Plasma membrane vesiculation in 3T3 and Sv3T3 cells: I. morphological and biochemical characterization

Scott, Robert E.; Perkins, Roger; Zschunke, Michael A.; Hoerl, Bryan J.; Maercklein, Peter B.

doi:10.1242/jcs.35.1.229

Cited by 108 publications

(33 citation statements)

References 31 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Although the formation of PM vesicles results in some loss of membrane asymmetry, as indicated by ANXA binding to the outer leaflet (Figures S5 and S6), the proteins, however, associated with the PM vesicles do retain their correct orientation, indicating that the protein composition of the PM vesicles is likely not affected. The PM vesicle assay therefore allows full control over protein location and orientation for both membrane-binding proteins and integral membrane proteins, which is in striking contrast to other model systems like protein reconstituted GUVs. , Additionally, the PM vesicles are devoid of internal cellular structures and therefore can be used to study the interplay between the PM and protein dynamics.…”

Section: Results and Discussionmentioning

confidence: 99%

“…The PM vesicle assay therefore allows full control over protein location and orientation for both membrane-binding proteins and integral membrane proteins, which is in striking contrast to other model systems like protein reconstituted GUVs. 11,32 Additionally, the PM vesicles are devoid of internal cellular structures 33 and therefore can be used to study the interplay between the PM and protein dynamics.…”

Section: Resultsmentioning

confidence: 99%

See 1 more Smart Citation

Curvature- and Phase-Induced Protein Sorting Quantified in Transfected Cell-Derived Giant Vesicles

et al. 2019

View full text Add to dashboard Cite

Eukaryotic cells possess a dynamic network of membranes that vary in lipid composition. To perform numerous biological functions, cells modulate their shape and the lateral organization of proteins associated with membranes. The modulation is generally facilitated by physical cues that recruit proteins to specific regions of the membrane. Analyzing these cues is difficult due to the complexity of the membrane conformations that exist in cells. Here, we examine how different types of membrane proteins respond to changes in curvature and to lipid phases found in the plasma membrane. By using giant plasma membrane vesicles derived from transfected cells, the proteins were positioned in the correct orientation and the analysis was performed in plasma membranes with a biological composition. Nanoscale membrane curvatures were generated by extracting nanotubes from these vesicles with an optical trap. The viral membrane protein neuraminidase was not sensitive to curvature, but it did exhibit strong partitioning (coefficient of K = 0.16) disordered membrane regions. In contrast, the membrane repair protein annexin 5 showed a preference for nanotubes with a density up to 10–15 times higher than that on the more flat vesicle membrane. The investigation of nanoscale effects in isolated plasma membranes provides a quantitative platform for studying peripheral and integral membrane proteins in their natural environment.

show abstract

Section: Results and Discussionmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Curvature- and Phase-Induced Protein Sorting Quantified in Transfected Cell-Derived Giant Vesicles

et al. 2019

View full text Add to dashboard Cite

show abstract

“…For some cases, the additional activation of the heterologously expressed receptor can further increase the cellular release of plasma membrane vesicles. , Certain chemical compounds can induce the formation of plasma membrane vesicles. Examples are formaldehyde, related low-molecular weight aldehydes, and −SH blocking agents such as N-ethylmaleimide or dithiothreitol on the one side, − or less-harsh agents such as cytochalasin B known to disrupt the interaction between the cellular cytoskeleton and the plasma membrane on the other side. , The size of the vesicles can be tuned from the 100 nm to 10-μm range by adjusting the concentration of the disrupting agent and the time of incubation with living cells; the vesicles are isolated by a combination of centrifugation and filtration steps. ,− Individual plasma membrane vesicles have been drawn mechanically from living biological cells using optical tweezers. , …”

Section: Production and Isolation Of Single-vesiclesmentioning

confidence: 99%

Single-Vesicle Assays Using Liposomes and Cell-Derived Vesicles: From Modeling Complex Membrane Processes to Synthetic Biology and Biomedical Applications

2018

View full text Add to dashboard Cite

The plasma membrane is of central importance for defining the closed volume of cells in contradistinction to the extracellular environment. The plasma membrane not only serves as a boundary, but it also mediates the exchange of physical and chemical information between the cell and its environment in order to maintain intra- and intercellular functions. Artificial lipid- and cell-derived membrane vesicles have been used as closed-volume containers, representing the simplest cell model systems to study transmembrane processes and intracellular biochemistry. Classical examples are studies of membrane translocation processes in plasma membrane vesicles and proteoliposomes mediated by transport proteins and ion channels. Liposomes and native membrane vesicles are widely used as model membranes for investigating the binding and bilayer insertion of proteins, the structure and function of membrane proteins, the intramembrane composition and distribution of lipids and proteins, and the intermembrane interactions during exo- and endocytosis. In addition, natural cell-released microvesicles have gained importance for early detection of diseases and for their use as nanoreactors and minimal protocells. Yet, in most studies, ensembles of vesicles have been employed. More recently, new micro- and nanotechnological tools as well as novel developments in both optical and electron microscopy have allowed the isolation and investigation of individual (sub)micrometer-sized vesicles. Such single-vesicle experiments have revealed large heterogeneities in the structure and function of membrane components of single vesicles, which were hidden in ensemble studies. These results have opened enormous possibilities for bioanalysis and biotechnological applications involving unprecedented miniaturization at the nanometer and attoliter range. This review will cover important developments toward single-vesicle analysis and the central discoveries made in this exciting field of research.

show abstract

“…The cell membrane blebs when exposed to a sulfhydryl-blocking reagent . This cell-blebbing phenomenon has been adopted to study membrane raft phases using giant plasma membrane vesicles. , Despite the current utility of giant plasma membrane vesicles, their large size range and accompanying unacceptable polydispersivity are not suitable for therapeutic and diagnostic delivery, demanding fine-tuned preparation of EVs via sulfhydryl-blocking. By applying a previous observation and further optimizing it, we demonstrate that chemically induced cell-blebbing via sulfhydryl-blocking can be harnessed to produce nanosized EVs in a desirable quantity, resulting in nanovesicles induced by sulfhydryl-blocking (NIbS).…”

mentioning

confidence: 99%

Cancer Cell-Derived, Drug-Loaded Nanovesicles Induced by Sulfhydryl-Blocking for Effective and Safe Cancer Therapy

et al. 2018

View full text Add to dashboard Cite

Extracellular vesicles (EVs) pose great promise as therapeutic carriers due to their ideal size range and intrinsic biocompatibility. Limited scalability, poor quality control during production, and cumbersome isolation and purification processes have caused major setbacks in the progression of EV therapeutics to the clinic. Here, we overcome these setbacks by preparing cell-derived nanovesicles induced by sulfhydryl-blocking (NIbS), in the desirable size range for therapeutic delivery, that can be further loaded with the chemotherapeutic drug, doxorubicin (DOX), resulting in NIbS/DOX. Applicable to most cell types, this chemical blebbing approach enables efficient, quick, and simple harvest and purification as well as easily scalable production. Cellular uptake and intracellular release of DOX was improved using NIbS/DOX compared to a liposomal formulation. We also confirmed that in tumor-challenged C57BL/6 mice NIbS/DOX significantly slowed tumor growth and led to improved survival compared to treatment with free drug or liposomal drug. NIbS are a promising therapeutic carrier for improving cancer treatment outcomes since they are easy to prepare at a large scale, good candidates for drug loading, and capable of efficient administration of therapeutic agents with avoided nonspecific major distribution in vital organs. In addition, the utility of NIbS can be easily expanded to immunotherapy, gene therapy, and cell therapy when they are derived from applicable cell types.

show abstract

Plasma membrane vesiculation in 3T3 and Sv3T3 cells: I. morphological and biochemical characterization

Cited by 108 publications

References 31 publications

Curvature- and Phase-Induced Protein Sorting Quantified in Transfected Cell-Derived Giant Vesicles

Curvature- and Phase-Induced Protein Sorting Quantified in Transfected Cell-Derived Giant Vesicles

Single-Vesicle Assays Using Liposomes and Cell-Derived Vesicles: From Modeling Complex Membrane Processes to Synthetic Biology and Biomedical Applications

Cancer Cell-Derived, Drug-Loaded Nanovesicles Induced by Sulfhydryl-Blocking for Effective and Safe Cancer Therapy

Contact Info

Product

Resources

About