yf and to soil layer T1, with the dissolution of carbonate, the acid insoluble phase also showed a significant tendency of chemical weathering. However, the U and Th in the Y acid insoluble phase were not leached with the decomposition of the acid insoluble phase but were redistributed among the residual phases. For the geochemical behaviors of U and Th in the evolution of soil profile (T1~T12), they were subjected to the occurrence speciation of U and Th in T1 and the change of U and Th occurrence speciation with the upward direction of soil profile. The U and Th released from the carrier minerals were mainly redistributed among the residual solid phases, which weakened the intensity of their further loss. This study deepens the understanding of the geochemical behavior of radionuclides in karst environment and provides reference for the treatment of radioactive pollution in karst areas.

RNF125 attenuates hepatocellular carcinoma progression by downregulating SRSF1-ERK pathway.

Hepatocellular carcinoma (HCC) is one of the most deadly malignant cancers worldwide. Research into the crucial genes responsible for maintaining the aggressive behaviour of cancer cells is important for the clinical treatment of HCC. The purpose of this study was to determine whether the E3 ubiquitin ligase Ring Finger Protein 125 (RNF125) plays a role in the proliferation and metastasis of HCC. RNF125 expression in human HCC samples and cell lines was investigated using TCGA dataset mining, qRT‒PCR, western blot, and immunohistochemistry assays. In addition, 80 patients with HCC were studied for the clinical value of RNF125. Furthermore, the molecular mechanism by which RNF125 contributes to hepatocellular carcinoma progression was determined with mass spectrometry (MS), coimmunoprecipitation (Co-IP), dual-luciferase reporter assays, and ubiquitin ladder assays. We found that RNF125 was markedly downregulated in HCC tumour tissues, which was associated with a poor prognosis for patients with HCC. Moreover, the overexpression of RNF125 inhibited HCC proliferation and metastasis both in vitro and in vivo, whereas the knockdown of RNF125 exerted antithetical effects. Mechanistically, mass spectrometry analysis revealed a protein interaction between RNF125 and SRSF1, and RNF125 accelerated the proteasome-mediated degradation of SRSF1, which impeded HCC progression by inhibiting the ERK signalling pathway. Furthermore, RNF125 was detected to be the downstream target of miR-103a-3p. In this study, we identified that RNF125 is a tumour suppressor in HCC and inhibits HCC progression by inhibiting the SRSF1/ERK pathway. These findings provide a promising treatment target for HCC.

Comprehensive analysis of transcriptome-wide M6A methylation for hepatic ischaemia reperfusion injury in mice.

N6-Methyladenosine (m6A) plays key roles in the regulation of biological functions and cellular mechanisms for ischaemia reperfusion (IR) injury in different organs. However, little is known about the underlying mechanisms of m6A-modified mRNAs in hepatic IR injury. In mouse models, liver samples were subjected to methylated RNA immunoprecipitation with high-throughput sequencing (MeRIP-seq) and RNA sequencing (RNA-seq). In total, 16917 m6A peaks associated with 4098 genes were detected in the sham group, whereas 21,557 m6A peaks associated with 5322 genes were detected in the IR group. There were 909 differentially expressed m6A peaks, 863 differentially methylated transcripts and 516 differentially m6A modification genes determined in both groups. The distribution of m6A peaks was especially enriched in the coding sequence and 3'UTR. Furthermore, we identified a relationship between differentially m6A methylated genes (fold change≥1.5/≤ 0.667, p value≤0.05) and differentially expressed genes (fold change≥1.5 and p value≤0.05) to obtain three overlapping predicted target genes (Fnip2, Phldb2, and Pcf11). Our study revealed a transcriptome-wide map of m6A mRNAs in hepatic IR injury and might provide a theoretical basis for future research in terms of molecular mechanisms.

Atom-based sensing technique of microwave electric and magnetic fields via a single rubidium vapor cell.

We present an atom-based approach for determining microwave electric and magnetic fields by using a single rubidium vapor cell in a microwave waveguide. For a 87Rb cascade three-level system employed in our experiment, a weak probe laser driving the lower transition, 5S1/2→5P3/2, is first used to measure the microwave magnetic field based on the atomic Rabi resonance. When a counter-propagating strong coupling laser is subsequently turned on to drive the Rydberg transition, 5P3/2→67D5/2, the same probe laser is then used as a Rydberg electromagnetically induced transparency (EIT) probe to measure the microwave electric field by investigating the resonant microwave dressed Autler-Townes splitting (ATS). By tuning the hyperfine transition frequency of the ground state using an experimentally feasible static magnetic field, we first achieved a measurement of the microwave electric and magnetic field strength at the same microwave frequency of 6.916 GHz. Based on the ideal relationship between the electric and magnetic field components, we obtained the equivalent microwave magnetic fields by fitting the inversion to the measured microwave electric fields, which demonstrated that the results were in agreement with the experimental measurement of the microwave magnetic fields in the same microwave power range. This study provides new experimental evidence for quantum-based microwave measurements of electric and magnetic fields by a single sensor in the same system.

HLTF promotes hepatocellular carcinoma progression by enhancing SRSF1 stability and activating ERK/MAPK pathway.

Helicase-like transcription factor (HLTF) has been found to be involved in the progression of several tumors, but the role of HLTF in hepatocellular carcinoma (HCC) progression has not been studied. Here, our study explored the underlying mechanism of HLTF in HCC progression for the first time. Database analysis and clinical sample examination indicated that HLTF was upregulated in HCC tissues and was related to poor clinicopathological features in patients. Upregulation of HLTF accelerated the growth and metastasis of HCC cells both in vitro and in vivo. Bioinformatics analysis and subsequent experiments revealed that ERK/MAPK signaling pathway activation was vital to HLTF-mediated proliferation and metastasis in HCC cells. Moreover, HLTF was demonstrated to interact with SRSF1 and contribute to its protein stability to activate the ERK/MAPK signaling pathway and enhance HCC growth and metastasis. In addition, miR-511-5p was expressed at a low level in HCC tissues, was negatively correlated HLTF, and regulated HLTF expression. Our study shows that HLTF plays an oncogenic role in HCC progression and provides a novel biomarker and therapeutic target for the diagnosis and treatment of HCC.

Comprehensive analysis of the prognostic value and functions of prefoldins in hepatocellular carcinoma.

Prefoldins (PFDNs), a group of proteins known to be associated with cytoskeletal rearrangement, are involved in tumor progression in various cancer types. However, little is known about the roles of PFDNs in hepatocellular carcinoma (HCC). Herein, we investigated the transcriptional and survival data of PFDNs from The Cancer Genome Atlas (TCGA) database. Gene Ontology (GO), Gene Set Enrichment Analysis (GSEA), and single-sample gene set enrichment analysis (ssGSEA) were used to evaluate the potential functions of PFDN1/2/3/4. We also detected the expression of PFDN1/2/3/4 via immunohistochemistry (IHC), Western blotting, and real-time PCR in our clinical samples. We found that the PFDN family showed elevated expression in HCC tissues, while only PFDN1/2/3/4 were found to be significantly correlated with poor prognosis of patients with HCC in the TCGA database. Further investigation was associated with PFDN1-4. We found that the expression of PFDN1/2/3/4 was significantly associated with advanced clinicopathologic features. Apart from the TCGA database, IHC, real-time PCR, and immunoblotting identified the overexpression of PFDN1/2/3/4 in HCC tissues and HCC cell lines. Taken together, these results indicated that PFDN1/2/3/4 might be novel prognostic biomarkers and treatment targets for patients with HCC.

LncRNA Hnf4αos exacerbates liver ischemia/reperfusion injury in mice via Hnf4αos/Hnf4α duplex-mediated PGC1α suppression.

LncRNAs are involved in the pathophysiologic processes of multiple diseases, but little is known about their functions in hepatic ischemia/reperfusion injury (HIRI). As a novel lncRNA, the pathogenetic significance of hepatic nuclear factor 4 alpha, opposite strand (Hnf4αos) in hepatic I/R injury remains unclear. Here, differentially expressed Hnf4αos and Hnf4α antisense RNA 1 (Hnf4α-as1) were identified in liver tissues from mouse ischemia/reperfusion models and patients who underwent liver resection surgery. Hnf4αos deficiency in Hnf4αos-KO mice led to improved liver function, alleviated the inflammatory response and reduced cell death. Mechanistically, we found a regulatory role of Hnf4αos-KO in ROS metabolism through PGC1α upregulation. Hnf4αos also promoted the stability of Hnf4α mRNA through an RNA/RNA duplex, leading to the transcriptional activation of miR-23a and miR-23a depletion was required for PGC1α function in hepatoprotective effects on HIRI. Together, our findings reveal that Hnf4αos elevation in HIRI leads to severe liver damage via Hnf4αos/Hnf4α/miR-23a axis-mediated PGC1α inhibition.

NNMT promotes the progression of intrahepatic cholangiocarcinoma by regulating aerobic glycolysis via the EGFR-STAT3 axis.

Nicotinamide N-methyltransferase (NNMT), a member of the N-methyltransferase family, plays an important role in tumorigenesis. However, its expression and biological functions in intrahepatic cholangiocarcinoma (iCCA) remain to be established. In our study, we identified NNMT as an oncogene in iCCA and provided mechanistic insights into the roles of NNMT in iCCA progression. High NNMT expression in iCCA tissues was identified using western blotting and immunohistochemistry (IHC). We identified a significantly higher NNMT expression level in human iCCA tissues than that in adjacent normal tissues. Increased NNMT expression promoted iCCA cell proliferation and metastasis in vitro and in vivo. Mechanistically, NNMT inhibited the level of histone methylation in iCCA cells by consuming the methyl donor S-adenosyl methionine (SAM), thereby promoting the expression of epidermal growth factor receptor (EGFR). EGFR may activate the aerobic glycolysis pathway in iCCA cells by activating the STAT3 signaling pathway. In conclusion, we identified NNMT as an oncogene in iCCA and provided mechanistic insights into the roles of NNMT in iCCA progression.

Identification of CFHR4 as a Potential Prognosis Biomarker Associated With lmmune Infiltrates in Hepatocellular Carcinoma.

Background: Complement factor H-related 4 (CFHR4) is a protein-coding gene that plays an essential role in multiple diseases. However, the prognostic value of CFHR4 in hepatocellular carcinoma (HCC) is unknown. Methods: Using multiple databases, we investigated CFHR4 expression levels in HCC and multiple cancers. The relationship between CFHR4 expression levels and clinicopathological variables was further analyzed. Various potential biological functions and regulatory pathways of CFHR4 in HCC were identified by performing a Gene Ontology (GO) analysis, Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis and Gene Set Enrichment Analysis (GSEA). Single-sample gene set enrichment analysis (ssGSEA) was performed to confirm the correlation between CFHR4 expression and immune cell infiltration. The correlations between CFHR4 expression levels in HCC and N6-methyladenosine (m6A) modifications and the competing endogenous RNA (ceRNA) regulatory networks were confirmed in TCGA cohort. Results: CFHR4 expression levels were significantly decreased in HCC tissues. Low CFHR4 expression in HCC tissues was significantly correlated with the patients' sex, race, age, TNM stage, pathological stage, tumor status, residual tumor, histologic grade and alpha fetal protein (AFP) level. GO and KEGG analyses revealed that differentially expressed genes related to CFHR4 may be involved in the synaptic membrane, transmembrane transporter complex, gated channel activity, chemical carcinogenesis, retinol metabolism, calcium signaling pathway, PPAR signaling pathway, insulin and gastric acid secretion. GSEA revealed that the FCGR-activated reaction, PLK1 pathway, ATR pathway, MCM pathway, cascade reactions of PI3K and FGFR1, reactant-mediated MAPK activation and FOXM1 pathway were significantly enriched in HCC with low CFHR4 expression. Moreover, CFHR4 expression was inversely correlated the levels of infiltrating Th2 cells, NK CD56bright cells and Tfh cells. In contrast, we observed positive correlations with the levels of infiltrating DCs, neutrophils, Th17 cells and mast cells. CFHR4 expression showed a strong correlation with various immunomarker groups in HCC. In addition, high CFHR4 expression significantly prolonged the overall survival (OS), disease-specific survival (DSS) and progression-free interval (PFI). We observed a substantial correlation between the expression of CFHR4 and multiple N6-methyladenosine genes in HCC and constructed potential CFHR4-related ceRNA regulatory networks. Conclusions: CFHR4 might be a potential therapeutic target for improving the HCC prognosis and is closely related to immune cell infiltration.

Live imaging of postembryonic developmental processes in C. elegans.

Live imaging is an important tool to track dynamic processes such as neuronal patterning events. Here, we describe a protocol for time-lapse microscopy analysis using neuronal migration and dendritic growth as examples. This protocol can provide detailed information for understanding cellular dynamics during postembryonic development in Caenorhabditis elegans (C. elegans). For complete details on the use and execution of this protocol, please refer to Feng et al. (2020), Li et al. (2021), and Wang et al. (2021).

Serum amyloid A-to-albumin ratio as a potential biomarker to predict the activity, severity, and poor prognosis of systemic lupus erythematosus.

OBJECTIVES: To evaluate the predictive value of serum amyloid A-to-albumin ratio (SAR) for active systemic lupus erythematosus (SLE), severe active SLE, and poor prognosis of SLE. METHODS: One hundred and eighty-six patients with SLE undergoing treatment in our hospital were selected. The demographic characteristics, clinical data, and disease prognosis of all patients were collected and analyzed. RESULTS: There were significant differences in SLEDAI, total glyceride (TG), serum amyloid A (SAA), SAR, urinary microalbumin-to-creatinine ratio (ACR), erythrocyte sedimentation rate (ESR), albumin (ALB), complement 3 (C3), anti-dsDNA, anti-Sm positive rate, and anti-dsDNA positive rate between active SLE and stable SLE patients. TG, SAR, C3, ACR, and positive anti-dsDNA were independent influencing factors of active SLE, and the odds ratio (OR) values were 2.342, 10.921, 0.832, 1.451, and 2.476, respectively. The area under curves (AUCs) of SAA, ALB, and SAR for predicting active SLE and severe active SLE were 0.743, 0.724, 0.787, 0.711, 0.686, and 0.733, respectively. The AUC of SAR for predicting the poor prognosis of active SLE was 0.719. High SAR, high ACR, low C3, and positive anti-dsDNA were high risk factors for poor prognosis. Kaplan-Meier (K-M) survival analysis showed that patients with high SAR, high ACR, low C3, and positive anti-dsDNA had shorter continuous remission time than that with low SAR, low ACR, high C3, and negative anti-dsDNA. CONCLUSION: SAR had high predictive value for active SLE, severe active SLE, and poor prognosis of SLE. High SAR may be a potential marker for predicting the activity and prognosis of Chinese patients with SLE.

New wine in old bottles: current progress on P5 ATPases.

P5 ATPases are evolutionarily conserved P-type transporters. Despite their important roles in the endoplasmic reticulum (ER) and in lysosomes, the substrate specificities and transporting mechanisms of P5 ATPases have remained mysterious. Recently, several studies have provided genetic, biochemical, and structural evidence to help elucidate the physiological functions and substrates of P5 ATPases. Here, we summarize this progress and discuss the potential transport mechanisms of the P5 ATPases-in particular, P5A ATPase-for further study.

P5A ATPase controls ER translocation of Wnt in neuronal migration.

The Wnt family contains conserved secretory proteins required for developmental patterning and tissue homeostasis. However, how Wnt is targeted to the endoplasmic reticulum (ER) for processing and secretion remains poorly understood. Here, we report that CATP-8/P5A ATPase directs neuronal migration non-cell autonomously in Caenorhabditis elegans by regulating EGL-20/Wnt biogenesis. CATP-8 likely functions as a translocase to translocate nascent EGL-20/Wnt polypeptide into the ER by interacting with the highly hydrophobic core region of EGL-20 signal sequence. Such regulation of Wnt biogenesis by P5A ATPase is common in C. elegans and conserved in human cells. These findings describe the physiological roles of P5A ATPase in neural development and identify Wnt proteins as direct substrates of P5A ATPase for ER translocation.

Assessment of multiple pathways involved in the inhibitory effect of HCG22 on oral squamous cell carcinoma progression.

LncRNAs have been proposed to be associated with the tumorigenesis and progression of oral squamous cell carcinoma (OSCC). LncRNA HLA complex group 22 (HCG22) was reported to be lowly expressed and associated with poor prognosis in head and neck squamous cell carcinoma (HNSCC). However, the biological role and related mechanism of HCG22 in OSCC have not been characterized. HCG22 expression in OSCC cells was detected by qRT-PCR. Cell proliferation, invasion, and apoptosis were evaluated by Bromodeoxyuridine (BrdU) proliferation assay, Transwell invasion assay, and flow cytometry analysis, respectively. The protein levels of proliferating cell nuclear antigen (PCNA), E-cadherin, Vimentin, Bcl-2, Bax, protein kinase B (Akt), phosphorylated Akt (p-Akt), mammalian target of rapamycin (mTOR), phosphorylated mTOR (p-mTOR), and ß-catenin were detected by western blot. Cell growth evaluation was performed using in vitro colony formation assay and in vivo tumor xenograft assay. We found that HCG22 was weakly expressed in OSCC cells. HCG22 overexpression inhibited cell proliferation and invasion and induced apoptosis in OSCC cells. The levels of PCNA, Vimentin, and Bcl-2 were decreased and E-cadherin and Bax expression was elevated in OSCC cells after HCG22 overexpression. Additionally, HCG22 overexpression inhibited the Akt, mTOR and Wnt/ß-catenin pathways. Activation of Akt, mTOR, and Wnt/ß-catenin pathways attenuated the anti-tumor property of HCG22 in OSCC cells. Furthermore, HCG22 overexpression inhibited the growth of OSCC cells in vitro and in vivo. In conclusion, HCG22 exerted anti-tumor property in OSCC by inhibiting the Akt, mTOR, and Wnt/ß-catenin pathways.

In vivo imaging of a PVD neuron in Caenorhabditis elegans.

The nematode Caenorhabditis elegans nociceptive PVD neurons have highly ordered dendritic branches, making this an ideal model to study the development and organization of dendrites. A ser-2-promoter-driven GFP reporter line wyIs592[ser-2prom-3p::myr-GFP] provides a comprehensive visualization of PVD anatomy. Here, we describe the detailed procedures for imaging a PVD neuron using wyIs592 at late L4 larval stage in vivo by confocal microscopy. This protocol can also be applied to imaging other cells in C. elegans. For complete details on the use and execution of this protocol, please refer to Feng et al. (2020).

The effect of rheumatoid factor on three commercial immunoassays for serum cystatin C.

Cystatin C (CysC) can be used to diagnose early changes in renal insufficiency. However, there are little researches to study whether there is an interference between the level of CysC and RF. Thus, we conducted this study to investigate it. We randomly selected 30 patients with high RF (RF concentration: 552.05 ± 476.23 IU/mL) and 33 healthy subjects with RF concentration <11.1 IU/mL and CysC were measured with different reagents and instruments; Interference experiment was also be included. The results showed that the measured CysC concentration increases with increasing RF concentration in a dose-dependent manner and CysC levels are falsely increased by RF interference depends on the reagents used (CysC reagent: Whitman, Nanjing, China). Reagent manufacturers should fully consider RF interference when developing CysC reagents, and evaluate them before they are sold. When selecting CysC reagents, we should evaluate RF interference to the measurement to avoid misleading results.

CATP-8/P5A ATPase Regulates ER Processing of the DMA-1 Receptor for Dendritic Branching.

Dendrite morphogenesis is essential for a neuron to establish its receptive field and is, thus, the anatomical basis for the proper functioning of the nervous system. The molecular mechanisms governing dendrite branching are not fully understood. Using the multi-dendritic PVD neuron in the nematode Caenorhabditis elegans, we identify CATP-8/P5A ATPase as a key regulator of dendrite branching that controls the translocation of the DMA-1 receptor to the endoplasmic reticulum (ER). The specific signal peptide of DMA-1 and the ATPase activity of CATP-8 are essential for this process. Our results reveal that P5A ATPase may regulate protein translocation in the ER.

The E3 Ubiquitin Ligase SYVN1 Ubiquitinates Atlastins to Remodel the Endoplasmic Reticulum Network.

Atlastin (ATL) is a class of dynamin-like GTPases shaping endoplasmic reticulum (ER) by mediating homotypic membrane fusion. Defect of ATLs leads to abnormal ER structure and hereditary spastic paraplegia (HSP), a neurodegenerative disease with progressive spasticity. How ATLs are regulated to maintain the ER dynamics is not clear. Here, we found that SYVN1, an E3 ubiquitin ligase on the ER membrane, regulates ER shape and COPII exporting by mediating ubiquitination on ATLs, especially ATL1. ATL1 is ubiquitinated by SYVN1 strongly on K285 and mildly on K287. Ubiquitination on ATL1 does not result in protein degradation but inhibits ATL1 GTPase activity. SYVN1 overexpression compensates the excessive ER network fusion caused by ATL1 overexpression. Accordingly, the role of SYVN1 and ATL1 in regulating ER morphology is also recapitulated in Caenorhabditis elegans. Taken together, our study reveals a different role of SYVN1 in ER remodeling through mediating ubiquitination on ATLs.