Roles of host proteases in the entry of SARS-CoV-2

The spike protein(S)of SARS-CoV-2 is responsible for viral attachment and entry,thus a major factor for host suscep-tibility,tissue tropism,virulence and pathogenicity.The S is divided with S1 and S2 region,and the S1 contains the receptor-binding domain(RBD),while the S2 contains the hydrophobic fusion domain for the entry into the host cell.Numerous host proteases have been implicated in the activation of SARS-CoV-2 S through various c leavage sites.In this article,we review host proteases including furin,trypsin,transmembrane protease serine 2(TMPRSS2)and cathepsins in the activation of SARS-CoV-2 S.Many betacoronaviruses including SARS-CoV-2 have polybasic residues at the S1/S2 site which is subjected to the cleavage by furin.The S1/S2 cleavage facilitates more assessable RBD to the receptor ACE2,and the binding triggers further conformational changes and exposure of the S2'site to proteases such as type Il transmembrane serine proteases(TTPRs)including TMPRSS2.In the presence of TMPRSS2 on the target cells,SARS-CoV-2 can utilize a direct entry route by fusion of the viral envelope to the cellular membrane.In the absence of TMPRSS2,SARS-CoV-2 enter target cells via endosomes where multiple cathepsins cleave the S for the successful entry.Additional host proteases involved in the cleavage of the S were discussed.This article also includes roles of 3C-like protease inhibitors which have inhibitory activity against cathepsin L in the entry of SARS-CoV-2,and discussed the dual roles of such inhibitors in virus replication.

Highly Sensitive Poly-N-isopropylacrylamide Microgel-based Electrochemical Biosensor for the Detection of SARS-COV-2 Spike Protein

Objective Late 2019 witnessed the outbreak and widespread transmission of coronavirus disease 2019(COVID-19),a new,highly contagious disease caused by novel severe acute respiratory syndrome coronavirus 2(SARS-CoV-2).Consequently,considerable attention has been paid to the development of new diagnostic tools for the early detection of SARS-CoV-2.Methods In this study,a new poly-N-isopropylacrylamide microgel-based electrochemical sensor was explored to detect the SARS-CoV-2 spike protein(S protein)in human saliva.The microgel was composed of a copolymer of N-isopropylacrylamide and acrylic acid,and gold nanoparticles were encapsulated within the microgel through facile and economical fabrication.The electrochemical performance of the sensor was evaluated through differential pulse voltammetry.Results Under optimal experimental conditions,the linear range of the sensor was 10-13-10-9 mg/m L,whereas the detection limit was 9.55 fg/mL.Furthermore,the S protein was instilled in artificial saliva as the infected human saliva model,and the sensing platform showed satisfactory detection capability.Conclusion The sensing platform exhibited excellent specificity and sensitivity in detecting spike protein,indicating its potential application for the time-saving and inexpensive detection of SARS-CoV-2.

Recombinant Vaccinia Virus is an Effective and Non-perturbing Vector for Human Dendritic Cells Transfected with Epstein-Barr Virus Latent Membrane Protein 2A

ObjectiveTo study the effects of dendritic cells (DC) transfected with recombinant vaccinia virus encoding Epstein Barr virus (EBV) latent membrane protein 2A(LMP2A) gene,and to provide evidence for further investigation on the therapeutic vaccines against EBV associated malignancies. MethodsMature DC were transfected with EBV LMP2A recombinant vaccinia virus (rVV LMP2A). Before and after the transfection,the expression of surface antigens on mature DC including CD1a,CD83,CD40,CD80,HLA DR was measured by fluorescence activated cell sorter (FACS) and the function of DC to stimulate allogeneic T cells proliferation was measured by mixed leukocyte reactions (MLR). ResultsLMP2A protein was highly expressed (66.1 %) in DC after the transfection of rVV LMP2A. No significant changes in the primary surface antigens expression and in the MLR were detected during the transfection. Transfected DC still had strong potential in stimulating the proliferation of allogeneic T cells. ConclusionRecombinant vaccinia virus was an effective and non perturbing vector to mediate the transfection of LMP2A into DC. The functions of mature DC were not affected significantly by the transfection of Vac LMP2A. This study could provide evidence for the further immunotherapy of EBV associated malignancies,e.g. nasopharyngeal carcinoma (NPC).

血清IL-2、sST2表达与特发性膜性肾病免疫抑制剂治疗反应性的相关性

目的探讨特发性膜性肾病患者血清白介素-2(IL-2)、可溶性生长刺激表达基因2蛋白(sST2)表达水平与免疫抑制剂治疗反应性的相关性。方法选取2020年1月至2022年10月于医院接受免疫抑制剂治疗的135例特发性膜性肾病患者,于入院时检测患者血清IL-2、sST2,并于治疗完成后测定24 h尿蛋白定量,依据患者治疗反应性分为缓解组与未缓解组。对比两组患者一般资料及入院时血清IL-2、sST2水平,采用点二列相关性分析血清IL-2、sST2水平与特发性膜性肾病免疫抑制剂治疗反应性的关系,并绘制受试者工作特征(ROC)曲线评估血清IL-2、sST2水平预测特发性膜性肾病免疫抑制剂治疗反应性的价值。结果135例特发性膜性肾病患者中共有132例完成规律治疗,经免疫抑制剂治疗6个月后,101例患者疾病缓解,纳入缓解组,其余31例患者纳入未缓解组。未缓解组年龄、入院时肾功能分级、疾病分期、血清IL-2、sST2水平均高于缓解组,差异有统计学意义(P<0.05);点二列相关性分析显示,血清IL-2、sST2水平与特发性膜性肾病免疫抑制剂治疗反应性不良风险呈正相关(r 1=0.428,P 1<0.001;r 2=0.344,P 2<0.001);绘制ROC曲线,结果显示,血清IL-2、sST2预测特发性膜性肾病免疫抑制剂治疗反应性不良的曲线下面积均>0.7,具有一定预测价值,且联合预测价值更高。结论血清IL-2、sST2表达水平与特发性膜性肾病患者免疫抑制剂治疗反应性密切相关,二者表达水平越高,治疗反应性越差,且联合检测可作为预测特发性膜性肾病患者免疫抑制剂治疗反应性的敏感指标。

Analysis of Sperm Membrane Protein Relevant to Antisperm Antibody by Two-Dimensional Gel Electrophoresis and Western Blotting

Objective To identify the sperm membrane proteins that are associated with antisperm antibody Methods Using antisperm antibody positive serum through unidimensional polyacrylamide gel electrophoresis and 2-dimensional gel electrophoresis followed by Western blot analysis to determine the molecular weights (MW) and isoelectric points (pI) of sperm membrane proteins that are associated with antisperm antibody. Results Eight kinds of MW with more than ten sperm membrane proteins can be recognized by antisperm antibody positive serum, of which the MWs and pI were 23 kD, 31 kD, 32 kD, 34 kD, 41 kD, 51 kD, 60 kD, 78 kD and 5.3, 5.5,5.7, 5.0, 5.3, 5.8, 6.0, 5.5～6.2, 4.6,5.1,5.5～5.8 respectively. The identification ratios of the sperm membrane proteins on 78 kD (60.7%), 60 kD (71.4%), 51 kD (14.9%) and 23 kD (14.29%) were higher. Conclusion The sperm membrane proteins with MW of 78 kD, 60 kD, 51 kD and 23 kD were associated with antisperm antibody and immunological infertility. Two- dimensional gel electrophoresis and Western blotting can precisely identify the sperm membrane proteins that are associated with antisperm antibody.

Mutations in spike protein and allele variations in ACE2 impact targeted therapy strategies against SARS-CoV-2

Coronavirus disease 2019(COVID-19), which is caused by severe acute respiratory syndrome coronavirus 2(SARS-Co V-2), has spread rapidly worldwide with high rates of transmission and substantial mortality. To date, however, no effective treatments or enough vaccines for COVID-19 are available. The roles of angiotensin converting enzyme 2(ACE2) and spike protein in the treatment of COVID-19 are major areas of research. In this study, we explored the potential of ACE2 and spike protein as targets for the development of antiviral agents against SARS-Co V-2. We analyzed clinical data, genetic data, and receptor binding capability.Clinical data revealed that COVID-19 patients with comorbidities related to an abnormal reninangiotensin system exhibited more early symptoms and poorer prognoses. However, the relationship between ACE2 expression and COVID-19progression is still not clear. Furthermore, if ACE2 is not a good targetable protein, it would not be applicable across a wide range of populations. The spike-S1 receptor-binding domain that interacts with ACE2 showed various amino acid mutations based on sequence analysis. We identified two spike-S1 point mutations(V354 F and V470 A) by receptorligand docking and binding enzyme-linked immunosorbent assays. These variants enhanced the binding of the spike protein to ACE2 receptors and were potentially associated with increased infectivity. Importantly, the number of patients infected with the V354 F and V470 A mutants has increased with the development of the SARS-Co V-2 pandemic. These results suggest that ACE2 and spike-S1 are likely not ideal targets for the design of peptide drugs to treat COVID-19 in different populations.

特发性膜性肾病患者血清ROCK2、renalase水平及其诊断价值

目的探讨Rho相关卷曲螺旋形成蛋白激酶2(ROCK2)、肾胺酶(renalase)在特发性膜性肾病(IMN)患者血清中的表达变化及其对IMN的诊断价值。方法选取2019年1月—2020年11月河北以岭医院肾病科收治IMN患者120例为IMN组,另选取同期医院进行健康体检的志愿者120例为健康对照组。酶联免疫吸附法(ELISA)检测受试者血清ROCK2、renalase水平,Pearson法分析ROCK2、renalase水平与部分临床指标的相关性;采用受试者工作特征(ROC)曲线分析血清ROCK2、renalase及二者联合诊断IMN的价值。结果IMN组ROCK2、renalase水平均显著高于健康对照组(t/P=12.837/<0.001、11.066/<0.001)。Ⅰ期、Ⅱ期、Ⅲ期、Ⅳ期IMN患者血清ROCK2逐次升高(F/P=77.154/<0.001),而Ⅰ期、Ⅱ期、Ⅲ期血清renalase水平逐次升高,但Ⅳ期血清renalase水平低于Ⅲ期(F/P=163.042/<0.001)。Alb与血清ROCK2、renalase水平呈负相关(r/P=-0.302/0.009、-0.402/<0.001),PLA2R抗体、BUN、SCr、24 h尿蛋白与血清ROCK2、renalase水平均呈正相关(r/P=0.336/<0.001、0.264/0.011、0.315/0.007、0.320/<0.001,0.359/<0.001、0.284/0.010、0.420/<0.001、0.412/<0.001);ROC曲线显示:血清ROCK2、renalase及二者联合诊断IMN的AUC分别为0.908、0.907、0.965,二者联合优于各自单独预测效能(Z=3.597,3.755,P均<0.001)。结论ROCK2与renalase在IMN患者血清中均高表达,二者联合诊断IMN的价值较高。

穿山龙总皂苷对膜性肾病大鼠肾组织M型PLA2R和IgG4表达的影响及其机制

目的 分析穿山龙总皂苷对膜性肾病大鼠肾组织M型磷脂酶A2受体(Phospholipase A2 receptor, PLA2R)和免疫球蛋白G亚型4(Immunoglobulin G4,IgG4)表达影响及可能机制。方法 将40只SPF级雄性SD大鼠按随机数字表法分为对照组、模型组、贝那普利组(10 mg/kg)、低和高剂量穿山龙总皂苷组(80 mg/kg、160 mg/kg),每组各8只。除对照组,其余4组采用Border法制备膜性肾病模型,造模成功后,贝那普利组灌胃给予贝那普利10 mg/(kg·d),低和高剂量穿山龙总皂苷组分别灌胃给予穿山龙总皂苷80 mg/(kg·d)、160 mg/(kg·d),对照组、模型组灌胃给予10 ml/(kg·d)生理盐水。连续给药4周后,检测24 h尿蛋白、白蛋白、血肌酐、血尿素氮、尿酸水平,HE染色观察肾脏病理改变,蛋白免疫印迹法检测肾脏中M型PLA2R、IgG4、磷酸化磷脂酰肌醇3-激酶(Phosphorylated phosphoinositide 3-kinase, p-PI3K)、磷酸化蛋白激酶B(Phosphorylated protein kinase B,p-AKT)、核因子E2相关因子2(Nuclear factor E2-related factor 2,Nrf2)、血红素加氧酶(Heme oxygenase-1,HO-1)表达水平。结果 与模型组比较,贝那普利组、高剂量穿山龙总皂苷组白蛋白水平明显升高,血肌酐、血尿素氮、尿酸水平明显降低,差异均有统计学意义(P>0.05)。与模型组比较,贝那普利组、低剂量和高剂量穿山龙总皂苷组肾脏病理改变明显改善,24 h尿蛋白水平及肾脏中M型PLA2R、IgG4、p-PI3K、p-AKT表达水平明显降低,肾脏中Nrf2、HO-1表达水平明显增加,差异均有统计学意义(P<0.05)。结论 穿山龙总皂苷对膜性肾病大鼠的肾脏具有保护作用,其机制可能与降低PLA2R、IgG4表达,抑制PI3K/AKT通路,激活Nrf2/HO-1通路相关。

JTE-522-induced apoptosis in human gastric adenocarinoma cell line AGS cells by caspase activation accompanying cytochrome C release,membrane translocation of Bax and loss of mitochondrial membrane potential

瞄准：在 JTE-522-induced apoptosis 调查 mitochondrial 小径的角色并且调查在细胞色素 C 版本， caspase 活动和 mitochondrial 膜潜力(Deltapsim ) 的损失之间的关系。方法：房间文化，数的房间， ELISA 试金， TUNEL，流动 cytometry，西方的污点和 fluorometric 试金被采用在 AGS 房间和相关分子的机制在房间增长和 apoptosis 上调查 JTE-522 的效果。结果：JTE-522 禁止了 AGS 细胞的生长并且导致了 apoptosis。Caspases 8 并且 9 在从 procaspase 和 caspase 活动由劈开产品的外观判定了劈开特定的 fluorogenic 底层的 apoptosis 期间被激活。阐明是否 caspases 的激活 8 和 9 为 apoptosis 正式就职被要求，我们在 apoptosis 上检验了 caspase 特定的禁止者的效果。结果证明 caspase 禁止者显著地禁止了 JTE-522 导致的 apoptosis。另外，与 Rhodamin 123 的举起的减少伴随的细胞色素 C 的 Bax 和 cytosolic 版本的膜 translocation，在 apoptosis 的一个早阶段被检测。而且， Bax translocation，细胞色素 C 版本，和 caspase 9 激活被 Z-VAD.fmk 和 Z-IETD-CHO 堵住。结论：现在的数据显示在 caspases 的激活之间的一个关键协会 8， 9，细胞色素 C 版本， Bax 的膜 translocation， Deltapsim 的损失和在 AGS 房间的 JTE-522-induced apoptosis。

Bioabsorbable Barrier Membrane Combined with rhBMP-2 Improved Bone Formation in an Experimental Model of Compromised Healing But Was Not Superior to rhBMP-2 Alone

Objective: Bioabsorbable barrier membranes placed over alveolar ridge bone defects are routinely used in dental surgery to promote bone formation. Combining these osteoconductive membranes with osteoinductive Bone Morphogenetic Proteins could prove useful in long bone fracture treatment. The hypothesis was tested in a clinically relevant model of compromised healing. Methods: Four groups of 8 rabbits underwent unilateral mid-tibial osteotomy, excision of periosteum and endosteum, and plate fixation. One group had rhBMP-2 deposited between the bone ends and Membrane wrapped around the osteotomy, the second group had Membrane wrapped around the osteotomy, the third group had rhBMP-2 placed between the bone ends, and the fourth group received no additional treatment. Results: After 7 weeks, callus size and blood flow were significantly higher in the Membrane+rhBMP-2 group than in the rhBMP-2 treated group, but torsion to failure test showed no significant difference. Membrane treatment and no treatment led to non-union. Conclusion: Absorbable barrier membrane combined with rhBMP-2 enhances bone formation, but has no advantage to rhBMP-2 alone. Membrane alone wrapped around the osteotomy was unable to prevent non-union formation.

Extracorporeal membrane oxygenation for coronavirus disease 2019-associated acute respiratory distress syndrome:Report of two cases and review of the literature

BACKGROUND Coronavirus disease 2019(COVID-19),caused by severe acute respiratory syndrome coronavirus-2,is a worldwide pandemic.Some COVID-19 patients develop severe acute respiratory distress syndrome and progress to respiratory failure.In such cases,extracorporeal membrane oxygenation(ECMO)treatment is a necessary life-saving procedure.CASE SUMMARY Two special COVID-19 cases—one full-term pregnant woman and one elderly(72-year-old)man—were treated by veno-venous(VV)-ECMO in the Second People’s Hospital of Zhongshan,Zhongshan City,Guangdong Province,China.Both patients had developed refractory hypoxemia shortly after hospital admission,despite conventional support,and were therefore managed by VV-ECMO.Although both experienced multiple ECMO-related complications on top of the COVID-19 disease,their conditions improved gradually.Both patients were weaned successfully from the ECMO therapy.At the time of writing of this report,the woman has recovered completely and been discharged from hospital to home;the man remains on mechanical ventilation,due to respiratory muscle weakness and suspected lung fibrosis.As ECMO itself is associated with various complications,it is very important to understand and treat these complications to achieve optimal outcome.CONCLUSION VV-ECMO can provide sufficient gas exchange for COVID-19 patients with acute respiratory distress syndrome.However,it is crucial to understand and treat ECMO-related complications.

Membrane Proteins as Potential Colon Cancer Biomarkers: Verification of 4 Candidates from a Secretome Dataset

Colorectal cancer (CRC) is an important health issue in Taiwan. There were over ten thousand newly diagnosed CRC patients each year. The outcome of late stage CRC still remains to be improved, and tumor markers are expected to improve CRC detection and management. From a colorectal cancer cell secretome database, we chose four proteins as candidates for clinical verification, including tumor-associated calcium signal transducer 2 (TROP2, TACSTD2), transmembrane 9 superfamily member 2 (TM9SF2), and tetraspanin-6 (TSPAN6), and tumor necrosis factor receptor superfamily member 16 (NGFR). Different groups of 30 CRC patients’ tissue samples collected from Chang Gung Memorial Hospital were analyzed by immunohistochemistry (IHC) for the four proteins, and the results were scored by pathologist. For all the four candidate proteins, marked differences of IHC score existed between tumor and adjacent non-tumor counterpart. However, there were only trends between higher protein expression levels and worse outcome. Three proteins (TROP2, TM9SF2 and NGFR) had trends between higher tissue expression and tumor stage or lymph node metastasis. Our study revealed that tissue expression of four proteins (TROP2, TM9SF2, TSPAN6, and NGFR) was markedly different between tumor and adjacent non-tumor counterparts. Overexpression of all these four proteins showed some trends with poorer survival.

<i>Morinda citrifolia</i>(Noni) Fruit Juice Inhibits SARS-CoV-2 Spike Protein Binding of Angiotensin-Converting Enzyme 2 (ACE2)

COVID-19 is a global pandemic that has claimed millions of lives. This disease is caused by a coronavirus, SARS-CoV-2, which requires the binding of its spike protein to angiotensin-converting enzyme 2 (ACE2) for infection of the host cell. Morinda citrifolia (noni) fruit juice has antiviral activity that involves enhancement of immune system function. SARS-CoV-2 spike-ACE2 interaction experiments were carried out to further investigate the antiviral properties of noni juice and its major iridoids. Noni juice inhibited binding by approximately 69%. Scandoside was the most active of the three iridoids evaluated, reducing average spike protein-ACE2 interaction by 79.25%. The iridoids worked synergistically towards inhibiting spike protein binding when assayed together, improving activity by more than 22% above the expected level. But the modest activity of the most abundant iridoid, deacetylasperulosidic acid, indicates that other phytochemicals (i.e. scopoletin, quercetin, rutin and kaempferol) are also involved. Our results suggest that the presence of several biological active phytochemicals in noni juice enhances resistance to SARS-CoV-2 by interfering with its ability to bind ACE2. This is a new and significant anti-viral mechanism of noni juice that does not directly involve its immunomodulatory properties.

磷酸钙复合重组人骨形态发生蛋白2修复和重建胫骨感染性骨缺损

背景:尽管Masquelet技术在临床的应用取得了广泛成功,但目前优化该技术各个环节的研究仍在不断开展,其中加快植骨后骨愈合速度、缩短骨愈合时间是医生关注的焦点。目的:观察磷酸钙复合重组人骨形态发生蛋白2材料修复胫骨感染性骨缺损的效果。方法:选择2017年6月至2022年6月简阳市人民医院收治的感染性胫骨缺损患者31例,均接受Masquelet技术分期治疗,二期手术根据植骨材料的不同分为对照组(n=15)与研究组(n=16),对照组植入异体骨/自体骨颗粒,研究组植入磷酸钙复合重组人骨形态发生蛋白2/自体骨颗粒。二期术后6个月,检测患者外周血白细胞数、C-反应蛋白、血沉等炎性指标,记录影像学骨愈合时间、骨愈合X射线评分、骨缺损愈合分级和邻近关节功能,观察是否存在钉道感染、植入吸收、取骨区疼痛、感染等情况。结果与结论:①两组患者二期术后6个月外周血白细胞计数、血沉、C-反应蛋白水平均低于一期术前(P<0.05),两组间各指标比较差异无显著性意义(P>0.05);②研究组骨愈合时间短于对照组(P<0.05);③研究组患者二期术后6个月的骨愈合X射线评分高于对照组(P<0.05),骨缺损愈合优良率与邻近关节功能优良率均高于对照组(P<0.05);两组间感染复发率与并发症发生率比较差异均无显著性意义(P>0.05);④结果表明,Masquelet技术二期术中应用磷酸钙复合重组人骨形态发生蛋白2治疗胫骨感染性骨缺损的愈合效果良好、安全性高。

Universal COVID-19 Vaccine Targeting SARS-CoV-2 Envelope Protein

The coronavirus disease 2019 (COVID-19) pandemic, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), had caused over 382 million cases and over 2.7 million deaths globally as of 23 March 2021. By that date, at least 10 SARS-CoV-2 variants had emerged. The transmissibility and lethality of the variants are higher than those of the Wuhan reference strain. Therefore, a universal vaccine for the reference strain and all variants (present and future) is indispensable. The coronavirus envelope (E) protein is an integral membrane protein crucial to the viral lifecycle and the pathogenesis of coronaviruses. The SARS-CoV-2 E protein has a postsynaptic density protein 95/Drosophila disc large tumor suppressor/zonula occludens-1 (PDZ) binding motif (PBM), and its interaction with PDZ-domain-2 of the human tight junction protein may interrupt the integrity of lung epithelium. Furthermore, the SARS-CoV-2 E protein itself is a homopentameric cation channel viroporin, which may be involved in viral release. This protein is thus a potential target for the development of a universal COVID-19 vaccine, because of its highly conserved amino acid sequence. The variant mutations occur mainly in the spike protein, and conservation of E protein remained in most Variants of Concern (VOC). Only one of the extant VOC have mutations in the E protein that P71L mutation occurs in the South African variant 501Y.V2 (B.1.351). If a vaccine is designed to target E protein, two scenarios are possible: 1) SARS-CoV-2 maintains a highly conserved E protein amino acid sequence, rendering the virus consistently or permanently susceptible to the vaccine;or 2) the E protein mutates and new variants evolve accordingly. In scenario 2, the tertiary structure and function of the E protein homopentameric cation channel viroporin, PBM, or other aspects affecting pathogenicity would be attenuated. Either scenario would thus ameliorate the pandemic. I therefore propose that a vaccine targeting the SARS-CoV-2 E protein would be effective against the Wuhan reference strain and all current and future SARS-CoV-2 variants. Efforts to create E protein-based vaccines are ongoing. Further research and clinical trials are needed to realize this universal COVID-19 vaccine.

Ciclopirox inhibits SARS-CoV-2 replication by promoting the degradation of the nucleocapsid protein

The nucleocapsid protein(NP)plays a crucial role in SARS-CoV-2 replication and is the most abundant structural protein with a long half-life.Despite its vital role in severe acute respiratory syndrome coronavirus 2(SARS-CoV-2)assembly and host inflammatory response,it remains an unexplored target for drug development.In this study,we identified a small-molecule compound(ciclopirox)that promotes NP degradation using an FDA-approved library and a drug-screening cell model.Ciclopirox significantly inhibited SARS-CoV-2 replication both in vitro and in vivo by inducing NP degradation.Ciclopirox induced abnormal NP aggregation through indirect interaction,leading to the formation of condensates with higher viscosity and lower mobility.These condensates were subsequently degraded via the autophagy-lysosomal pathway,ultimately resulting in a shortened NP half-life and reduced NP expression.Our results suggest that NP is a potential drug target,and that ciclopirox holds substantial promise for further development to combat SARS-CoV-2 replication.

Artificial Nucleic Acid Tractor-Directed Simultaneous Depletion of Oncogenic Membrane Proteins Without Hijacking Proteolysis-Specific Actuator

Targeted protein degradation(TPD)is an emerging tool for degrading proteins of interest,which affords an attractive modality for cancer therapy.However,the present TPD technologies must engage a proteolysis-specific actuator to initiate degradation of targeted proteins in the proteasome or lysosome.Herein,we report an artificial tractor that can induce endocytosis-mediated protein depletion without hijacking a proteolysis-specific actuator.In this design,bispecific aptamer chimeras(BSACs)are established,which can bridge human epidermal growth factor receptor 2(ErbB-2),an important biomarker in a common important biomarker in cancer,with membrane proteins of interest.Taking advantage of the property of aptamer-induced endocytosis and digestion of ErbB-2,another membrane protein is translocated into the lysosome in a hitchhike-like manner,resulting in lysosomal proteolysis along with ErbB-2.This strategy frees the TPD from the fundamental limitation of proteolysis-specific actuator and allows simultaneous regulation of the quantity and function of two oncogenic receptors in a cell-type-specific manner,expanding the application scope of TPD-based therapeutics.

The endoplasmic reticulum membrane protein complex subunit Emc6 is essential for rhodopsin localization and photoreceptor cell survival

The endoplasmic reticulum(ER)membrane protein complex(EMC)is responsible for monitoring the biogenesis and synthetic quality of membrane proteins with tail-anchored or multiple transmembrane domains.The EMC subunit EMC6 is one of the core members of EMC and forms an enclosed hydrophilic vestibule in cooperation with EMC3.Despite studies demonstrating that deletion of EMC3 led to rhodopsin mislocalization in rod photoreceptors of mice,the precise mechanism leading to the failure of rhodopsin trafficking remains unclear.Here,we generated the first rod photoreceptor-specific knockout of Emc6(RKO)and cone photoreceptor-specific knockout of Emc6(CKO)mouse models.Deficiency of Emc6 in rod photoreceptors led to progressive shortening of outer segments(OS),impaired visual function,mislocalization and reduced expression of rhodopsin,and increased gliosis in rod photoreceptors.In addition,CKO mice displayed the progressive death of cone photoreceptors and abnormal localization of cone opsin protein.Subsequently,proteomics analysis of the RKO mouse retina illustrated that several cilium-related proteins,particularly anoctamin-2(ANO2)and transmembrane protein 67(TMEM67),were significantly down-regulated prior to OS degeneration.Detrimental rod photoreceptor cilia and mislocalized membrane disc proteins were evident in RKO mice.Our data revealed that in addition to monitoring the synthesis of rhodopsin-dominated membrane disc proteins,EMC6 also impacted rod photoreceptors'ciliogenesis by regulating the synthesis of membrane proteins associated with cilia,contributing to the mislocalization of membrane disc proteins.

Structural basis of SARS-CoV-2 3CL^pro and anti-COVID-19 drug discovery from medicinal plants

The recent pandemic of coronavirus disease 2019(COVID-19)caused by SARS-CoV-2 has raised global health concerns.The viral 3-chymotrypsin-like cysteine protease(3CL^pro)enzyme controls coronavirus replication and is essential for its life cycle.3CL^pro is a proven drug discovery target in the case of severe acute respiratory syndrome coronavirus(SARS-CoV)and Middle East respiratory syndrome coronavirus(MERS-CoV).Recent studies revealed that the genome sequence of SARS-CoV-2 is very similar to that of SARS-CoV.Therefore,herein,we analysed the 3CL^pro sequence,constructed its 3D homology model,and screened it against a medicinal plant library containing 32,297 potential anti-viral phytochemicals/traditional Chinese medicinal compounds.Our analyses revealed that the top nine hits might serve as potential anti-SARS-CoV-2 lead molecules for further optimisation and drug development process to combat COVID-19.

Charge‑Transfer Resonance and Electromagnetic Enhancement Synergistically Enabling MXenes with Excellent SERS Sensitivity for SARS‑CoV‑2 S Protein Detection

The outbreak of coronavirus disease 2019 has seriously threatened human health.Rapidly and sensitively detecting SARSCoV-2 viruses can help control the spread of viruses.However,it is an arduous challenge to apply semiconductor-based substrates for virus SERS detection due to their poor sensitivity.Therefore,it is worthwhile to search novel semiconductor-based substrates with excellent SERS sensitivity.Herein we report,for the first time,Nb2C and Ta2C MXenes exhibit a remarkable SERS enhancement,which is synergistically enabled by the charge transfer resonance enhancement and electromagnetic enhancement.Their SERS sensitivity is optimized to 3.0×10^6 and 1.4×10^6 under the optimal resonance excitation wavelength of 532 nm.Additionally,remarkable SERS sensitivity endows Ta2C MXenes with capability to sensitively detect and accurately identify the SARS-CoV-2 spike protein.Moreover,its detection limit is as low as 5×10^−9 M,which is beneficial to achieve real-time monitoring and early warning of novel coronavirus.This research not only provides helpful theoretical guidance for exploring other novel SERS-active semiconductor-based materials but also provides a potential candidate for the practical applications of SERS technology.