TMEM52B Isoforms P18 and P20 Differentially Promote the Oncogenesis and Metastasis of Nasopharyngeal Carcinoma.

Transmembrane protein 52B (TMEM52B), a newly identified tumor-related gene, has been reported to regulate various tumors, yet its role in nasopharyngeal carcinoma (NPC) remains unclear. Transcriptomic analysis of NPC cell lines reveals frequent overexpression of TMEM52B, and immunohistochemical results show that TMEM52B is associated with advanced tumor stage, recurrence, and decreased survival time. Depleting TMEM52B inhibits the proliferation, migration, invasion, and oncogenesis of NPC cells in vivo. TMEM52B encodes two isoforms, TMEM52B-P18 and TMEM52B-P20, differing in their N-terminals. While both isoforms exhibit similar pro-oncogenic roles and contribute to drug resistance in NPC, TMEM52B-P20 differentially promotes metastasis. This functional discrepancy may be attributed to their distinct subcellular localization; TMEM52B-P18 is confined to the cytoplasm, while TMEM52B-P20 is found both at the cell membrane and in the cytoplasm. Mechanistically, cytoplasmic TMEM52B enhances AKT phosphorylation by interacting with phosphoglycerate kinase 1 (PGK1), fostering NPC growth and metastasis. Meanwhile, membrane-localized TMEM52B-P20 promotes E-cadherin ubiquitination and degradation by facilitating its interaction with the E3 ubiquitin ligase NEDD4, further driving NPC metastasis. In conclusion, the TMEM52B-P18 and TMEM52B-P20 isoforms promote the metastasis of NPC cells through different mechanisms. Drugs targeting these TMEM52B isoforms may offer therapeutic benefits to cancer patients with varying degrees of metastasis.

DOM hydrophilic components of organic fertilizers increased the soil nitrogen retention capacity and succession of the microbial community.

Introduction: Application of organic fertilizers affects soil properties and microbial communities, which in turn alters soil N transformation processes. Unfortunately, it is not clear how the difference in the character of the organic fertilizer DOM affects the soil nitrogen retention capacity and its microbial processes. Methods: According to the principle of equal nutrients, the treatments of chemical fertilizer alone (treatment CF), chemical fertilizer with organic fertilizer DOM hydrophilic components (treatment H), and chemical fertilizer with organic fertilizer DOM hydrophobic components (treatment P) were set up, where the characteristics of soil nitrogen transformation and changes in microbial community structure were studied with soil culture conditions for 24 days. Results: It was discovered that the addition of organic fertilizer DOM components (H and P) slowed nitrification rate and increased protease activity resulting in a higher NH4+-N content compared to the CF treatment. The DOM addition (H and P) increased the microbial biomass nitrogen (MBN) levels in the soil and increased the soil nitrogen pool capacity. Conclusions: Moreover, the carbon use efficiency of the hydrophilic components is higher than that of the hydrophobic components, resulting in its further increase in nitrogen reservoir capacity and higher nitrogen retention capacity. Network analysis showed that the addition of organic fertilizer DOM hydrophilic components increased network complexity and synergy between microorganisms. In combination with random forest analysis, it was shown that Sphingomonas and Massilia were key species influencing soil nitrogen retention capacity and nitrogen availability characteristics.

Identification of hyperoxidized PRDX3 as a ferroptosis marker reveals ferroptotic damage in chronic liver diseases.

Ferroptosis, a regulated cell death pathway driven by accumulation of phospholipid peroxides, has been challenging to identify in physiological conditions owing to the lack of a specific marker. Here, we identify hyperoxidized peroxiredoxin 3 (PRDX3) as a marker for ferroptosis both in vitro and in vivo. During ferroptosis, mitochondrial lipid peroxides trigger PRDX3 hyperoxidation, a posttranslational modification that converts a Cys thiol to sulfinic or sulfonic acid. Once hyperoxidized, PRDX3 translocates from mitochondria to plasma membranes, where it inhibits cystine uptake, thereby causing ferroptosis. Applying hyperoxidized PRDX3 as a marker, we determined that ferroptosis is responsible for death of hepatocytes in mouse models of both alcoholic and nonalcoholic fatty liver diseases, the most prevalent chronic liver disorders. Our study highlights the importance of ferroptosis in pathophysiological conditions and opens the possibility to treat these liver diseases with drugs that inhibit ferroptosis.

Responses of soil microbial biomass nitrogen to organic fertilizer with different degrees of maturity and regu-lation to soil mineral nitrogen.

By combining the composting process with soil culture experiment, we conducted an experiment with four treatments, including conventional chemical fertilizer (CK), chemical fertilizer + compost maturity reaching 50% germination index (GI, the same below) organic fertilizer (CO1), chemical fertilizer + compost maturity reaching 80% GI organic fertilizer (CO2), chemical fertilizer + compost maturity reaching 100% GI organic ferti-lizer (CO3). We measured soil microbial biomass nitrogen (MBN), mineral nitrogen (NH4+-N, NO3--N), net nitrification rate, microbial biomass carbon (MBC), dissolved organic carbon (DOC), soil urease and soil protease, aiming to reveal the regulatory effect of soil MBN on mineral nitrogen. The results showed that organic fertilizer application significantly increased MBN and NH4+-N concentrations by 50.1%-62.4% and 109.9%-147.1%, reduced NO3--N concentration and net nitrification rate by 23.3%-46.8%, and 26.2%-51.5%, and enhanced MBC, DOC, urease and protease activities by 33.8%-69.6%, 7.4%-20.8%, 11.2%-69.0% and 9.4%-25.1%, respectively. The change ranges of CO2 and CO3 were significantly higher than CO1. Redundancy analysis (RDA) and structural equation model (SEM) results showed that the application of organic fertilizer with higher degree of maturity (GI≥80%) positively regulated soil MBC, MBN, NH4+-N, and the activities of urease and protease, but had a negative effect on soil net nitrification rate. The combined application of chemical fertilizers and high decomposed organic fertilizers could significantly increase soil MBN and NH4+-N contents, as well as soil urease and protease activities, but reduce soil net nitrification rate. To efficiently utilize organic solid wastes, it is recommended to use chemical fertilizer in combination of organic fertilizers with 80% decomposing degree in practical production to reduce the cost in both economy and time.

Humification process enhancement through relative abundance promotion of Talaromyces and Coprinopsis by inoculated Phanerochaete chrysosporium during the secondary fermentation of composting.

To explore the mechanism of Phanerochaete chrysosporium (P. chrysosporium) inoculation driving the humification process of maize straw composting, the treatments without P. chrysosporium inoculation (T1) and that with P. chrysosporium inoculation (T2) were carried out separately during the secondary fermentation of the co-composting of maize straw and rapeseed cake. The key microorganisms were determined by evaluating the succession of the fungal community and its relationship with humification process parameters. The results showed that P. chrysosporium inoculation (T2) reduced fungal diversity but increased the relative abundance of Coprinopsis and Talaromyces. At the end of the composting (day 36), the relative abundance of Talaromyces and Coprinopsis in T2 increased by 1223.7% and 30.2%, respectively, compared with T1. Combined CCA and SEMs analyses demonstrated the microbially driven mechanisms that enhance the humification process of composting, that is, P. chrysosporium inoculation promoted lignin continuous degradation by promoting the relative abundance of Talaromyces and Coprinopsis during the secondary fermentation of composting; meanwhile, P. Chrysosporium inoculation further intensified the biological process of humification in composting.

CircRNA-mediated regulation of brown adipose tissue adipogenesis.

Adipose tissue represents a candidate target for the treatment of metabolic illnesses, such as obesity. Brown adipose tissue (BAT), an important heat source within the body, promotes metabolic health through fat consumption. Therefore, the induction of white fat browning may improve lipid metabolism. Currently, the specific roles of circRNA in BAT and white adipose tissue (WAT) remain elusive. Herein, we conducted circRNA expression profiling of mouse BAT and WAT using RNA-seq. We identified a total of 12,183 circRNAs, including 165 upregulated and 79 downregulated circRNAs between BAT and WAT. Differentially expressed (DE) circRNAs were associated with the mitochondrion, mitochondrial part, mitochondrial inner membrane, mitochondrial envelope, therefore, these circRNAs may affect the thermogenesis and lipid metabolism of BAT. Moreover, DE circRNAs were enriched in browning- and thermogenesis-related pathways, including AMPK and HIF-1 signaling. In addition, a novel circRNA, circOgdh, was found to be highly expressed in BAT, formed by back-splicing of the third and fourth exons of the Ogdh gene, and exhibited higher stability than linear Ogdh. circOgdh was mainly distributed in the cytoplasm and could sponge miR-34a-5p, upregulating the expression of Atgl, a key lipolysis gene, which enhanced brown adipocyte lipolysis and suppressed lipid droplet accumulation. Our findings offer in-depth knowledge of the modulatory functions of circRNAs in BAT adipogenesis.

FAF1 blocks ferroptosis by inhibiting peroxidation of polyunsaturated fatty acids.

Iron-dependent peroxidation of polyunsaturated fatty acids (PUFAs) leads to ferroptosis. While detoxification reactions removing lipid peroxides in phospholipids such as that catalyzed by glutathione peroxidase 4 (GPX4) protect cells from ferroptosis, the mechanism through which cells prevent PUFA peroxidation was not completely understood. We previously identified Fas-associated factor 1 (FAF1) as a protein directly interacting with free PUFAs through its UAS domain. Here we report that this interaction is crucial to protect cells from ferroptosis. In the absence of FAF1, cultured cells became sensitive to ferroptosis upon exposure to physiological levels of PUFAs, and mice developed hepatic injury upon consuming a diet enriched in PUFA. Mechanistically, we demonstrate that FAF1 assembles a globular structure that sequesters free PUFAs into a hydrophobic core, a reaction that prevents PUFA peroxidation by limiting its access to iron. Our study suggests that peroxidation of free PUFAs contributes to ferroptosis, and FAF1 acts upstream of GPX4 to prevents initiation of ferroptosis by limiting peroxidation of free PUFAs.

Identification of TRAMs as sphingolipid-binding proteins using a photoactivatable and clickable short-chain ceramide analog.

Ceramide is a lipid molecule that regulates diverse physiological and pathological reactions in part through inverting the topology of certain transmembrane proteins. This topological inversion is achieved through regulated alternative translocation (RAT), which reverses the direction by which membrane proteins are translocated across the endoplasmic reticulum during translation. However, owing to technical challenges in studying protein-ceramide interaction, it remains unclear how ceramide levels are sensed in cells to trigger RAT. Here, we report the synthesis of pac-C7-Cer, a photoactivatable and clickable short-chain ceramide analog that can be used as a probe to study protein-ceramide interactions. We demonstrate that translocating chain-associated membrane protein 2 (TRAM2), a protein known to control RAT of transmembrane 4 L6 subfamily member 20, and TRAM1, a homolog of TRAM2, interacted with molecules derived from pac-C7-Cer. This interaction was competed by naturally existing long-chain ceramide molecules. We showed that binding of ceramide and its analogs to TRAM2 correlated with their ability to induce RAT of transmembrane 4 L6 subfamily member 20. In addition to probing ceramide-TRAM interactions, we provide evidence that pac-C7-cer could be used for proteome-wide identification of ceramide-binding proteins. Our study provides mechanistic insights into RAT by identifying TRAMs as potential ceramide-binding proteins and establishes pac-C7-Cer as a valuable tool for future study of ceramide-protein interactions.

Rab18 binds PLIN2 and ACSL3 to mediate lipid droplet dynamics.

Lipid droplet (LD) is a vital organelle governing lipid homeostasis and Rab18 has been linked to lipid metabolism. However, the mechanisms of Rab18-mediated LD dynamics in myoblast cells remain elusive. Here, we report that Rab18 plays an important role in oleic acid (OA)-induced LD accumulation in mouse myoblast C2C12 cells. Rab18 was translocated from the endoplasmic reticulum (ER) to LDs during LD accumulation, which was regulated by perilipin 2 (PLIN2), a major LD protein. LD-associated Rab18 bound with the C terminus of PLIN2 and the LD localization of Rab18 was diminished when PLIN2 was depleted. Moreover, loss of function of Rab18 led to reduced triacylglycerol (TAG) level and fewer but larger LDs. In contrast, overexpression of Rab18 resulted in elevated TAG content and LD number. Furthermore, LD-associated Rab18 interacted with acyl-CoA synthetase long-chain family member 3 (ACSL3), which in turn promoted the LD localization of this protein. These data show that Rab18 interacts with PLIN2 and forms a complex with PLIN2 and ACSL3, which plays a critical role in LD accumulation and dynamics of myoblast cells.

Identification and Pleiotropic Effect Analysis of GSE5 on Rice Chalkiness and Grain Shape.

Chalkiness is one of several major restricting factors for the improvement of rice quality. Although many chalkiness-related quantitative trait loci have been mapped, only a small number of genes have been cloned to date. In this study, the candidate gene GSE5 of a major quantitative trait locus (QTL) for rice chalkiness, qDEC5, was identified by map-based cloning. Phenotyping and haplotype analysis of proActin:GSE5 transgenic line, gse5-cr mutant, and 69 rice varieties further confirmed that GSE5 had the pleiotropic effects and regulated both chalkiness and grain shape. Genetic analysis showed GSE5 was a dominant gene for grain length and a semi-dominant gene for grain width and chalkiness. The DNA interval closely linked to GSE5 was introgressed to Zhenshan 97B (ZB) based on molecular marker-assisted selection, and the improved ZB showed lower chalkiness and longer but smaller grains, which showed that GSE5 played an important role in breeding rice varieties with high yield and good quality. Transcriptomics, proteomics, and qRT-PCR analyses showed that thirty-nine genes associated with carbon and protein metabolism are regulated by GSE5 to affect the formation of chalkiness, including some newly discovered genes, such as OsCESA9, OsHSP70, OsTPS8, OsPFK04, OsSTA1, OsERdj3A, etc. The low-chalkiness lines showed higher amino sugar and nucleotide sugar metabolism at 10 days after pollination (DAP), lower carbohydrate metabolism at 15 DAP, and lower protein metabolism at 10 and 15 DAP. With heat shock at 34/30°C, rice chalkiness increased significantly; OsDjC10 and OsSUS3 were upregulated at 6 and 12 DAP, respectively, and OsGSTL2 was downregulated at 12 DAP. Our results identified the function and pleiotropic effects of qDEC5 dissected its genetic characteristics and the expression profiles of the genes affecting the chalkiness formation, and provided a theoretical basis and application value to harmoniously pursue high yield and good quality in rice production.

Identification of small ORF-encoded peptides in mouse serum.

Identification of the coding elements in the genome is fundamental to interpret the development of living systems and species diversity. Small peptides (length < 100 amino acids) have played an important role in regulating the biological metabolism, but their identification has been limited by their size and abundance. Serum is the most important body fluid and is full of small peptides. In this study, we have established a small ORF-encoded peptides (SEPs) database from mouse GENCODE release. This database provides about half a million putative translated SEPs in mouse. We also extract serum proteins from wild type and ob/ob mice, and collect the low molecular weight proteins for mass spectrometric analysis. More than 50 novel SEPs have been discovered. Several SEPs are further verified by biochemical method with newly raised antibodies. These novel SEPs enhance the knowledge about the complexity of serum and provide new clues for the annotation and functional analysis of genes, especially the noncoding elements in the genome.

Morphologically and Functionally Distinct Lipid Droplet Subpopulations.

Lipid droplet (LD), a multi-functional organelle, is often found to associate with other cellular membranous structures and vary in size in a given cell, which may be related to their functional diversity. Here we established a method to separate LD subpopulations from isolated CHO K2 LDs into three different size categories. The subpopulation with smallest LDs was nearly free of ER and other membranous structures while those with larger LDs contained intact ER. These distinct subpopulations of LDs differed in their protein composition and ability to recruit proteins. This method was also applicable to LDs obtained from other sources, such as Huh7 cells, mouse liver and brown adipose tissue, et al. We developed an in vitro assay requiring only isolated LDs, Coenzyme A, and ATP to drive lipid synthesis. The LD subpopulation nearly depleted of ER was able to incorporate fatty acids into triacylglycerol and phospholipids. Together, our data demonstrate that LDs in a given cell are heterogeneous in size and function, and suggest that LDs are one of cellular lipid synthetic organelles.

Comparative Proteomic Study of Fatty Acid-treated Myoblasts Reveals Role of Cox-2 in Palmitate-induced Insulin Resistance.

Accumulated studies demonstrate that saturated fatty acids (FAs) such as palmitic acid (PA) inhibit insulin signaling in skeletal muscle cells and monounsaturated fatty acids such as oleic acid (OA) reverse the effect of PA on insulin signaling. The detailed molecular mechanism of these opposite effects remains elusive. Here we provide a comparative proteomic study of skeletal myoblast cell line C2C12 that were untreated or treated with PA, and PA plus OA. A total of 3437 proteins were quantified using SILAC in this study and 29 proteins fall into the pattern that OA reverses PA effect. Expression of some these proteins were verified using qRT-PCR and Western blot. The most significant change was cyclooxygenase-2 (Cox-2). In addition to whole cell comparative proteomic study, we also compared lipid droplet (LD)-associated proteins and identified that Cox-2 was one of three major altered proteins under the FA treatment. This finding was then confirmed using immunofluorescence. Finally, Cox-2 selective inhibitor, celecoxib protected cells from PA-reduced insulin signaling Akt phosphorylation. Together, these results not only provide a dataset of protein expression change in FA treatment but also suggest that Cox-2 and lipid droplets (LDs) are potential players in PA- and OA-mediated cellular processes.

A reagentless electrochemiluminescent immunosensor for apurinic/apyrimidinic endonuclease 1 detection based on the new Ru(bpy)3(2+)/bi-arginine system.

Apurinic/apyrimidinic endonuclease 1 (APE-1), a kind of multifunctional protein widely-distributed in the body, plays an essential role in the DNA base excision repair and serves as multiple possible roles in the response of human cancer to radiotherapy and chemotherapy. In this work, an ultrasensitive solid-state electrochemiluminescence (ECL) immunosensor is designed to determine APE-1 based on the new Ru(bpy)3(2+)/bi-arginine system. The bi-arginine (bi-Arg) is decorated on the Au nanoparticles functionalized magnetic Fe3O4/reduced graphene oxide (bi-Arg/Au@Fe3O4-rGO) according to the self-assembling and covalent cross-linking interaction to obtain the functionalized nanocomposite of bi-Arg/Au@Fe3O4-rGO. Herein, the bi-Arg/Au@Fe3O4-rGO plays not only an amplification label to enhance the ECL signal of Ru(bpy)3(2+) due to the coreactant of bi-Arg but also an ideal nanocarrier to load numerous secondary antibody. Based on sandwich-type immunoassay format, this proposed method offers a linear range of 1.0fgmL(-1)-5.0pgmL(-1) and an estimated detection limit of 0.3fgmL(-1) for the APE-1. Moreover, the reagentless ECL immunosensor also exhibits high sensitivity, excellent selectivity and good stability, which has greatly potential development and application in clinical diagnostics, immunology and biomedical research.