RESUMO
Integrins are validated drug targets with six approved therapeutics. However, small-molecule inhibitors to three integrins failed in late-stage clinical trials for chronic indications. Such unfavorable outcomes may in part be caused by partial agonism, i.e., the stabilization of the high-affinity, extended-open integrin conformation. Here, we show that the failed, small-molecule inhibitors of integrins αIIbß3 and α4ß1 stabilize the high-affinity conformation. Furthermore, we discovered a simple chemical feature present in multiple αIIbß3 antagonists that stabilizes integrins in their bent-closed conformation. Closing inhibitors contain a polar nitrogen atom that stabilizes, via hydrogen bonds, a water molecule that intervenes between a serine residue and the metal in the metal-ion-dependent adhesion site (MIDAS). Expulsion of this water is a requisite for transition to the open conformation. This change in metal coordination is general to integrins, suggesting broad applicability of the drug-design principle to the integrin family, as validated with a distantly related integrin, α4ß1.
Assuntos
Desenho de Fármacos , Integrina alfa4beta1 , Conformação Proteica , Serina , ÁguaRESUMO
Advances in neuroendocrinology have led to major discoveries since the 19th century, identifying adaptive loops for maintaining homeostasis. One of the most remarkable discoveries was the concept of neurosteroids, according to which the brain is not only a target but also a source of steroid production. The identification of new membrane steroid targets now underpins the neuromodulatory effects of neurosteroids such as pregnenolone, which is involved in functions mediated by the GPCR CB1 receptor. Structural analysis of steroids is a key feature of their interactions with the phospholipid membrane, receptors and resulting activity. Therefore, mass spectrometry-based methods have been developed to elucidate the metabolic pathways of steroids, the ultimate approach being metabolomics, which allows the identification of a large number of metabolites in a single sample. This approach should enable us to make progress in understanding the role of neurosteroids in the functioning of physiological and pathological processes.
Assuntos
Neuroesteroides , Neuroesteroides/metabolismo , Pregnenolona/metabolismo , Esteroides , Encéfalo/metabolismoRESUMO
SignificanceEpidermal growth factor receptor (EGFR) is one of the most important membrane receptors that transduce growth signals into cells to sustain cell growth, proliferation, and survival. EGFR signal termination is initiated by EGFR internalization, followed by trafficking through endosomes, and degradation in lysosomes. How this process is regulated is still poorly understood. Here, we show that hepatocyte growth factor regulated tyrosine kinase substrate (HGS), a key protein in the EGFR trafficking pathway, is dynamically modified by a single sugar N-acetylglucosamine. This modification inhibits EGFR trafficking from endosomes to lysosomes, leading to the accumulation of EGFR and prolonged signaling. This study provides an important insight into diseases with aberrant growth factor signaling, such as cancer, obesity, and diabetes.
Assuntos
Endossomos/metabolismo , Lisossomos/metabolismo , Transdução de Sinais , Acilação/genética , Endossomos/genética , Receptores ErbB/genética , Receptores ErbB/metabolismo , Células Hep G2 , Humanos , Lisossomos/genética , Transporte Proteico/genéticaRESUMO
On-demand engineering of cell membrane receptors to nongenetically intervene in cellular behaviors is still a challenge. Herein, a membraneless enzyme biofuel cell-based self-powered biosensor (EBFC-SPB) was developed for autonomously and precisely releasing Zn2+ to initiate DNAzyme-based reprogramming of cell membrane receptors, which further mediates signal transduction to regulate cellular behaviors. The critical component of EBFC-SPB is a hydrogel film on a biocathode which is prepared using a Fe3+-cross-linked alginate hydrogel film loaded with Zn2+ ions. In the working mode in the presence of glucose/O2, the hydrogel is decomposed due to the reduction of Fe3+ to Fe2+, accompanied by rapid release of Zn2+ to specifically activate a Zn2+-responsive DNAzyme nanodevice on the cell surface, leading to the dimerization of homologous or nonhomologous receptors to promote or inhibit cell proliferation and migration. This EBFC-SPB platform provides a powerful "sensing-actuating-treating" tool for chemically regulating cellular behaviors, which holds great promise in precision biomedicine.
Assuntos
Técnicas Biossensoriais , Zinco , Zinco/química , Zinco/metabolismo , Receptores de Superfície Celular/metabolismo , DNA Catalítico/metabolismo , DNA Catalítico/química , Humanos , Hidrogéis/química , Proliferação de Células/efeitos dos fármacos , Fontes de Energia Bioelétrica , Alginatos/química , Movimento Celular/efeitos dos fármacosRESUMO
This work demonstrates that targeting ligand density on nanoparticles can affect interactions between the nanoconstructs and cell membrane receptors. We discovered that when the separation between covalently grafted DNA aptamers on gold nanostars was comparable to the distance between binding sites on a receptor dimer (matched density; MD), nanoconstructs exhibited a higher selectivity for binding to the dimeric form of the protein. Single-particle dynamics of MD nanoconstructs showed slower rotational rates and larger translational footprints on cancer cells expressing more dimeric forms of receptors (dimer+) compared with cells having more monomeric forms (dimer-). In contrast, nanoconstructs with either increased (nonmatched density; NDlow) or decreased ligand spacing (NDhigh) had minimal changes in dynamics on either dimer+ or dimer- cells. Real-time, single-particle analyses can reveal the importance of nanoconstruct ligand density for the selective targeting of membrane receptors in live cells.
Assuntos
Nanopartículas , Ligantes , Membrana Celular/metabolismo , Nanopartículas/química , Polímeros/metabolismo , Sítios de LigaçãoRESUMO
Macrophages, as highly phenotypic plastic immune cells, play diverse roles in different pathological conditions. Changing and controlling the phenotypes of macrophages is considered a novel potential therapeutic intervention. Meanwhile, specific transmembrane proteins anchoring on the surface of the macrophage membrane are relatively conserved, supporting its functional properties, such as inflammatory chemotaxis and tumor targeting. Thus, a series of drug delivery systems related to specific macrophage membrane proteins are commonly used to treat chronic inflammatory diseases. This review summarizes macrophages-based strategies for chronic diseases, discusses the regulation of macrophage phenotypes and their polarization processes, and presents how to design and apply the site-specific targeted drug delivery systems in vivo based on the macrophages and their derived membrane receptors. It aims to provide a better understanding of macrophages in immunoregulation and proposes macrophages-based targeted therapeutic approaches for chronic diseases.
Assuntos
Sistemas de Liberação de Medicamentos , Neoplasias , Humanos , Fenótipo , Macrófagos/metabolismo , Neoplasias/tratamento farmacológico , Neoplasias/metabolismo , Doença CrônicaRESUMO
Cyclic nucleotide-gated (CNG) ion channels of olfactory neurons are tetrameric membrane receptors that are composed of two A2 subunits, one A4 subunit, and one B1b subunit. Each subunit carries a cyclic nucleotide-binding domain in the carboxyl terminus, and the channels are activated by the binding of cyclic nucleotides. The mechanism of cooperative channel activation is still elusive. Using a complete set of engineered concatenated olfactory CNG channels, with all combinations of disabled binding sites and fit analyses with systems of allosteric models, the thermodynamics of microscopic cooperativity for ligand binding was subunit- and state-specifically quantified. We show, for the closed channel, that preoccupation of each of the single subunits increases the affinity of each other subunit with a Gibbs free energy (ΔΔG) of â¼-3.5 to â¼-5.5 kJ â mol-1, depending on the subunit type, with the only exception that a preoccupied opposite A2 subunit has no effect on the other A2 subunit. Preoccupation of two neighbor subunits of a given subunit causes the maximum affinity increase with ΔΔG of â¼-9.6 to â¼-9.9 kJ â mol-1 Surprisingly, triple preoccupation leads to fewer negative ΔΔG values for a given subunit as compared to double preoccupation. Channel opening increases the affinity of all subunits. The equilibrium constants of closed-open isomerizations systematically increase with progressive liganding. This work demonstrates, on the example of the heterotetrameric olfactory CNG channel, a strategy to derive detailed insights into the specific mutual control of the individual subunits in a multisubunit membrane receptor.
Assuntos
AMP Cíclico/metabolismo , GMP Cíclico/metabolismo , Canais de Cátion Regulados por Nucleotídeos Cíclicos/química , Canais de Cátion Regulados por Nucleotídeos Cíclicos/metabolismo , Ativação do Canal Iônico , Termodinâmica , Animais , Sítios de Ligação , Canais de Cátion Regulados por Nucleotídeos Cíclicos/genética , Ligantes , Oócitos/metabolismo , Conformação Proteica , Subunidades Proteicas , Xenopus laevis/crescimento & desenvolvimento , Xenopus laevis/metabolismoRESUMO
BACKGROUND: Labeling efficiency is a crucial parameter in fluorescence applications, especially when studying biomolecular interactions. Current approaches for estimating the yield of fluorescent labeling have critical drawbacks that usually lead them to be inaccurate or not quantitative. RESULTS: We present a method to quantify fluorescent-labeling efficiency that addresses the critical issues marring existing approaches. The method operates in the same conditions of the target experiments by exploiting a ratiometric evaluation with two fluorophores used in sequential reactions. We show the ability of the protocol to extract reliable quantification for different fluorescent probes, reagents concentrations, and reaction timing and to optimize labeling performance. As paradigm, we consider the labeling of the membrane-receptor TrkA through 4'-phosphopantetheinyl transferase Sfp in living cells, visualizing the results by TIRF microscopy. This investigation allows us to find conditions for demanding single and multi-color single-molecule studies requiring high degrees of labeling. CONCLUSIONS: The developed method allows the quantitative determination and the optimization of staining efficiency in any labeling strategy based on stable reactions.
Assuntos
Técnicas de Cultura de Células , Corantes Fluorescentes , Microscopia , Coloração e RotulagemRESUMO
Antiangiogenic therapies are considered a promising strategy against solid tumors. Their aim is to inhibit the formation of new blood vasculature, thereby reducing the oxygen and nutrient supply to prevent further tumor growth and spreading. However, the strategy has seen limitations, as survival benefits are modest and often accompanied with increased tumor aggressiveness in form of invasion and metastasis. Antiangiogenic induced changes in the tumor microenvironment, such as hypoxia, mechanical stress or extracellular acidification can activate different receptors of tumoral and stromal cells and induce an extensive remodeling of the entire tumor microenvironment, with the overall goal to invade nearby tissues and regain access to the vasculature. In this regard, receptor tyrosine kinases have been studied intensively and especially the inhibition of c-Met has given promising results, characterized by a reduction in invasiveness and prolonged survival. Receptors that sense changes in the extracellular matrix like integrins or proteoglycans can also induce downstream signaling that stimulates the expression of remodeling factors such as new matrix components, enzymes or chemoattractants. Targeting multiple receptors and sensors of cancer cells simultaneously might represent an effective second line treatment that prevents the formation of malignant side effects.
Assuntos
Neoplasias , Microambiente Tumoral , Humanos , Microambiente Tumoral/fisiologia , Inibidores da Angiogênese/farmacologia , Inibidores da Angiogênese/uso terapêutico , Matriz Extracelular/metabolismo , Neoplasias/tratamento farmacológico , Neoplasias/etiologia , Neoplasias/metabolismo , Transdução de SinaisRESUMO
A surface-enhanced Raman scattering (SERS)/fluorescence dual-mode nanoprobe was proposed to assess anti-diabetic drug actions from the expression level of the epidermal growth factor receptor (EGFR), which is a significant biomarker of breast cancers. The nanoprobe has a raspberry shape, prepared by coating a dye-doped silica nanosphere with a mass of SERS tags, which gives high gains in fluorescence imaging and SERS measurement. The in situ detection of EGFR on the cell membrane surfaces after drug actions was achieved by using this nanoprobe, and the detection results agree with the enzyme-linked immunosorbent assay (ELISA) kit. Our study suggests that rosiglitazone hydrochloride (RH) may be a potential drug for diabetic patients with breast cancer, while the anti-cancer effect of metformin hydrochloride (MH) is debatable since MH slightly promotes the EGFR expression of MCF-7 cells in this study. This sensing platform endows more feasibility for highly sensitive and accurate feedback of pesticide effects at the membrane protein level.
Assuntos
Neoplasias da Mama , Feminino , Humanos , Neoplasias da Mama/tratamento farmacológico , Ensaio de Imunoadsorção Enzimática , Receptores ErbB , Imagem Óptica , FluorescênciaRESUMO
The capacity to regulate the signaling amplitude of membrane receptors in a user-defined manner would open various opportunities for precise biological study and therapy. While partial agonists enabled downtuning of cellular responses, they required esoteric optimization of the ligand-receptor interface, limiting their practical applications. Herein, we developed an aptamer-functionalized, tweezer-like nanodevice to dynamically modulate the cellular behavior through control over the distance between receptors in the dimer with no need to involve complicated structural analysis. By combining a reversible conformation switch with aptamer-based molecular recognition, this nanodevice showed excellent performance on dynamic regulation of CD28 receptor-mediated T cell immunity. With the modular design, this nanodevice could be extended to dynamically modulate the activity of other membrane receptors (e.g., c-Met), expecting to offer a new paradigm for precise study and manipulation of specific molecular events in complex biological systems.
Assuntos
Aptâmeros de Nucleotídeos , DNA , Aptâmeros de Nucleotídeos/química , Antígenos CD28 , DNA/química , Ligantes , Oligonucleotídeos , Transdução de SinaisRESUMO
Melatonin (N-acetyl-5-methoxytryptamine, MEL), its kynurenic (N1-acetyl-N2-formyl-5-methoxykynurenine, AFMK) and indolic derivatives (6-hydroxymelatonin, 6(OH)MEL and 5-methoxytryptamine, 5-MT) are endogenously produced in human epidermis. Melatonin, produced by the pineal gland, brain and peripheral organs, displays a diversity of physiological functions including anti-inflammatory, immunomodulatory, and anti-tumor capacities. Herein, we assessed their regulatory effect on melanogenesis using amelanotic (A375, Sk-Mel-28) and highly pigmented (MNT-1, melanotic) human melanoma cell lines. We discovered that subjected compounds decrease the downstream pathway of melanin synthesis by causing a significant drop of cyclic adenosine monophosphate (cAMP) level, the microphthalmia-associated transcription factor (MITF) and resultant collapse of tyrosinase (TYR) activity, and melanin content comparatively to N-phenylthiourea (PTU, a positive control). We observed a reduction in pigment in melanosomes visualized by the transmission electron microscopy. Finally, we assessed the role of G-protein-coupled seven-transmembrane-domain receptors. Obtained results revealed that nonselective MT1 and MT2 receptor antagonist (luzindole) or selective MT2 receptor antagonist (4-P-PDOT) did not affect dysregulation of the melanin pathway indicating a receptor-independent mechanism. Our findings, together with the current state of the art, provide a convenient experimental model to study the complex relationship between metabolites of melatonin and the control of pigmentation serving as a future and rationale strategy for targeted therapies of melanoma-affected patients.
Assuntos
Melanoma , Melatonina , Humanos , Melatonina/metabolismo , Melaninas , 5-Metoxitriptamina , Receptor MT2 de Melatonina , Melanoma/metabolismo , Monofenol Mono-OxigenaseRESUMO
The angiotensin-converting enzyme 2 (ACE2) has been identified as entry receptor on cells enabling binding and infection with the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) via trimeric spike (S) proteins protruding from the viral surface. It has been suggested that trimeric S proteins preferably bind to plasma membrane areas with high concentrations of possibly multimeric ACE2 receptors to achieve a higher binding and infection efficiency. Here we used direct stochastic optical reconstruction microscopy (dSTORM) in combination with different labeling approaches to visualize the distribution and quantify the expression of ACE2 on different cells. Our results reveal that endogenous ACE2 receptors are present as monomers in the plasma membrane with densities of only 1-2 receptors µm-2 . In addition, binding of trimeric S proteins does not induce the formation of ACE2 oligomers in the plasma membrane. Supported by infection studies using vesicular stomatitis virus (VSV) particles bearing S proteins our data demonstrate that a single S protein interaction per virus particle with a monomeric ACE2 receptor is sufficient for infection, which provides SARS-CoV-2 a high infectivity.
Assuntos
Enzima de Conversão de Angiotensina 2 , COVID-19 , Humanos , Enzima de Conversão de Angiotensina 2/metabolismo , SARS-CoV-2/metabolismo , Peptidil Dipeptidase A/metabolismo , Proteínas de Transporte/metabolismo , Ligação ProteicaRESUMO
BACKGROUND: Legume species are an important plant model because of their protein-rich physiology. The adaptability and productivity of legumes are limited by major biotic and abiotic stresses. Responses to these stresses directly involve plasma membrane receptor proteins known as receptor-like kinases and receptor-like proteins. Evaluating the homology relations among RLK and RLP for seven legume species, and exploring their presence among synteny blocks allow an increased understanding of evolutionary relations, physical position, and chromosomal distribution in related species and their shared roles in stress responses. RESULTS: Typically, a high proportion of RLK and RLP legume proteins belong to orthologous clusters, which is confirmed in this study, where between 66 to 90% of the RLKs and RLPs per legume species were classified in orthologous clusters. One-third of the evaluated syntenic blocks had shared RLK/RLP genes among both legumes and non-legumes. Among the legumes, between 75 and 98% of the RLK/RLP were present in syntenic blocks. The distribution of chromosomal segments between Phaseolus vulgaris and Vigna unguiculata, two species that diverged ~ 8 mya, were highly similar. Among the RLK/RLP synteny clusters, seven experimentally validated resistance RLK/RLP genes were identified in syntenic blocks. The RLK resistant genes FLS2, BIR2, ERECTA, IOS1, and AtSERK1 from Arabidopsis and SLSERK1 from Solanum lycopersicum were present in different pairwise syntenic blocks among the legume species. Meanwhile, only the LYM1- RLP resistant gene from Arabidopsis shared a syntenic blocks with Glycine max. CONCLUSIONS: The orthology analysis of the RLK and RLP suggests a dynamic evolution in the legume family, with between 66 to 85% of RLK and 83 to 88% of RLP belonging to orthologous clusters among the species evaluated. In fact, for the 10-species comparison, a lower number of singleton proteins were reported among RLP compared to RLK, suggesting that RLP positions are more physically conserved compared to RLK. The identification of RLK and RLP genes among the synteny blocks in legumes revealed multiple highly conserved syntenic blocks on multiple chromosomes. Additionally, the analysis suggests that P. vulgaris is an appropriate anchor species for comparative genomics among legumes.
Assuntos
Arabidopsis , Fosfotransferases , Filogenia , Glycine max , SinteniaRESUMO
Targeting mitochondrial quality control with melatonin has been found promising for attenuating diabetic cardiomyopathy (DCM), although the underlying mechanisms remain largely undefined. Activation of SIRT6 and melatonin membrane receptors exerts cardioprotective effects while little is known about their roles during DCM. Using high-fat diet-streptozotocin-induced diabetic rat model, we found that prolonged diabetes significantly decreased nocturnal circulatory melatonin and heart melatonin levels, reduced the expressions of cardiac melatonin membrane receptors, and decreased myocardial SIRT6 and AMPK-PGC-1α-AKT signaling. 16 weeks of melatonin treatment inhibited the progression of DCM and the following myocardial ischemia-reperfusion (MI/R) injury by reducing mitochondrial fission, enhancing mitochondrial biogenesis and mitophagy via re-activating SIRT6 and AMPK-PGC-1α-AKT signaling. After the induction of diabetes, adeno-associated virus carrying SIRT6-specific small hairpin RNA or luzindole was delivered to the animals. We showed that SIRT6 knockdown or antagonizing melatonin receptors abolished the protective effects of melatonin against mitochondrial dysfunction as evidenced by aggravated mitochondrial fission and reduced mitochondrial biogenesis and mitophagy. Additionally, SIRT6 shRNA or luzindole inhibited melatonin-induced AMPK-PGC-1α-AKT activation as well as its cardioprotective actions. Collectively, we demonstrated that long-term melatonin treatment attenuated the progression of DCM and reduced myocardial vulnerability to MI/R injury through preserving mitochondrial quality control. Melatonin membrane receptor-mediated SIRT6-AMPK-PGC-1α-AKT axis played a key role in this process. Targeting SIRT6 with melatonin treatment may be a promising strategy for attenuating DCM and reducing myocardial vulnerability to ischemia-reperfusion injury in diabetic patients.
Assuntos
Cardiomiopatias Diabéticas/prevenção & controle , Melatonina/farmacologia , Mitocôndrias Cardíacas/efeitos dos fármacos , Traumatismo por Reperfusão Miocárdica/prevenção & controle , Miócitos Cardíacos/efeitos dos fármacos , Biogênese de Organelas , Sirtuínas/metabolismo , Proteínas Quinases Ativadas por AMP/metabolismo , Animais , Diabetes Mellitus Experimental/complicações , Diabetes Mellitus Tipo 2/complicações , Cardiomiopatias Diabéticas/enzimologia , Cardiomiopatias Diabéticas/etiologia , Cardiomiopatias Diabéticas/patologia , Proteína Forkhead Box O3/metabolismo , Masculino , Mitocôndrias Cardíacas/enzimologia , Mitocôndrias Cardíacas/ultraestrutura , Traumatismo por Reperfusão Miocárdica/enzimologia , Traumatismo por Reperfusão Miocárdica/patologia , Miócitos Cardíacos/enzimologia , Miócitos Cardíacos/ultraestrutura , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Ratos Sprague-Dawley , Transdução de Sinais , Sirtuínas/genética , Fatores de TempoRESUMO
Epithelial membrane protein 3 (EMP3) is a tetraspan membrane protein overexpressed in isocitrate dehydrogenase-wild-type (IDH-wt) glioblastoma (GBM). Several studies reported high EMP3 levels as a poor prognostic factor in GBM patients. Experimental findings based on glioma and non-glioma models have demonstrated the role of EMP3 in the regulation of several membrane proteins known to drive IDH-wt GBM. In this review, we summarize what is currently known about EMP3 biology. We discuss the regulatory effects that EMP3 exerts on a variety of oncogenic receptors and discuss how these mechanisms may relate to IDH-wt GBM. Lastly, we enumerate the open questions towards EMP3 function in IDH-wt GBM.
Assuntos
Glioblastoma/metabolismo , Glicoproteínas de Membrana/metabolismo , Biomarcadores Tumorais/metabolismo , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/metabolismo , Expressão Gênica/genética , Regulação Neoplásica da Expressão Gênica/genética , Glioblastoma/genética , Glioma/genética , Glioma/metabolismo , Humanos , Isocitrato Desidrogenase/genética , Isocitrato Desidrogenase/metabolismo , Glicoproteínas de Membrana/fisiologia , PrognósticoRESUMO
Neuropathic pain is a refractory disease that occurs across the world and pharmacotherapy has limited efficacy and/or safety. This disease imposes a significant burden on both the somatic and mental health of patients; indeed, some patients have referred to neuropathic pain as being 'worse than death'. The pharmacological agents that are used to treat neuropathic pain at present can produce mild effects in certain patients, and induce many adverse reactions, such as sedation, dizziness, vomiting, and peripheral oedema. Therefore, there is an urgent need to discover novel drugs that are safer and more effective. Natural compounds from medical plants have become potential sources of analgesics, and evidence has shown that glycosides alleviated neuropathic pain via regulating oxidative stress, transcriptional regulation, ion channels, membrane receptors and so on. In this review, we summarize the epidemiology of neuropathic pain and the existing therapeutic drugs used for disease prevention and treatment. We also demonstrate how glycosides exhibit an antinociceptive effect on neuropathic pain in laboratory research and describe the antinociceptive mechanisms involved to facilitate the discovery of new drugs to improve the quality of life of patients experiencing neuropathic pain.
Assuntos
Glicosídeos/farmacologia , Glicosídeos/uso terapêutico , Neuralgia/tratamento farmacológico , Analgésicos/farmacologia , Analgésicos/uso terapêutico , Animais , Biomarcadores , Modelos Animais de Doenças , Suscetibilidade a Doenças , Regulação da Expressão Gênica/efeitos dos fármacos , Glicosídeos/química , Humanos , Ativação do Canal Iônico/efeitos dos fármacos , Camundongos , Neuralgia/diagnóstico , Neuralgia/epidemiologia , Neuralgia/etiologia , Estresse Oxidativo/efeitos dos fármacos , Manejo da Dor , Relação Estrutura-Atividade , Resultado do TratamentoRESUMO
Modifications of the composition or organization of the cancer cell membrane seem to be a promising targeted therapy. This approach can significantly enhance drug uptake or intensify the response of cancer cells to chemotherapeutics. There are several methods enabling lipid bilayer modifications, e.g., pharmacological, physical, and mechanical. It is crucial to keep in mind the significance of drug resistance phenomenon, ion channel and specific receptor impact, and lipid bilayer organization in planning the cell membrane-targeted treatment. In this review, strategies based on cell membrane modulation or reorganization are presented as an alternative tool for future therapeutic protocols.
Assuntos
Membrana Celular , Sistemas de Liberação de Medicamentos , Neoplasias , Membrana Celular/metabolismo , Membrana Celular/patologia , Humanos , Neoplasias/tratamento farmacológico , Neoplasias/metabolismo , Neoplasias/patologiaRESUMO
Synthetic molecular robots can execute sophisticated molecular tasks at nanometer resolution. However, a molecular robot capable of controlling cellular behavior remains unexplored. Herein, we report a self-propelled DNA robot operating on the cell membrane to control the migration of a cell. Driven by DNAzyme catalytic activity, the DNA robot could autonomously and stepwise move on the membrane-floating cell-surface receptors in a stochastic manner and simultaneously trigger the receptor-dimerization to activate downstream signaling for cell motility. The cell membrane-associated continuous motion and operation of a DNA robot allowed for the ultrasensitive regulation of MET/AKT signaling and cytoskeleton remodeling to enhance cell migration. Finally, we designed distinct conditional DNA robots to orthogonally manipulate the cell migration in a coculture of mixed cell populations. We have developed a novel strategy to engineer a cell-driving molecular robot, representing a promising avenue for precise cell manipulation with nanoscale resolution.
Assuntos
Membrana Celular/metabolismo , DNA Catalítico/metabolismo , DNA/metabolismo , Células A549 , Membrana Celular/química , Movimento Celular , DNA/química , HumanosRESUMO
We introduce concentration-dependent number and brightness (cdN&B), a fluorescence fluctuation technique that can be implemented on a standard confocal microscope and can report on the thermodynamics of membrane protein association in the native plasma membrane. It uses transient transfection to enable measurements of oligomer size as a function of receptor concentration over a broad range, yielding the association constant. We discuss artifacts in cdN&B that are concentration-dependent and can distort the oligomerization curves, and we outline procedures that can correct for them. Using cdN&B, we characterize the association of neuropilinâ 1 (NRP1), a protein that plays a critical role in the development of the embryonic cardiovascular and nervous systems. We show that NRP1 associates into a tetramer in a concentration-dependent manner, and we quantify the strength of the association. This work demonstrates the utility of cdN&B as a powerful tool in biophysical chemistry.