Small interfering RNAs(siRNA)provide a novel and highly specific therapy due to their ability to effectively silence target genes,to date six siRNA therapeutics are approved for clinical use.Even so,some critical chal...Small interfering RNAs(siRNA)provide a novel and highly specific therapy due to their ability to effectively silence target genes,to date six siRNA therapeutics are approved for clinical use.Even so,some critical challenges remain to overcome in the therapeutic application of siRNAs,with delivery issues at the forefront.Among them,endo/lysosomal barrier is one of the important but often-neglected limitations hindering the delivery of siRNA therapeutics.In this review,we summarize the promising strategies that facilitate siRNAs overcoming endo/lysosomal barriers based on the cellular uptake and intracellular transport pathways,including promoting escape once endocytosis into the endo/lysosomes and bypassing lysosomes via endosome-Golgi-endoplasmic reticulum(ER)pathway or nonendocytosis pathway,and discuss the principal considerations and the future directions of promoting endo/lysosomal escape in the development of therapeutic siRNAs.展开更多
Efficient siRNA delivery is highly desirable for disease treatment.However,the application of conventional nanoparticles is limited by the inability to escape from endo-lysosomes.Herein,we report a strategy using smal...Efficient siRNA delivery is highly desirable for disease treatment.However,the application of conventional nanoparticles is limited by the inability to escape from endo-lysosomes.Herein,we report a strategy using small-molecule drugs to enhance siRNA endo-lysosomal release,addressing this challenge.We encapsulated gentamicin(GM)into the marketed Onpattro■ formulation to establish LNP-siRNA/GM nanoparticles that promote siRNA endo-lysosomal escape through endosomal disruption,mechanistically exhibiting unique functionality and synergistic effects of LNP-siRNA/GM to improve cancer therapy.Besides,GM induced reactive oxygen species(ROS)and phospholipids accumulation in endolysosomes,as well as the physical characteristics of lipid nanoparticles(LNPs)were preserved.We also revealed that GM causes endo-lysosomal swelling and disrupts the endosomal membrane to enable siRNA release,as confirmed by Galectin 3 recruitment and acridine orange release.This approach achieved∼81%mRNA-EGFR silencing,which is more than LNP-siEGFR(∼56.23%)by enhancing siRNA endo-lysosomal escape efficiency.Meanwhile,LNP-siEGFR/GM exhibited significant biological activities in HepG2 cells,driven by the synergistic effects of siEGFR and GM with the VEGF and CXCL12 downregulation of,and ROS and phospholipids upregulation.Furthermore,tumor growth was notably suppressed after intravenous injection of LNP-siEGFR/GM in tumor-bearing nude mice.The combination of EGFR-siRNA and GM could also greatly inhibit angiogenesis,be antiproliferative,and induce tumor cells apoptosis.Therefore,this GM and siRNA co-delivery system would provide an efficient strategy for siRNA endosomal escape,significantly improving knockdown in various LNPs based siRNA delivery systems and efficiently enhancing cancer therapy.展开更多
With the identification of more than a dozen novel Hermansky-Pudlak Syndrome （HPS） proteins in vesicle trafficking in higher eukaryotes, a new class of trafficking pathways has been described. It mainly consists of ...With the identification of more than a dozen novel Hermansky-Pudlak Syndrome （HPS） proteins in vesicle trafficking in higher eukaryotes, a new class of trafficking pathways has been described. It mainly consists of three newly-defined protein com- plexes, BLOC-l, -2, and -3. Compelling evidence indicates that these complexes together with two other well-known complexes, AP3 and HOPS, play important roles in endosomal transport. The interactions between these complexes form a network in protein trafficking via endosomes and cytoskeleton. Each node of this network has intra-complex and extra-complex interactions. These complexes are connected by direct interactions between the subunits from different complexes or by indirect interactions through coupling nodes that interact with two or more subunits from different complexes. The dissection of this network facilitates the understanding of a dynamic but elaborate transport machinery in protein/membrane trafficking. The disruption of this network may lead to abnormal trafficking or defective organellar development as described in patients with Hermansky-Pudlak syndrome.展开更多
Human immunodeficiency virus-1(HIV-1)encodes simply 15 proteins and thus depends on multiple host cellular factors for virus reproduction.Spastin,a microtubule severing protein,is an identified HIV-1 dependency factor...Human immunodeficiency virus-1(HIV-1)encodes simply 15 proteins and thus depends on multiple host cellular factors for virus reproduction.Spastin,a microtubule severing protein,is an identified HIV-1 dependency factor,but the mechanism regulating HIV-1 is unclear.Here,the study showed that knockdown of spastin inhibited the production of the intracellular HIV-1 Gag protein and new virions through enhancing Gag lysosomal degradation.Further investigation showed that increased sodium tolerance 1(IST1),the subunit of endosomal sorting complex required for transport(ESCRT),could interact with the MIT domain of spastin to regulate the intracellular Gag production.In summary,spastin is required for HIV-1 replication,while spastin-IST1 interaction facilitates virus production by regulating HIV-1 Gag intracellular trafficking and degradation.Spastin may serve as new target for HIV-1 prophylactic and therapy.展开更多
Hepatitis B virus(HBV)produces and releases various particle types,including complete virions,subviral particles with envelope proteins,and naked capsids.Recent studies demonstrate that HBV exploits distinct intracell...Hepatitis B virus(HBV)produces and releases various particle types,including complete virions,subviral particles with envelope proteins,and naked capsids.Recent studies demonstrate that HBV exploits distinct intracellular membrane trafficking pathways,including the endosomal vesicle trafficking and autophagy pathway,to assemble and release viral and subviral particles.Herein,we summarize the findings about the distinct roles of autophagy and endosomal membrane trafficking and the interaction of both pathways in HBV replication,assembly,and release.展开更多
Rabies virus(RABV) is a highly neurotropic virus that follows clathrin-mediated endocytosis and p H-dependent pathway for trafficking and invasion into endothelial cells. Early(Rab5, EEA1) and late(Rab7, LAMP1) endoso...Rabies virus(RABV) is a highly neurotropic virus that follows clathrin-mediated endocytosis and p H-dependent pathway for trafficking and invasion into endothelial cells. Early(Rab5, EEA1) and late(Rab7, LAMP1) endosomal proteins play critical roles in endosomal sorting, maturity and targeting various molecular cargoes, but their precise functions in the early stage of RABV neuronal infection remain elusive. In this study, the relationship between enigmatic entry of RABV with these endosomal proteins into neuronal and SH-SY5 Y cells was investigated.Immunofluorescence, TCID_(50) titers, electron microscopy and western blotting were carried out to determine the molecular interaction of the nucleoprotein(N) of RABV with early or late endosomal proteins in these cell lines. The expression of N was also determined by down-regulating Rab5 and Rab7 in both cell lines through RNA interference. The results were indicative that N proficiently colocalized with Rab5/EEA1 and Rab7/LAMP1 in both cell lines at 24 and 48 h post-infection, while N titers significantly decreased in early infection of RABV. Down-regulation of Rab5 and Rab7 did not inhibit N expression, but it prevented productive infection via blocking the normal trafficking of RABV in a low pH environment. Ultrathin sections of cells studied by electron microscope also verified the close association of RABV with Rab5 and Rab7 in neurons. From the data it was concluded that primary entry of RABV strongly correlates with the kinetics of Rab-proteins present on early and late vesicles, which provides helpful clues to explain the early events of RABV in nerve cells.展开更多
Objectives:Improper activation of Wnt/β-catenin signaling has been implicated in human diseases.Beyond the well-studied glycogen synthase kinase 3p(GSK3p)and casein kinase 1(CK1),other kinases affecting Wnt/β-cateni...Objectives:Improper activation of Wnt/β-catenin signaling has been implicated in human diseases.Beyond the well-studied glycogen synthase kinase 3p(GSK3p)and casein kinase 1(CK1),other kinases affecting Wnt/β-catenin signaling remain to be defined.Methods:To identify the kinases that modulate Wnt/β-catenin signaling,we applied a kinase small interfering RNA(siRNA)library screen approach.Luciferase assays,immunoblotting,and real-time polymerase chain reaction(PCR)were performed to confirm the regulation o f the Wnt/β-catenin signaling pathway by cyclin-dependent kinase 11(CDK11)and to investigate the underlying mechanism.Confocal immunofluorescence,coimmunoprecipitation(co-IP),and scratch wound assays were used to demonstrate colocalization,detect protein interactions,and explore the function of CDK11.Results:CDK11 was found to be a significant candidate kinase participating in the negative control of Wnt/P-catenin signaling.Down-regulation of CDK11 led to the accumulation of Wnt/β-catenin signaling receptor complexes,in a manner dependent on intact adenomatosis polyposis coli(APC)protein.Further analysis showed that CDK11 modulation of Wnt/P-catenin signaling engaged the endolysosomal machinery,and CDK11 knockdown enhanced the colocalization of Wnt/β-catenin signaling receptor complexes with early endosomes and decreased colocalization with lysosomes.Mechanistically,CDK11 was found to function in Wnt/β-catenin signaling by regulating microtubule stability.Depletion of CDK11 down-regulated acetyl-a-tubulin.Moreover,co-IP assays demonstrated that CDK11 interacts with the a-tubulin deacetylase SIRT2,whereas SIRT2 down-regulation in CDK11-depleted cells reversed the accumulation of Wnt/(3-catenin signaling receptor complexes.CDK11 was found to suppress cell migration through altered W nt/β-catenin signaling.Conclusions:CDK11 is a negative modulator of Wnt/β-catenin signaling that stabilizes microtubules,thus resulting in the dysregulation of receptor complex trafficking from early endosomes to lysosomes.展开更多
To further enhance the transfection efficiency of a micelleplex system based on monomethoxy poly（ethylene glycol）-block-poly（e-caprolactone）-block-poly（L-lysine） （MPEG-b-PCL-b-PLL）, cholesterol （Chol） moieti...To further enhance the transfection efficiency of a micelleplex system based on monomethoxy poly（ethylene glycol）-block-poly（e-caprolactone）-block-poly（L-lysine） （MPEG-b-PCL-b-PLL）, cholesterol （Chol） moieties are attached to the e-termini of PLL segments to obtain MPEG-b-PCL-b-PLL/Chol. The structure and morphology of the copolymer are studied by IH-NMR, TEM and DLS （dynamic light scattering）. The cytotoxicity, cell uptake, endosomal release and mRNA knockdown are studied by MTT （3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide） assay, flow cytometry, CLSM （confocal laser scanning microscopy） and RT-PCR （real-time polymerase chain reaction）. The results show that compared to their precursor MPEG-b-PCL-b-PLL, the cholesterol-grafted copolymer shows significantly lower toxicity, more rapid cellular endocytosis and endosome escape, and consequently displays enhanced siRNA transfection efficiency even at a lower N/P ratio. These improvements are ascribed to enhanced interaction of the cholesterol moieties with both cellular membrane and endosomal membrane. Moreover, effect of the PLL block length is examined. The final conclusion is that long enough PLL segments and incorporation of proper fraction of cholesterol onto the PLL segments benefit the enhancement of siRNA transfection efficiency.展开更多
Endosomes are characterized by the presence of various phosphoinositides that are essential for defining the membrane properties.However,the interplay between endosomal phosphoinositides metabolism and innate immunity...Endosomes are characterized by the presence of various phosphoinositides that are essential for defining the membrane properties.However,the interplay between endosomal phosphoinositides metabolism and innate immunity is yet to be fully understood.Here,our findings highlight the evolutionary continuity of RAB-10/Rab10’s involvement in regulating innate immunity.Upon infection of Caenorhabditis elegans with Pseudomonas aeruginosa,an increase in RAB-10 activity was observed in the intestine.Conversely,when RAB-10 was absent,the intestinal diacylglycerols(DAGs)decreased,and the animal’s response to the pathogen was impaired.Further research revealed that UNC-16/JIP3 acts as an RAB-10 effector,facilitating the recruitment of phospholipase EGL-8 to endosomes.This leads to a decrease in endosomal phosphatidylinositol 4,5-bisphosphate(PI(4,5)P2)and an elevation of DAGs,as well as the activation of the PMK-1/p38 MAPK innate immune pathway.It is noteworthy that the dimerization of UNC-16 is a prerequisite for its interaction with RAB-10(GTP)and the recruitment of EGL-8.Moreover,we ascertained that the rise in RAB-10 activity,due to infection,was attributed to the augmented expression of LET-413/Erbin,and the nuclear receptor NHR-25/NR5A1/2 was determined to be indispensable for this increase.Hence,this study illuminates the significance of endosomal PI(4,5)P2 catabolism in boosting innate immunity and outlines an NHR-25-mediated mechanism for pathogen detection in intestinal epithelia.展开更多
Recent discovery of PYR/PYL/RCAR-type abscisic acid （ABA） receptors has become one of most significant advances in plant science in the past decade. In mammals, endosomal sorting acts as an important pathway to down...Recent discovery of PYR/PYL/RCAR-type abscisic acid （ABA） receptors has become one of most significant advances in plant science in the past decade. In mammals, endosomal sorting acts as an important pathway to downregulate different types of receptors, but its role in plant hormone signaling is poorly understood. Here, we report that an ubiquitin E2-1ike protein, VPS23A, which is a key component of ESCRT-I, negatively regulates ABA signaling. VPS23A has epistatic relationship with PYR/PYL/RCAR-type ABA receptors and disruption of VPS23A enhanced the activity of key kinase OST1 in the ABA signaling pathway under ABA treatment. Moreover, VPS23A interacts with PYR1/PYLs and K63-1inked diubiquitin, and PYL4 possesses K63-1inked ubiquitinated modification in vivo. Further analysis revealed that VPS23A affects the subcellular localization of PYR 1 and the stability of PYL4. Taken together, our results suggest that VPS23A affects PYR1/ PYL4 via vacuole-mediated degradation, providing an advanced understanding of both the turnover of ABA receptors and ESCRTs in plant hormone signaling.展开更多
The 76 amino acid protein ubiquitin （Ub） is highly conserved in all eukaryotic species. It plays important roles in many cellular processes by covalently attaching to the target proteins. The best known function of ...The 76 amino acid protein ubiquitin （Ub） is highly conserved in all eukaryotic species. It plays important roles in many cellular processes by covalently attaching to the target proteins. The best known function of Ub is marking substrate proteins for degra- dation by the 26S proteasome. In fact, other consequences of ubiquitination have been discovered in yeast and mammals, such as membrane trafficking, DNA repair, chromatin modification, and protein kinase activation. The common mechanism underlying these processes is that Ub serves as a signal to sort proteins to the vacuoles or lysosomes for degradation as opposed to 26S proteasome-dependent degradation. To date, several reports have indicated that a similar function of Ub also exists in plants. This review focuses on a summary and analysis of the recent research progress on Ub acting as a signal to mediate endocytosis and endosomal trafficking in plants.展开更多
There are many challenges in developing efficient and target specific delivery systems of small molecule and nucleic acid drugs. Cell membrane presents one of the major barriers for the penetration of hydrophilic macr...There are many challenges in developing efficient and target specific delivery systems of small molecule and nucleic acid drugs. Cell membrane presents one of the major barriers for the penetration of hydrophilic macromolecules across the plasma membrane. Nanocar- riers have been designed to enhance their cellular uptake via endocytosis but following their cellular uptake, endosomal escape is the rate limiting step which restricts the value associated with the enhanced uptake by nanocarriers. Viruses are an excellent model for efficient cytosolic delivery by nanocarriers. Viruses exploit intra- cellular cues to release the genome to cytosol. In this review, we first discuss different endocytic uptake path- ways and endosomal escape mechanisms. We then summarize the existing tools for studying the intracellular trafficking of nanocarriers. Finally, we highlight the important design elements of recent virus-based nanocar- tiers for efficient cellular uptake and endosomal escape.展开更多
Dendrimer,such as dendrigraft poly-L-lysine(DGL)polymers,with high surface charge density,well-defined structure,and narrow poly-dispersity is often employed as a gene vector,but its transfection efficiency is still p...Dendrimer,such as dendrigraft poly-L-lysine(DGL)polymers,with high surface charge density,well-defined structure,and narrow poly-dispersity is often employed as a gene vector,but its transfection efficiency is still partially inhibited due to poor endosomal escape ability.Herein,we used a surface modification strategy to enhance the endosomal escape ability of DGL polymers,and thus improved its gene transfection efficiency.A library of phenylboronic acid(PBA)modified DGL polymers(PBA-DGLs)was designed to screen efficient small interfering RNA(siRNA)vectors.The lead candidate screened from the library shown a capability of inducing nearly 90% gene silencing in MDA-MB-231 cells.The study of the transfection mechanism revealed that PBA modification not only improves siRNA cellular uptake,but,more importantly,endows DGL polymers the ability of endosomal escape.One of the top candidates from polyplexes was further shielded with hyaluronic acid to construct targeted nanoparticles,and the yielding nanoparticles significantly suppressed the tumor growth in a breast cancer model by effective siRNA delivery.This research provides a general and effective strategy to enhance the endosomal escape and transfection efficiency of dendrimer.展开更多
Poly-histidine peptides such as H6(HHHHHH)are used in protein biotechnologies as purification tags,protein-assembling agents and endosomal-escape entities.The pleiotropic properties of such peptides make them appealin...Poly-histidine peptides such as H6(HHHHHH)are used in protein biotechnologies as purification tags,protein-assembling agents and endosomal-escape entities.The pleiotropic properties of such peptides make them appealing to design protein-based smart materials or nanoparticles for imaging or drug delivery to be produced in form of recombinant proteins.However,the clinical applicability of H6-tagged proteins is restricted by the potential immunogenicity of these segments.In this study,we have explored several humanized histidine-rich peptides in tumor-targeted modular proteins,which can specifically bind and be internalized by the target cells through the tumoral marker CXCR4.We were particularly interested in exploring how protein purification,self-assembling and endosomal escape perform in proteins containing the variant histidine-rich tags.Among the tested candidates,the peptide H5 E(HEHEHEHEH)is promising as a good promoter of endosomal escape of the associated fulllength protein upon endosomal internalization.The numerical modelling of cell penetration and endosomal escape of the tested proteins has revealed a negative relationship between the amount of protein internalized into target cells and the efficiency of cytoplasmic release.This fact demonstrates that the His-mediated,proton sponge-based endosomal escape saturates at moderate amounts of internalized protein,a fact that might be critical for the design of protein materials for cytosolic molecular delivery.展开更多
A major impediment in the development of chitosan nanoparticles （CTS NPs） as effective drug delivery vesicles is their rapid clearance from blood and endosome entrapment. To overcome these problems, a convenient and...A major impediment in the development of chitosan nanoparticles （CTS NPs） as effective drug delivery vesicles is their rapid clearance from blood and endosome entrapment. To overcome these problems, a convenient and promising template system was developed by decorating poly（methacrylic acid） （PMAA） to the surface of 10-hydroxy camptothecin （HCPT）-loaded CTS NPs （HCPT-CTS/ PMAA NPs）. The results show that the presence of negatively charged PMAA significantly elongated the blood circulation time of HCPT-CTS NPs from 12 to 24 h, and reduced the blood clearance （C1） from 30.57 to 6.72 mL/h in vivo. The calculated area under curve （AUC0-24h） and terminal elimination half-life （tl/2） of HCPT-CTS/PMAA NPs were 4.37-fold and 2.48-fold compared with those of HCPT-CTS NPs. Furthermore, the positively charged HCPT-CTS/PMAA NPs triggered by tumor acidic microenvironment （pH 6.5） result in a 453-fold higher cellular uptake than the negatively charged counterparts at pH 7.4. Additionally, HCPT-CTS/PMAA NPs have the ability to escape endosomal entrapment via ＂proton sponge effect＂ after incubation with HepG2 cells for 3 h at pH 6.5. Taken together, these findings open up a convenient, low-cost, but effective way to prepare HCPT-CTS/PMAA NPs as a candidate for developing vectors with enhanced long blood circulation and endosomal escape ability in future clinical experiments.展开更多
Endosomes are crucial sites for intracellular material sorting and transportation.Endosomal transport is a critical process involved in the selective uptake,processing,and intracellular transport of substances.The equ...Endosomes are crucial sites for intracellular material sorting and transportation.Endosomal transport is a critical process involved in the selective uptake,processing,and intracellular transport of substances.The equilibrium between endocytosis and circulation mediated by the endosome-centered transport pathway plays a significant role in cell homeostasis,signal transduction,and immune response.In recent years,there have been hints linking endosomal transport abnormalities to neurodegenerative diseases,including Alzheimer’s disease.Nonetheless,the related mechanisms remain unclear.Here,we provide an overview of endosomal-centered transport pathways and highlight potential physiological processes regulated by these pathways,with a particular focus on the correlation of endosomal trafficking disorders with common pathological features of neurodegenerative diseases.Additionally,we summarize potential therapeutic agents targeting endosomal trafficking for the treatment of neurodegenerative diseases.展开更多
Intracellular delivery of biologicals such as peptides,proteins,and nucleic acids presents a great opportunity for innovative therapeutics.However,the endosome entrapment remains a major bottleneck in the intracellula...Intracellular delivery of biologicals such as peptides,proteins,and nucleic acids presents a great opportunity for innovative therapeutics.However,the endosome entrapment remains a major bottleneck in the intracellular delivery of biomacromolecules,largely limiting their therapeutic potential.Here,we converted a cell-penetrating peptide(CPP),low molecular weight protamine(LMWP),to endosomal escape peptides(EEPs)by masking LMWP with a pH-responsive counter-ionic peptide.The resulting masked CPPs(mLMWP and mLMWP2)effectively promoted the escape of peptide/protein cargoes from endosomes into the cytoplasm.Consequential lysosome repair and lysophagy were initiated upon the endolysosomal leakage.Minimal reactive oxygen species(ROS)elevation or cell death was observed.Based on mLMWP2,we constructed an intracellular protein delivery system containing an antibody as a targeting module,mLMWP2 as an endosomal escape module,and the desired protein cargo.With the HER2-targeting delivery system,we efficiently translocated cyclization recombination enzyme(Cre)and BH3-interacting domain death agonist(BID)into the cytosol of HER2^(+)cells to exert their biological activity.Thereby,the modular delivery system shows its potential as a promising tool for scientific studies and therapeutic applications.展开更多
Prevacuolar compartments （PVCs） and endosomal compartments are membrane-bound organelles mediating protein traffic to vacuoles in the secretory and endocytic pathways of plant cells. Over the years, great progress h...Prevacuolar compartments （PVCs） and endosomal compartments are membrane-bound organelles mediating protein traffic to vacuoles in the secretory and endocytic pathways of plant cells. Over the years, great progress has been made towards our understanding in these two compartments in plant cells. In this review, we will summarize our contributions toward the identification and characterization of plant prevacuolar and endosomal compartments. Our studies will serve as important steps in future molecular characterization of PVC biogenesis and PVC-mediated protein traffickinq in plant cells.展开更多
Endosomal compartments sort and deliver exogenous lipoprotein-derived cholesterol to the endoplasmic reticulum for regulating cellular cholesterol homeostasis.A large number of studies have focused on the removal of e...Endosomal compartments sort and deliver exogenous lipoprotein-derived cholesterol to the endoplasmic reticulum for regulating cellular cholesterol homeostasis.A large number of studies have focused on the removal of endosomal cholesterol,since its accumulation leads to devastating human diseases.Recent studies suggest that cytoplasmic sterol-binding proteins may be involved in endosomal cholesterol transport.In particular,endosome/lysosome-localized or-associated cholesterol-binding proteins may serve as key mediators of cholesterol removal in a non-vesicular manner.Further characterization of these cholesterol-binding proteins will shed light on the molecular mechanisms that regulate endosomal cholesterol sorting.展开更多
Attaching DNA/RNA to nanomaterials is the basis for nucleic acid-based assembly and drug delivery.Herein,we report that small interfering RNA(siRNA)effectively coordinates with ligand-free lanthanide nanoparticles(NaG...Attaching DNA/RNA to nanomaterials is the basis for nucleic acid-based assembly and drug delivery.Herein,we report that small interfering RNA(siRNA)effectively coordinates with ligand-free lanthanide nanoparticles(NaGdF4 NPs),and forms siRNA/NaGdF4 spherical nucleic acids(SNA).The coordination is primarily attributed to the interaction between Gd and phosphate backbone of the siRNA.Surprisingly,an efficient encapsulation and rapid endosomal escape of siRNA from the endosome/lysosome were achieved,due to its flexible ability to bound to phospholipid head of endosomal membrane,thereby disrupting the membrane structure.Resorting to the dual properties of NaGdF4 NPs,siRNA loading,and endosomal escape,siRNA targeting programmed cell death-ligand 1(siPD-L1)/NaGdF4 SNA triggers significant gene silencing in vitro and in vivo,and effectively represses the tumor growth in both CT26 tumor model and 4T1 orthotopic murine model.展开更多
基金supported by National Natural Science Foundation of China(No.82173769)the National Key R&D Program of China(No.2021YFE0106900)+1 种基金Applied Basic Research Multiinvestment Foundation of Tianjin(No.21JCYBJC01540)the Science&Technology Development Fund of Tianjin Education Commission for Higher Education(No.2023ZD019)。
文摘Small interfering RNAs(siRNA)provide a novel and highly specific therapy due to their ability to effectively silence target genes,to date six siRNA therapeutics are approved for clinical use.Even so,some critical challenges remain to overcome in the therapeutic application of siRNAs,with delivery issues at the forefront.Among them,endo/lysosomal barrier is one of the important but often-neglected limitations hindering the delivery of siRNA therapeutics.In this review,we summarize the promising strategies that facilitate siRNAs overcoming endo/lysosomal barriers based on the cellular uptake and intracellular transport pathways,including promoting escape once endocytosis into the endo/lysosomes and bypassing lysosomes via endosome-Golgi-endoplasmic reticulum(ER)pathway or nonendocytosis pathway,and discuss the principal considerations and the future directions of promoting endo/lysosomal escape in the development of therapeutic siRNAs.
基金supported by National Natural Science Foundation of China(81502688)Cooperation Research Funding of Capital Medical University(2020KJ000514)+1 种基金Cooperation Research Funding of Capital Medical University(2023KJ000814)R&D Program of Beijing Municipal Education Commission(KM202210025024).
文摘Efficient siRNA delivery is highly desirable for disease treatment.However,the application of conventional nanoparticles is limited by the inability to escape from endo-lysosomes.Herein,we report a strategy using small-molecule drugs to enhance siRNA endo-lysosomal release,addressing this challenge.We encapsulated gentamicin(GM)into the marketed Onpattro■ formulation to establish LNP-siRNA/GM nanoparticles that promote siRNA endo-lysosomal escape through endosomal disruption,mechanistically exhibiting unique functionality and synergistic effects of LNP-siRNA/GM to improve cancer therapy.Besides,GM induced reactive oxygen species(ROS)and phospholipids accumulation in endolysosomes,as well as the physical characteristics of lipid nanoparticles(LNPs)were preserved.We also revealed that GM causes endo-lysosomal swelling and disrupts the endosomal membrane to enable siRNA release,as confirmed by Galectin 3 recruitment and acridine orange release.This approach achieved∼81%mRNA-EGFR silencing,which is more than LNP-siEGFR(∼56.23%)by enhancing siRNA endo-lysosomal escape efficiency.Meanwhile,LNP-siEGFR/GM exhibited significant biological activities in HepG2 cells,driven by the synergistic effects of siEGFR and GM with the VEGF and CXCL12 downregulation of,and ROS and phospholipids upregulation.Furthermore,tumor growth was notably suppressed after intravenous injection of LNP-siEGFR/GM in tumor-bearing nude mice.The combination of EGFR-siRNA and GM could also greatly inhibit angiogenesis,be antiproliferative,and induce tumor cells apoptosis.Therefore,this GM and siRNA co-delivery system would provide an efficient strategy for siRNA endosomal escape,significantly improving knockdown in various LNPs based siRNA delivery systems and efficiently enhancing cancer therapy.
基金This work was supported in part by the National Science Fund for Distinguished Young Scholars (No. 30525007)National Basic Research Program of China (No. 2006CB504103+1 种基金 No. 2006CB500704)Hi-Tech Research and Development Program of China (No. 2006AA02Z322)
文摘With the identification of more than a dozen novel Hermansky-Pudlak Syndrome （HPS） proteins in vesicle trafficking in higher eukaryotes, a new class of trafficking pathways has been described. It mainly consists of three newly-defined protein com- plexes, BLOC-l, -2, and -3. Compelling evidence indicates that these complexes together with two other well-known complexes, AP3 and HOPS, play important roles in endosomal transport. The interactions between these complexes form a network in protein trafficking via endosomes and cytoskeleton. Each node of this network has intra-complex and extra-complex interactions. These complexes are connected by direct interactions between the subunits from different complexes or by indirect interactions through coupling nodes that interact with two or more subunits from different complexes. The dissection of this network facilitates the understanding of a dynamic but elaborate transport machinery in protein/membrane trafficking. The disruption of this network may lead to abnormal trafficking or defective organellar development as described in patients with Hermansky-Pudlak syndrome.
基金We greatly appreciate Prof.Charles Wood(University of Nebraska,Lincoln,USA)for the gift of the infectious molecular clones(pNL4.3,pNL4.3ΔEnvEGFP,and pVSV-G).the National Natural Science Foundation of China(81571987)Natural Science Foundation of Tianjin Municipal Science and Technology Commission(20JCQNJC01750,21JCQNJC01600).
文摘Human immunodeficiency virus-1(HIV-1)encodes simply 15 proteins and thus depends on multiple host cellular factors for virus reproduction.Spastin,a microtubule severing protein,is an identified HIV-1 dependency factor,but the mechanism regulating HIV-1 is unclear.Here,the study showed that knockdown of spastin inhibited the production of the intracellular HIV-1 Gag protein and new virions through enhancing Gag lysosomal degradation.Further investigation showed that increased sodium tolerance 1(IST1),the subunit of endosomal sorting complex required for transport(ESCRT),could interact with the MIT domain of spastin to regulate the intracellular Gag production.In summary,spastin is required for HIV-1 replication,while spastin-IST1 interaction facilitates virus production by regulating HIV-1 Gag intracellular trafficking and degradation.Spastin may serve as new target for HIV-1 prophylactic and therapy.
基金supported by the Deutsche Forschungsgemeinschaft(RTG1949/2)the National Natural Science Foundation of China(82202497).
文摘Hepatitis B virus(HBV)produces and releases various particle types,including complete virions,subviral particles with envelope proteins,and naked capsids.Recent studies demonstrate that HBV exploits distinct intracellular membrane trafficking pathways,including the endosomal vesicle trafficking and autophagy pathway,to assemble and release viral and subviral particles.Herein,we summarize the findings about the distinct roles of autophagy and endosomal membrane trafficking and the interaction of both pathways in HBV replication,assembly,and release.
基金supported by the National Key Research and Development Program of China(Grant No.216YFD0500402)Natural Science Foundation of China(Grants No.31272579 and 31472208)
文摘Rabies virus(RABV) is a highly neurotropic virus that follows clathrin-mediated endocytosis and p H-dependent pathway for trafficking and invasion into endothelial cells. Early(Rab5, EEA1) and late(Rab7, LAMP1) endosomal proteins play critical roles in endosomal sorting, maturity and targeting various molecular cargoes, but their precise functions in the early stage of RABV neuronal infection remain elusive. In this study, the relationship between enigmatic entry of RABV with these endosomal proteins into neuronal and SH-SY5 Y cells was investigated.Immunofluorescence, TCID_(50) titers, electron microscopy and western blotting were carried out to determine the molecular interaction of the nucleoprotein(N) of RABV with early or late endosomal proteins in these cell lines. The expression of N was also determined by down-regulating Rab5 and Rab7 in both cell lines through RNA interference. The results were indicative that N proficiently colocalized with Rab5/EEA1 and Rab7/LAMP1 in both cell lines at 24 and 48 h post-infection, while N titers significantly decreased in early infection of RABV. Down-regulation of Rab5 and Rab7 did not inhibit N expression, but it prevented productive infection via blocking the normal trafficking of RABV in a low pH environment. Ultrathin sections of cells studied by electron microscope also verified the close association of RABV with Rab5 and Rab7 in neurons. From the data it was concluded that primary entry of RABV strongly correlates with the kinetics of Rab-proteins present on early and late vesicles, which provides helpful clues to explain the early events of RABV in nerve cells.
基金grants from the National Natural Science Foundation of China(Grant No.81530084,81874200,and 81572750)the Hunan Science and Technology Department(Grant No.2018RS3028)+1 种基金Central South University(Grant No.20170033010007)The Strategy-Orientated Special Project of Central South University(Grant No.ZLXD2017003).
文摘Objectives:Improper activation of Wnt/β-catenin signaling has been implicated in human diseases.Beyond the well-studied glycogen synthase kinase 3p(GSK3p)and casein kinase 1(CK1),other kinases affecting Wnt/β-catenin signaling remain to be defined.Methods:To identify the kinases that modulate Wnt/β-catenin signaling,we applied a kinase small interfering RNA(siRNA)library screen approach.Luciferase assays,immunoblotting,and real-time polymerase chain reaction(PCR)were performed to confirm the regulation o f the Wnt/β-catenin signaling pathway by cyclin-dependent kinase 11(CDK11)and to investigate the underlying mechanism.Confocal immunofluorescence,coimmunoprecipitation(co-IP),and scratch wound assays were used to demonstrate colocalization,detect protein interactions,and explore the function of CDK11.Results:CDK11 was found to be a significant candidate kinase participating in the negative control of Wnt/P-catenin signaling.Down-regulation of CDK11 led to the accumulation of Wnt/β-catenin signaling receptor complexes,in a manner dependent on intact adenomatosis polyposis coli(APC)protein.Further analysis showed that CDK11 modulation of Wnt/P-catenin signaling engaged the endolysosomal machinery,and CDK11 knockdown enhanced the colocalization of Wnt/β-catenin signaling receptor complexes with early endosomes and decreased colocalization with lysosomes.Mechanistically,CDK11 was found to function in Wnt/β-catenin signaling by regulating microtubule stability.Depletion of CDK11 down-regulated acetyl-a-tubulin.Moreover,co-IP assays demonstrated that CDK11 interacts with the a-tubulin deacetylase SIRT2,whereas SIRT2 down-regulation in CDK11-depleted cells reversed the accumulation of Wnt/(3-catenin signaling receptor complexes.CDK11 was found to suppress cell migration through altered W nt/β-catenin signaling.Conclusions:CDK11 is a negative modulator of Wnt/β-catenin signaling that stabilizes microtubules,thus resulting in the dysregulation of receptor complex trafficking from early endosomes to lysosomes.
基金supported by the National Natural Science Foundation of China (No. 21004062)"100 Talents Program" of the Chinese Academy of Sciences (No. KGCX2-YW-802)the Ministry of Science and Technology ofChina ("973 Project", No. 2009CB930102)
文摘To further enhance the transfection efficiency of a micelleplex system based on monomethoxy poly（ethylene glycol）-block-poly（e-caprolactone）-block-poly（L-lysine） （MPEG-b-PCL-b-PLL）, cholesterol （Chol） moieties are attached to the e-termini of PLL segments to obtain MPEG-b-PCL-b-PLL/Chol. The structure and morphology of the copolymer are studied by IH-NMR, TEM and DLS （dynamic light scattering）. The cytotoxicity, cell uptake, endosomal release and mRNA knockdown are studied by MTT （3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide） assay, flow cytometry, CLSM （confocal laser scanning microscopy） and RT-PCR （real-time polymerase chain reaction）. The results show that compared to their precursor MPEG-b-PCL-b-PLL, the cholesterol-grafted copolymer shows significantly lower toxicity, more rapid cellular endocytosis and endosome escape, and consequently displays enhanced siRNA transfection efficiency even at a lower N/P ratio. These improvements are ascribed to enhanced interaction of the cholesterol moieties with both cellular membrane and endosomal membrane. Moreover, effect of the PLL block length is examined. The final conclusion is that long enough PLL segments and incorporation of proper fraction of cholesterol onto the PLL segments benefit the enhancement of siRNA transfection efficiency.
基金The National Key R&D Program of China(2021YFA1300302)the National Natural Science Foundation of China(32320103007,32130027)+1 种基金the National Science Fund for Distinguished Young Scholars(31825017)the Major Research Plan of the Natural Science Foundation of China(92154001)to A.Shi.
文摘Endosomes are characterized by the presence of various phosphoinositides that are essential for defining the membrane properties.However,the interplay between endosomal phosphoinositides metabolism and innate immunity is yet to be fully understood.Here,our findings highlight the evolutionary continuity of RAB-10/Rab10’s involvement in regulating innate immunity.Upon infection of Caenorhabditis elegans with Pseudomonas aeruginosa,an increase in RAB-10 activity was observed in the intestine.Conversely,when RAB-10 was absent,the intestinal diacylglycerols(DAGs)decreased,and the animal’s response to the pathogen was impaired.Further research revealed that UNC-16/JIP3 acts as an RAB-10 effector,facilitating the recruitment of phospholipase EGL-8 to endosomes.This leads to a decrease in endosomal phosphatidylinositol 4,5-bisphosphate(PI(4,5)P2)and an elevation of DAGs,as well as the activation of the PMK-1/p38 MAPK innate immune pathway.It is noteworthy that the dimerization of UNC-16 is a prerequisite for its interaction with RAB-10(GTP)and the recruitment of EGL-8.Moreover,we ascertained that the rise in RAB-10 activity,due to infection,was attributed to the augmented expression of LET-413/Erbin,and the nuclear receptor NHR-25/NR5A1/2 was determined to be indispensable for this increase.Hence,this study illuminates the significance of endosomal PI(4,5)P2 catabolism in boosting innate immunity and outlines an NHR-25-mediated mechanism for pathogen detection in intestinal epithelia.
文摘Recent discovery of PYR/PYL/RCAR-type abscisic acid （ABA） receptors has become one of most significant advances in plant science in the past decade. In mammals, endosomal sorting acts as an important pathway to downregulate different types of receptors, but its role in plant hormone signaling is poorly understood. Here, we report that an ubiquitin E2-1ike protein, VPS23A, which is a key component of ESCRT-I, negatively regulates ABA signaling. VPS23A has epistatic relationship with PYR/PYL/RCAR-type ABA receptors and disruption of VPS23A enhanced the activity of key kinase OST1 in the ABA signaling pathway under ABA treatment. Moreover, VPS23A interacts with PYR1/PYLs and K63-1inked diubiquitin, and PYL4 possesses K63-1inked ubiquitinated modification in vivo. Further analysis revealed that VPS23A affects the subcellular localization of PYR 1 and the stability of PYL4. Taken together, our results suggest that VPS23A affects PYR1/ PYL4 via vacuole-mediated degradation, providing an advanced understanding of both the turnover of ABA receptors and ESCRTs in plant hormone signaling.
基金supported by a grant from the National BasicResearch Program of China (973 Program, 2011CB915402)the National Science Foundation of China (CNSF31030047/90717006)
文摘The 76 amino acid protein ubiquitin （Ub） is highly conserved in all eukaryotic species. It plays important roles in many cellular processes by covalently attaching to the target proteins. The best known function of Ub is marking substrate proteins for degra- dation by the 26S proteasome. In fact, other consequences of ubiquitination have been discovered in yeast and mammals, such as membrane trafficking, DNA repair, chromatin modification, and protein kinase activation. The common mechanism underlying these processes is that Ub serves as a signal to sort proteins to the vacuoles or lysosomes for degradation as opposed to 26S proteasome-dependent degradation. To date, several reports have indicated that a similar function of Ub also exists in plants. This review focuses on a summary and analysis of the recent research progress on Ub acting as a signal to mediate endocytosis and endosomal trafficking in plants.
文摘There are many challenges in developing efficient and target specific delivery systems of small molecule and nucleic acid drugs. Cell membrane presents one of the major barriers for the penetration of hydrophilic macromolecules across the plasma membrane. Nanocar- riers have been designed to enhance their cellular uptake via endocytosis but following their cellular uptake, endosomal escape is the rate limiting step which restricts the value associated with the enhanced uptake by nanocarriers. Viruses are an excellent model for efficient cytosolic delivery by nanocarriers. Viruses exploit intra- cellular cues to release the genome to cytosol. In this review, we first discuss different endocytic uptake path- ways and endosomal escape mechanisms. We then summarize the existing tools for studying the intracellular trafficking of nanocarriers. Finally, we highlight the important design elements of recent virus-based nanocar- tiers for efficient cellular uptake and endosomal escape.
基金the National Natural Science Foundation of China(Nos.81771968,21704061,and 82003166)Natural Science Foundation of Shanghai(No.21ZR1439200)+3 种基金Shanghai Sailing Program(No.17YF1411000)Shanghai Municipal Education Commission-Gaofeng Clinical Grant Support(No.20181705)Shanghai Municipal Commission of Health and Family Planning(No.201840020)the Medical-Engineering Joint Funds from the Shanghai Jiao Tong University(Nos.ZH2018ZDA05 and YG2016QN54).
文摘Dendrimer,such as dendrigraft poly-L-lysine(DGL)polymers,with high surface charge density,well-defined structure,and narrow poly-dispersity is often employed as a gene vector,but its transfection efficiency is still partially inhibited due to poor endosomal escape ability.Herein,we used a surface modification strategy to enhance the endosomal escape ability of DGL polymers,and thus improved its gene transfection efficiency.A library of phenylboronic acid(PBA)modified DGL polymers(PBA-DGLs)was designed to screen efficient small interfering RNA(siRNA)vectors.The lead candidate screened from the library shown a capability of inducing nearly 90% gene silencing in MDA-MB-231 cells.The study of the transfection mechanism revealed that PBA modification not only improves siRNA cellular uptake,but,more importantly,endows DGL polymers the ability of endosomal escape.One of the top candidates from polyplexes was further shielded with hyaluronic acid to construct targeted nanoparticles,and the yielding nanoparticles significantly suppressed the tumor growth in a breast cancer model by effective siRNA delivery.This research provides a general and effective strategy to enhance the endosomal escape and transfection efficiency of dendrimer.
基金Agencia Estatal de Investigación (AEI) and to Fondo Europeo de Desarrollo Regional (FEDER) (BIO2016-76063-R, AEI/FEDER, UE) to Villaverde A, AGAUR (2017SGR-229) to Villaverde A and 2017SGR-865 GRCISCⅢ (PI15/00272 co-founding FEDER) to Vázquez E and ISCⅢ (Co-founding FEDER) PIE15//00028 and PI18/00650 to Mangues R, and to EU COST Action CA 17140+3 种基金funded by the Ⅵ National R&D&I Plan 2008–2011, Iniciativa Ingenio 2010, Consolider Program, CIBER Actionsfinanced by the Instituto de Salud Carlos Ⅲ, with assistance from the European Regional Development Fundsupported by a predoctoral fellowship from AGAUR (2019 FI_B 00352)PERIS program from the Health Department of the Generalitat de Catalunya
文摘Poly-histidine peptides such as H6(HHHHHH)are used in protein biotechnologies as purification tags,protein-assembling agents and endosomal-escape entities.The pleiotropic properties of such peptides make them appealing to design protein-based smart materials or nanoparticles for imaging or drug delivery to be produced in form of recombinant proteins.However,the clinical applicability of H6-tagged proteins is restricted by the potential immunogenicity of these segments.In this study,we have explored several humanized histidine-rich peptides in tumor-targeted modular proteins,which can specifically bind and be internalized by the target cells through the tumoral marker CXCR4.We were particularly interested in exploring how protein purification,self-assembling and endosomal escape perform in proteins containing the variant histidine-rich tags.Among the tested candidates,the peptide H5 E(HEHEHEHEH)is promising as a good promoter of endosomal escape of the associated fulllength protein upon endosomal internalization.The numerical modelling of cell penetration and endosomal escape of the tested proteins has revealed a negative relationship between the amount of protein internalized into target cells and the efficiency of cytoplasmic release.This fact demonstrates that the His-mediated,proton sponge-based endosomal escape saturates at moderate amounts of internalized protein,a fact that might be critical for the design of protein materials for cytosolic molecular delivery.
文摘A major impediment in the development of chitosan nanoparticles （CTS NPs） as effective drug delivery vesicles is their rapid clearance from blood and endosome entrapment. To overcome these problems, a convenient and promising template system was developed by decorating poly（methacrylic acid） （PMAA） to the surface of 10-hydroxy camptothecin （HCPT）-loaded CTS NPs （HCPT-CTS/ PMAA NPs）. The results show that the presence of negatively charged PMAA significantly elongated the blood circulation time of HCPT-CTS NPs from 12 to 24 h, and reduced the blood clearance （C1） from 30.57 to 6.72 mL/h in vivo. The calculated area under curve （AUC0-24h） and terminal elimination half-life （tl/2） of HCPT-CTS/PMAA NPs were 4.37-fold and 2.48-fold compared with those of HCPT-CTS NPs. Furthermore, the positively charged HCPT-CTS/PMAA NPs triggered by tumor acidic microenvironment （pH 6.5） result in a 453-fold higher cellular uptake than the negatively charged counterparts at pH 7.4. Additionally, HCPT-CTS/PMAA NPs have the ability to escape endosomal entrapment via ＂proton sponge effect＂ after incubation with HepG2 cells for 3 h at pH 6.5. Taken together, these findings open up a convenient, low-cost, but effective way to prepare HCPT-CTS/PMAA NPs as a candidate for developing vectors with enhanced long blood circulation and endosomal escape ability in future clinical experiments.
基金supported by the National Natural Science Foundation of China(81901309)the Science and Technology Funding Project to Support the High-Quality Development of China Medical University(2023JH2/20200039)+1 种基金the Basic Scientific Research Project of Institutions of Higher Learning of Liaoning Province(LJKZ0775)the Joint Plan of Science and Technology of Liaoning Province(2023-MSLH-369)the Science and Technology Innovation Team Project of China Medical University(CXTD2022007).
文摘Endosomes are crucial sites for intracellular material sorting and transportation.Endosomal transport is a critical process involved in the selective uptake,processing,and intracellular transport of substances.The equilibrium between endocytosis and circulation mediated by the endosome-centered transport pathway plays a significant role in cell homeostasis,signal transduction,and immune response.In recent years,there have been hints linking endosomal transport abnormalities to neurodegenerative diseases,including Alzheimer’s disease.Nonetheless,the related mechanisms remain unclear.Here,we provide an overview of endosomal-centered transport pathways and highlight potential physiological processes regulated by these pathways,with a particular focus on the correlation of endosomal trafficking disorders with common pathological features of neurodegenerative diseases.Additionally,we summarize potential therapeutic agents targeting endosomal trafficking for the treatment of neurodegenerative diseases.
基金supported by supported by Beijing Municipal Science&Technology Commission(Z231100007223008,China)the National Key R&D Program of China(2017YFA0207900,China)+1 种基金Tsinghua University Initiative Scientific Research Program(2023Z11DSZ001,China)the Tsinghua-Peking Joint Center for Life Sciences.
文摘Intracellular delivery of biologicals such as peptides,proteins,and nucleic acids presents a great opportunity for innovative therapeutics.However,the endosome entrapment remains a major bottleneck in the intracellular delivery of biomacromolecules,largely limiting their therapeutic potential.Here,we converted a cell-penetrating peptide(CPP),low molecular weight protamine(LMWP),to endosomal escape peptides(EEPs)by masking LMWP with a pH-responsive counter-ionic peptide.The resulting masked CPPs(mLMWP and mLMWP2)effectively promoted the escape of peptide/protein cargoes from endosomes into the cytoplasm.Consequential lysosome repair and lysophagy were initiated upon the endolysosomal leakage.Minimal reactive oxygen species(ROS)elevation or cell death was observed.Based on mLMWP2,we constructed an intracellular protein delivery system containing an antibody as a targeting module,mLMWP2 as an endosomal escape module,and the desired protein cargo.With the HER2-targeting delivery system,we efficiently translocated cyclization recombination enzyme(Cre)and BH3-interacting domain death agonist(BID)into the cytosol of HER2^(+)cells to exert their biological activity.Thereby,the modular delivery system shows its potential as a promising tool for scientific studies and therapeutic applications.
基金Supported by grants from the Research Grants Council of Hong Kong(CUHK4156/01M,CUHK4260/02M,CUHK4307/03M,and CUHK4580/05M)National Science Foundation of China (30529001)+1 种基金CUHK Scheme C,UGCAoE(B-07/99)Germany/HK Joint Research Scheme to L.Jiang.
文摘Prevacuolar compartments （PVCs） and endosomal compartments are membrane-bound organelles mediating protein traffic to vacuoles in the secretory and endocytic pathways of plant cells. Over the years, great progress has been made towards our understanding in these two compartments in plant cells. In this review, we will summarize our contributions toward the identification and characterization of plant prevacuolar and endosomal compartments. Our studies will serve as important steps in future molecular characterization of PVC biogenesis and PVC-mediated protein traffickinq in plant cells.
基金supported by research grants from the Ara Parseghian Medical Research Foundationthe National Health and Medical Research Council of Australia(#510271).
文摘Endosomal compartments sort and deliver exogenous lipoprotein-derived cholesterol to the endoplasmic reticulum for regulating cellular cholesterol homeostasis.A large number of studies have focused on the removal of endosomal cholesterol,since its accumulation leads to devastating human diseases.Recent studies suggest that cytoplasmic sterol-binding proteins may be involved in endosomal cholesterol transport.In particular,endosome/lysosome-localized or-associated cholesterol-binding proteins may serve as key mediators of cholesterol removal in a non-vesicular manner.Further characterization of these cholesterol-binding proteins will shed light on the molecular mechanisms that regulate endosomal cholesterol sorting.
基金supported by the Beijing Nova Program from Beijing Municipal Science&Technology Commission(No.Z201100006820005)the Beijing-Tianjin-Hebei Basic Research Cooperation Project(No.19JCZDJC64100)+2 种基金the National Key Research&Development Program of China(Nos.2018YFE0117800,2021YFA1201000,and 2021YFE0106900)the National Natural Science Foundation of China(Nos.32030060 and 31871003)the Natural Science Foundation of China international collaboration key project(No.51861135103).
文摘Attaching DNA/RNA to nanomaterials is the basis for nucleic acid-based assembly and drug delivery.Herein,we report that small interfering RNA(siRNA)effectively coordinates with ligand-free lanthanide nanoparticles(NaGdF4 NPs),and forms siRNA/NaGdF4 spherical nucleic acids(SNA).The coordination is primarily attributed to the interaction between Gd and phosphate backbone of the siRNA.Surprisingly,an efficient encapsulation and rapid endosomal escape of siRNA from the endosome/lysosome were achieved,due to its flexible ability to bound to phospholipid head of endosomal membrane,thereby disrupting the membrane structure.Resorting to the dual properties of NaGdF4 NPs,siRNA loading,and endosomal escape,siRNA targeting programmed cell death-ligand 1(siPD-L1)/NaGdF4 SNA triggers significant gene silencing in vitro and in vivo,and effectively represses the tumor growth in both CT26 tumor model and 4T1 orthotopic murine model.
