Chemical characterization and source apportionment of PM2.5 in a Northeastern China city during the epidemic period.

To investigate the influence of COVID-19 lockdown measures on PM2.5 and its chemical components in Shenyang, PM2.5 samples were continuously collected from January 1 to May 31, 2020. The samples were then analyzed for water-soluble inorganic ions, metal elements, organic carbon, and elemental carbon. The findings indicated a significant decrease in PM2.5 and its various chemical components during the lockdown period, compared to pre-lockdown levels (p < 0.05), suggesting a substantial improvement in air quality. Water-soluble inorganic ions (WSIIs) were identified as the primary contributors to PM2.5, accounting for 47% before the lockdown, 46% during the lockdown, and 37% after the lockdown. Ionic balance analysis revealed that PM2.5 exhibited neutral, weakly alkaline, and alkaline characteristics before, during, and after the lockdown, respectively. NH4+ was identified as the main balancing cation and was predominantly present in the form of NH4NO3 in the absence of complete neutralization of SO42- and NO3-. Moreover, the higher sulfur oxidation ratio (SOR) and nitrogen oxidation ratio (NOR), along with the significant increase in PM2.5/EC, suggested intense secondary transformation during the lockdown period. The elevated OC/EC ratio during the lockdown period implied higher secondary organic carbon (SOC), and the notable increase in SOC/EC ratio indicated a significant secondary transformation of total carbon. The enrichment factor (EF) results revealed that during the lockdown, 9 metal elements (As, Sn, Pb, Zn, Cu, Sb, Ag, Cd, and Se) were substantially impacted by anthropogenic emissions. Source analysis of PMF was employed to identify the sources of PM2.5 in Shenyang during the study period, and the analysis identified six factors: secondary sulfate and vehicle emissions, catering fume sources, secondary nitrate and coal combustion emissions, dust sources, biomass combustion, and industrial emissions, with secondary sulfate and vehicle emissions and catering fume sources contributing the most to PM2.5.

Subscaling of a cytosolic RNA binding protein governs cell size homeostasis in the multiple fission alga Chlamydomonas.

Coordination of growth and division in eukaryotic cells is essential for populations of proliferating cells to maintain size homeostasis, but the underlying mechanisms that govern cell size have only been investigated in a few taxa. The green alga Chlamydomonas reinhardtii (Chlamydomonas) proliferates using a multiple fission cell cycle that involves a long G1 phase followed by a rapid series of successive S and M phases (S/M) that produces 2n daughter cells. Two control points show cell-size dependence: the Commitment control point in mid-G1 phase requires the attainment of a minimum size to enable at least one mitotic division during S/M, and the S/M control point where mother cell size governs cell division number (n), ensuring that daughter distributions are uniform. tny1 mutants pass Commitment at a smaller size than wild type and undergo extra divisions during S/M phase to produce small daughters, indicating that TNY1 functions to inhibit size-dependent cell cycle progression. TNY1 encodes a cytosolic hnRNP A-related RNA binding protein and is produced once per cell cycle during S/M phase where it is apportioned to daughter cells, and then remains at constant absolute abundance as cells grow, a property known as subscaling. Altering the dosage of TNY1 in heterozygous diploids or through mis-expression increased Commitment cell size and daughter cell size, indicating that TNY1 is a limiting factor for both size control points. Epistasis placed TNY1 function upstream of the retinoblastoma tumor suppressor complex (RBC) and one of its regulators, Cyclin-Dependent Kinase G1 (CDKG1). Moreover, CDKG1 protein and mRNA were found to over-accumulate in tny1 cells suggesting that CDKG1 may be a direct target of repression by TNY1. Our data expand the potential roles of subscaling proteins outside the nucleus and imply a control mechanism that ties TNY1 accumulation to pre-division mother cell size.

Autophagosome biogenesis and organelle homeostasis in plant cells.

Autophagy is one of the major highly inducible degradation processes in response to plant developmental and environmental signals. In response to different stimuli, cellular materials, including proteins and organelles, can be sequestered into a double membrane autophagosome structure either selectively or non-selectively. The formation of an autophagosome as well as its delivery into the vacuole involves complex and dynamic membrane processes. The identification and characterization of the conserved autophagy-related (ATG) proteins and their related regulators have greatly advanced our understanding of the molecular mechanism underlying autophagosome biogenesis and function in plant cells. Autophagosome biogenesis is tightly regulated by the coordination of multiple ATG and non-ATG proteins, and selective cargo recruitment. This review updates our current knowledge of autophagosome biogenesis, with special emphasis on the core molecular machinery that drives autophagosome formation, and autophagosome-organelle interactions under abiotic stress conditions.

A Century Journey of Organelles Research in the Plant Endomembrane System.

We are entering an exciting century in the study of the plant organelles in the endomembrane system. Over the past century, especially within the past 50 years, tremendous advancements have been made in the complex plant cell to generate a much clearer and informative picture of plant organelles, including the molecular/morphological features, dynamic/spatial behavior, and physiological functions. Importantly, all these discoveries and achievements in the identification and characterization of organelles in the endomembrane system would not have been possible without: 1) the innovations and timely applications of various state-of-art cell biology tools and technologies for organelle biology research, 2) the continuous efforts in developing and characterizing new organelle markers by the plant biology community; and 3) the landmark studies on the identification and characterization of the elusive organelles. While molecular aspects and results for individual organelles have been extensively reviewed, the development of the techniques for organelle research in plant cell biology is less appreciated. As one of the ASPB Centennial Reviews on "organelle biology", here we aim to take a journey across a century of organelle biology research in plants by highlighting the important tools (or landmark technologies) and key scientists that contributed to visualize organelles. We then highlight the landmark studies leading to the identification and characterization of individual organelles in the plant endomembrane systems.

Indole Diterpene Derivatives from the Aspergillus flavus GZWMJZ-288, an Endophytic Fungus from Garcinia multiflora.

A new indole diterpene, 26-dihydroxyaflavininyl acetate (1), along with five known analogs (2-6) were isolated from the liquid fermentation of Aspergillus flavus GZWMJZ-288, an endophyte from Garcinia multiflora. The structures of these compounds were identified through NMR, MS, chemical reaction, and X-ray diffraction experiments. Enzyme inhibition activity screening found that compounds 1, 4, and 6 have a good binding affinity with NPC1L1, among which compound 6 exhibited a stronger binding ability than ezetimibe at a concentration of 10 µM. Moreover, compound 5 showed inhibitory activity against α-glucosidase with an IC50 value of 29.22 ± 0.83 µM, which is 13 times stronger than that of acarbose. The results suggest that these aflavinine analogs may serve as lead compounds for the development of drugs targeting NPC1L1 and α-glucosidase. The binding modes of the bioactive compounds with NPC1L1 and α-glucosidase were also performed through in silico docking studies.

Recent advances in plant endomembrane research and new microscopical techniques.

The endomembrane system consists of various membrane-bound organelles including the endoplasmic reticulum (ER), Golgi apparatus, trans-Golgi network (TGN), endosomes, and the lysosome/vacuole. Membrane trafficking between distinct compartments is mainly achieved by vesicular transport. As the endomembrane compartments and the machineries regulating the membrane trafficking are largely conserved across all eukaryotes, our current knowledge on organelle biogenesis and endomembrane trafficking in plants has mainly been shaped by corresponding studies in mammals and yeast. However, unique perspectives have emerged from plant cell biology research through the characterization of plant-specific regulators as well as the development and application of the state-of-the-art microscopical techniques. In this review, we summarize our current knowledge on the plant endomembrane system, with a focus on several distinct pathways: ER-to-Golgi transport, protein sorting at the TGN, endosomal sorting on multivesicular bodies, vacuolar trafficking/vacuole biogenesis, and the autophagy pathway. We also give an update on advanced imaging techniques for the plant cell biology research.

A positive feedback regulation of SnRK1 signaling by autophagy in plants.

SnRK1, an evolutionarily conserved heterotrimeric kinase complex that acts as a key metabolic sensor in maintaining energy homeostasis in plants, is an important upstream activator of autophagy that serves as a cellular degradation mechanism for the healthy growth of plants. However, whether and how the autophagy pathway is involved in regulating SnRK1 activity remains unknown. In this study, we identified a clade of plant-specific and mitochondria-localized FCS-like zinc finger (FLZ) proteins as currently unknown ATG8-interacting partners that actively inhibit SnRK1 signaling by repressing the T-loop phosphorylation of the catalytic α subunits of SnRK1, thereby negatively modulating autophagy and plant tolerance to energy deprivation caused by long-term carbon starvation. Interestingly, these AtFLZs are transcriptionally repressed by low-energy stress, and AtFLZ proteins undergo a selective autophagy-dependent pathway to be delivered to the vacuole for degradation, thereby constituting a positive feedback regulation to relieve their repression of SnRK1 signaling. Bioinformatic analyses show that the ATG8-FLZ-SnRK1 regulatory axis first appears in gymnosperms and seems to be highly conserved during the evolution of seed plants. Consistent with this, depletion of ATG8-interacting ZmFLZ14 confers enhanced tolerance, whereas overexpression of ZmFLZ14 leads to reduced tolerance to energy deprivation in maize. Collectively, our study reveals a previously unknown mechanism by which autophagy contributes to the positive feedback regulation of SnRK1 signaling, thereby enabling plants to better adapt to stressful environments.

Anlotinib Combined With Toripalimab as First-Line Therapy for Unresectable Hepatocellular Carcinoma: A Prospective, Multicenter, Phase II Study.

BACKGROUND: For patients with unresectable hepatocellular carcinoma (HCC), the first-line therapeutic options are still relatively limited, and treatment outcomes remain poor. We aimed to assess the efficacy and safety of anlotinib combined with toripalimab as first-line therapy for unresectable HCC. METHODS: In this single-arm, multicenter, phase II study (ALTER-H-003), patients with advanced HCC without previous systemic anticancer therapy were recruited. Eligible patients were given anlotinib (12 mg on days 1-14) combined with toripalimab (240 mg on day 1) in a 3-week cycle. The primary endpoint was the objective response rate (ORR) by immune-related Response Evaluation Criteria in Solid Tumours (irRECIST)/RECIST v1.1 and modified RECIST (mRECIST). Secondary endpoints included disease control rate (DCR), duration of response (DoR), progression-free survival (PFS), overall survival (OS), and safety. RESULTS: Between January 2020 and Jul 2021, 31 eligible patients were treated and included in the full analysis set. At data cutoff (January 10, 2023), the ORR was 29.0% (95% CI: 12.1%-46.0%) by irRECIST/RECIST v1.1, and 32.3% (95% CI: 14.8%-49.7%) by mRECIST criteria, respectively. Confirmed DCR and median DoR by irRECIST/RECIST v1.1 and mRECIST criteria were 77.4 % (95% CI: 61.8%-93.0%) and not reached (range: 3.0-22.5+ months), respectively. Median PFS was 11.0 months (95% CI: 3.4-18.5 months) and median OS was 18.2 months (95% CI: 15.8-20.5 months). Of the 31 patients assessed for adverse events (AEs), the most common grade ≥ 3 treatment-related AEs were hand-foot syndrome (9.7%, 3/31), hypertension (9.7%, 3/31), arthralgia (9.7%, 3/31), abnormal liver function (6.5%, 2/31), and decreased neutrophil counts (6.5%, 2/31). CONCLUSIONS: Anlotinib combined with toripalimab showed promising efficacy and manageable safety in Chinese patients with unresectable HCC in the first-line setting. This combination therapy may offer a potential new therapeutic approach for patients with unresectable HCC.

ABI5-FLZ13 module transcriptionally represses growth-related genes to delay seed germination in response to ABA.

The bZIP transcription factor ABSCISIC ACID INSENSITIVE5 (ABI5) is a master regulator of seed germination and post-germinative growth in response to abscisic acid (ABA), but the detailed molecular mechanism by which it represses plant growth remains unclear. In this study, we used proximity labeling to map the neighboring proteome of ABI5 and identified FCS-LIKE ZINC FINGER PROTEIN 13 (FLZ13) as a novel ABI5 interaction partner. Phenotypic analysis of flz13 mutants and FLZ13-overexpressing lines demonstrated that FLZ13 acts as a positive regulator of ABA signaling. Transcriptomic analysis revealed that both FLZ13 and ABI5 downregulate the expression of ABA-repressed and growth-related genes involved in chlorophyll biosynthesis, photosynthesis, and cell wall organization, thereby repressing seed germination and seedling establishment in response to ABA. Further genetic analysis showed that FLZ13 and ABI5 function together to regulate seed germination. Collectively, our findings reveal a previously uncharacterized transcriptional regulatory mechanism by which ABA mediates inhibition of seed germination and seedling establishment.

Arabidopsis AUTOPHAGY-RELATED2 is essential for ATG18a and ATG9 trafficking during autophagosome closure.

As a fundamental metabolic pathway, autophagy plays important roles in plant growth and development, particularly under stress conditions. A set of autophagy-related (ATG) proteins is recruited for the formation of a double-membrane autophagosome. Among them, the essential roles of ATG2, ATG18, and ATG9 have been well established in plant autophagy via genetic analysis; however, the underlying molecular mechanism for ATG2 in plant autophagosome formation remains poorly understood. In this study, we focused on the specific role of ATG2 in the trafficking of ATG18a and ATG9 during autophagy in Arabidopsis (Arabidopsis thaliana). Under normal conditions, YFP-ATG18a proteins are partially localized on late endosomes and translocated to ATG8e-labeled autophagosomes upon autophagic induction. Real-time imaging analysis revealed sequential recruitment of ATG18a on the phagophore membrane, showing that ATG18a specifically decorated the closing edges and finally disassociated from the completed autophagosome. However, in the absence of ATG2, most of the YFP-ATG18a proteins are arrested on autophagosomal membranes. Ultrastructural and 3D tomography analysis showed that unclosed autophagosome structures are accumulated in the atg2 mutant, displaying direct connections with the endoplasmic reticulum membrane and vesicular structures. Dynamic analysis of ATG9 vesicles suggested that ATG2 depletion also affects the association between ATG9 vesicles and the autophagosomal membrane. Furthermore, using interaction and recruitment analysis, we mapped the interaction relationship between ATG2 and ATG18a, implying a possible role of ATG18a in recruiting ATG2 and ATG9 to the membrane. Our findings unveil a specific role of ATG2 in coordinating ATG18a and ATG9 trafficking to mediate autophagosome closure in Arabidopsis.

A non-canonical role of ATG8 in Golgi recovery from heat stress in plants.

Above-optimal growth temperatures, usually referred to as heat stress (HS), pose a challenge to organisms' survival as they interfere with essential physiological functions and disrupt cellular organization. Previous studies have elucidated the complex transcriptional regulatory networks involved in plant HS responses, but the mechanisms of organellar remodelling and homeostasis during plant HS adaptations remain elusive. Here we report a non-canonical function of ATG8 in regulating the restoration of plant Golgi damaged by HS. Short-term acute HS causes vacuolation of the Golgi apparatus and translocation of ATG8 to the dilated Golgi membrane. The inactivation of the ATG conjugation system, but not of the upstream autophagic initiators, abolishes the targeting of ATG8 to the swollen Golgi, causing a delay in Golgi recovery after HS. Using TurboID-based proximity labelling, we identified CLATHRIN LIGHT CHAIN 2 (CLC2) as an interacting partner of ATG8 via the AIM-LDS interface. CLC2 is recruited to the cisternal membrane by ATG8 to facilitate Golgi reassembly. Collectively, our study reveals a hitherto unanticipated process of Golgi stack recovery from HS in plant cells and uncovers a previously unknown mechanism of organelle resilience involving ATG8.

VAMP724 and VAMP726 are involved in autophagosome formation in Arabidopsis thaliana.

Macroautophagy/autophagy, an evolutionarily conserved degradative process essential for cell homeostasis and development in eukaryotes, involves autophagosome formation and fusion with a lysosome/vacuole. The soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) proteins play important roles in regulating autophagy in mammals and yeast, but relatively little is known about SNARE function in plant autophagy. Here we identified and characterized two Arabidopsis SNAREs, AT4G15780/VAMP724 and AT1G04760/VAMP726, involved in plant autophagy. Phenotypic analysis showed that mutants of VAMP724 and VAMP726 are sensitive to nutrient-starved conditions. Live-cell imaging on mutants of VAMP724 and VAMP726 expressing YFP-ATG8e showed the formation of abnormal autophagic structures outside the vacuoles and compromised autophagic flux. Further immunogold transmission electron microscopy and electron tomography (ET) analysis demonstrated a direct connection between the tubular autophagic structures and the endoplasmic reticulum (ER) in vamp724-1 vamp726-1 double mutants. Further transient co-expression, co-immunoprecipitation and double immunogold TEM analysis showed that ATG9 (autophagy related 9) interacts and colocalizes with VAMP724 and VAMP726 in ATG9-positive vesicles during autophagosome formation. Taken together, VAMP724 and VAMP726 regulate autophagosome formation likely working together with ATG9 in Arabidopsis.Abbreviations: ATG, autophagy related; BTH, benzo-(1,2,3)-thiadiazole-7-carbothioic acid S-methyl ester; Conc A, concanamycin A; EM, electron microscopy; ER, endoplasmic reticulum; FRET, Förster/fluorescence resonance energy transfer; MS, Murashige and Skoog; MVB, multivesicular body; PAS, phagophore assembly site; PM, plasma membrane; PVC, prevacuolar compartment; SNARE, soluble N-ethylmaleimide-sensitive factor attachment protein receptor; TEM, transmission electron microscopy; TGN, trans-Golgi network; WT, wild-type.

A plant-unique protein BLISTER coordinates with core retromer to modulate endosomal sorting of plasma membrane and vacuolar proteins.

The retromer is a heteromeric protein complex that localizes to endosomal membranes and drives the formation of endosomal tubules that recycle membrane protein cargoes. In plants, the retromer plays essential and canonical functions in regulating the transport of vacuolar storage proteins and the recycle of endocytosed plasma membrane proteins (PM); however, the mechanisms underlying the regulation of assembly, protein stability, and membrane recruitment of the plant retromer complex remain to be elucidated. In this study, we identify a plant-unique endosomal regulator termed BLISTER (BLI), which colocalizes and associates with the retromer complex by interacting with the retromer core subunits VPS35 and VPS29. Depletion of BLI perturbs the assembly and membrane recruitment of the retromer core VPS26-VPS35-VPS29 trimer. Consequently, depletion of BLI disrupts retromer-regulated endosomal trafficking function, including transport of soluble vacuolar proteins and recycling of endocytosed PIN-FORMED (PIN) proteins from the endosomes back to the PM. Moreover, genetic analysis in Arabidopsis thaliana mutants reveals BLI and core retromer interact genetically in the regulation of endosomal trafficking. Taken together, we identified BLI as a plant-specific endosomal regulator, which functions in retromer pathway to modulate the recycling of endocytosed PM proteins and the trafficking of soluble vacuolar cargoes.

Cu-MOF@Pt 3D nanocomposites prepared by one-step wrapping method with peroxidase-like activity for colorimetric detection of glucose.

As an alternative to natural enzymes, artificial enzymes based on nanomaterials have attracted a lot of attention owing to their outstanding catalytic activity and high stability as well as low cost. Cu-MOF loaded with platinum nanoparticles (labeled Cu-MOF@Pt) was prepared by simple one-step wrapping method using platinum nanoparticles, copper nitrate trihydrate and 1,3,5-tricarboxybenzene. It was confirmed that Cu-MOF@Pt exhibits peroxidase-like activity, which can quickly catalyze the oxidation of 3,3',5,5'-tetramethylbenzidine (TMB) and produce blue oxidized TMB (oxTMB) in the presence of hydrogen peroxide (H2O2). Additionally, steady-state kinetics showed that Cu-MOF@Pt exhibits stronger appetency to TMB and H2O2 compared with horseradish peroxidase. Thanks to the peroxidase-like activity of Cu-MOF@Pt, a highly selective colorimetric method for glucose detection has been successfully established, the linear range is 2-15 mM (R2 =0.9999) and the Limit of Detection (LOD) is 0.42 mM, with a detection range that meets clinical needs. Moreover, its good intra- and inter-assay precision and excellent stability make the results of glucose detection very reproducible. The detection performance of 90.09% was still maintained at 4 â„ƒ for 2 months. In conclusion, a new nanocomposite was successfully prepared and its selective detection ability for glucose was proved, which established a good basis for the clinical development of new enzymes for biosensors.

Shedding Light on the Role of Phosphorylation in Plant Autophagy.

Autophagy is a highly conserved quality control process that maintains cellular health by eliminating deleterious cargoes. Compared with the extensive studies in yeast and mammalian models, the molecular details and significance of post-translational modifications (PTMs) in the autophagy process in plants remain less well defined. In this review, we discuss recent progress in our understanding of phosphorylation, one of the most extensively studied PTMs, in the regulation of autophagosome biogenesis and autophagic degradation in plants. Based on the plant mass spectrometric database, we summarize the experimentally verified phosphorylation sites of the core autophagy machinery in plants. Furthermore, we put forward several approaches to test the roles of phosphorylation in the regulation of plant autophagy.

MLKs kinases phosphorylate the ESCRT component FREE1 to suppress abscisic acid sensitivity of seedling establishment.

The FYVE domain protein required for endosomal sorting 1 (FREE1), which was previously identified as a plant-specific component of the endosomal sorting complex required for transport machinery, plays an essential role in endosomal trafficking. Moreover, FREE1 also functions as an important negative regulator in abscisic acid (ABA) signalling. Multiple phosphorylations and ubiquitination sites have been identified in FREE1, hence unveiling the factors involved in posttranslational regulation of FREE1 is critical for comprehensively understanding FREE1-related regulatory networks during plant growth. Here, we demonstrate that plant-specific casein kinase I members MUT9-like kinases 1-4 (MLKs 1-4)/Arabidopsis EL1-like 1-4 interact with and phosphorylate FREE1 at serine residue S582, thereby modulating the nuclear accumulation of FREE1. Consequently, mutation of S582 to non-phosphorylable residue results in reduced nuclear localization of FREE1 and enhanced ABA response. In addition, mlk123 and mlk134 triple mutants accumulate less FREE1 in the nucleus and display hypersensitive responses to ABA treatment, whereas overexpression of the nuclear-localized FREE1 can restore the ABA sensitivity of seedling establishment in mlks triple mutants. Collectively, our study demonstrates a previously unidentified function of MLKs in attenuating ABA signalling in the nucleus by regulating the phosphorylation and nuclear accumulation of FREE1.

An Update on Coat Protein Complexes for Vesicle Formation in Plant Post-Golgi Trafficking.

Endomembrane trafficking is an evolutionarily conserved process for all eukaryotic organisms. It is a fundamental and essential process for the transportation of proteins, lipids, or cellular metabolites. The aforementioned cellular components are sorted across multiple membrane-bounded organelles. In plant cells, the endomembrane mainly consists of the nuclear envelope, endoplasmic reticulum (ER), Golgi apparatus, trans-Golgi network or early endosome (TGN/EE), prevacuolar compartments or multivesicular bodies (PVCs/MVBs), and vacuole. Among them, Golgi apparatus and TGN represent two central sorting intermediates for cargo secretion and recycling from other compartments by anterograde or retrograde trafficking. Several protein sorting machineries have been identified to function in these pathways for cargo recognition and vesicle assembly. Exciting progress has been made in recent years to provide novel insights into the sorting complexes and also the underlying sorting mechanisms in plants. Here, we will highlight the recent findings for the adaptor protein (AP) complexes, retromer, and retriever complexes, and also their functions in the related coated vesicle formation in post-Golgi trafficking.

Mammalian Cells Exocytose Alkylated Gold Nanoparticles via Extracellular Vesicles.

Understanding the exocytosis of nanoparticles (NPs) from cells is valuable because it informs design rules of NPs that support desirable cellular retention for nanomedicine applications, but investigations into the mechanism for the exocytosis of NPs remain scarce. We elucidate the mechanism for the exocytosis of dodecyl-terminated, polyethylene glycol-coated gold NPs (termed "dodecyl-PEG-AuNP"). The Au core enables ultrastructural differentiation of the exocytosed NPs from the nearby extracellular vesicles (EVs). The PEG shell prevents interparticle agglomeration or aggregation that disfavors exocytosis. The minute amounts of alkyl chains on the PEG shell not only promote cellular uptake but also improve exocytosis by up to 4-fold higher probability and upregulate exocytosis- and vesicle-related genes. After entering Kera-308 keratinocytes and trafficking to multivesicular bodies and lysosomes, these NPs exit the cell predominantly via unconventional exocytosis, accompanied by enhanced secretion of sub-100 nm, CD81-enriched exosomes. The pathway for NP exocytosis and subpopulation of EVs that are secreted alongside the exocytosed NPs depends on dodecyl loading. This work provides insights into dissecting the mechanism of NP exocytosis and its relationship with EV secretion.

Mechanistic insights into an atypical interaction between ATG8 and SH3P2 in Arabidopsis thaliana.

In selective macroautophagy/autophagy, cargo receptors are recruited to the forming autophagosome by interacting with Atg8 (autophagy-related 8)-family proteins and facilitate the selective sequestration of specific cargoes for autophagic degradation. In addition, Atg8 interacts with a number of adaptors essential for autophagosome biogenesis, including ATG and non-ATG proteins. The majority of these adaptors and receptors are characterized by an Atg8-family interacting motif (AIM) for binding to Atg8. However, the molecular basis for the interaction mode between ATG8 and regulators or cargo receptors in plants remains largely unclear. In this study, we unveiled an atypical interaction mode for Arabidopsis ATG8f with a plant unique adaptor protein, SH3P2 (SH3 domain-containing protein 2), but not with the other two SH3 proteins. By structure analysis of the unbound form of ATG8f, we identified the unique conformational changes in ATG8f upon binding to the AIM sequence of a plant known autophagic receptor, NBR1. To compare the binding affinity of SH3P2-ATG8f with that of ATG8f-NBR1, we performed a gel filtration assay to show that ubiquitin-associated domain of NBR1 outcompetes the SH3 domain of SH3P2 for ATG8f interaction. Biochemical and cellular analysis revealed that distinct interfaces were employed by ATG8f to interact with NBR1 and SH3P2. Further subcellular analysis showed that the AIM-like motif of SH3P2 is essential for its recruitment to the phagophore membrane but is dispensable for its trafficking in endocytosis. Taken together, our study provides an insightful structural basis for the ATG8 binding specificity toward a plant-specific autophagic adaptor and a conserved autophagic receptor.Abbreviations: ATG, autophagy-related; AIM, Atg8-family interacting motif; BAR, Bin-Amphiphysin-Rvs; BFA, brefeldin A; BTH, benzo-(1,2,3)-thiadiazole-7-carbothioic acid S-methyl ester; CCV, clathrin-coated-vesicle; CLC2, clathrin light chain 2; Conc A, concanamycin A; ER, endoplasmic reticulum; LDS, LIR docking site; MAP1LC3/LC3, microtubule associated protein 1 light chain 3; LIR, LC3-interacting region; PE, phosphatidylethanolamine; SH3P2, SH3 domain containing protein 2; SH3, Src-Homology-3; UBA, ubiquitin-associated; UIM, ubiquitin-interacting motif.

AS1 expression in prostate cancer and its effects on proliferation and invasion of prostate cancer cells.

This paper aimed at investigating AS1 expression in prostate cancer (PCa) and its effects on the proliferation and invasion of prostate cancer cells (PCCs). The prostate tissues and the matched adjacent normal prostate tissues excised and preserved during radical prostatectomy in our hospital were collected. The LncRNA NCK1-AS1 expression was detected. PCa patients were followed up for three years to analyze their prognosis. The correlation of LncRNA NCK1-AS1 expression with clinicopathological features was analyzed. Human normal prostate cells and human PCCs were selected, in which LncRNA NCK1-AS1 expression was tested to screen and then transfect the cells. Cell proliferation, invasion and migration were detected. Cell cycles and apoptosis were analyzed. Compared with the adjacent normal tissues, LncRNA NCK1-AS1 was highly expressed in the prostate cancer tissues. Its expression was remarkably different in those with different stages of TNM and with lymphatic metastasis or not. The prognosis of patients with high LncRNA NCK1-AS1 expression was remarkably poorer than that of those with low expression. Compared with the human normal prostate cells, LncRNA NCK1-AS1 expression in the human PCCs remarkably rose, with the greatest difference in 22Rv1 cells. Compared with the Blank group, cell proliferation and the number of plate cloned cells remarkably reduced in the sh-NCK1-AS1 group. Additionally, in this group, the number of invasive and migratory cells remarkably reduced; the expression of invasion-related protein E-cadherin remarkably rose but that of MMP-2 remarkably reduced; cell cycles were arrested and the expression of cycle-related proteins (CDK4, CDK6, cyclin D1) remarkably reduced; the apoptotic rate and the expression of apoptosis-related protein Bax remarkably rose. LncRNA NCK1-AS1 is highly expressed in PCa, so its down-regulation can inhibit PCCs from proliferating and reduce the number of invasive cells.