Spatial Organization of Lipid Phases in Micropatterned Polymer-Supported Membranes

Roder, Friedrich; Birkholz, Oliver; Beutel, Oliver; Paterok, Dirk; Piehler, Jacob

doi:10.1021/ja310186g

Cited by 32 publications

(27 citation statements)

References 37 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Monomeric EGFP was obtained by the A206K mutation within the EGFP sequence of pEGFP-N1 (Clontech). An artificial transmembrane domain (TMD) with the sequence AS ALAALAALAALAALAALAALA KSSRL (ALA7) (as described by 64 ) extracellular fused to HaloTag (Promega) and mTagBFP (obtained from Vladislav Verkhusha, New York 65 ) and intracellularly fused to human STAT2 was cloned into pDisplay ™ (Invitrogen). For cloning of pSEMS-HaloTag®-IFNAR2, the gene coding for the HaloTag followed by the genes of full length IFNAR2 or IFNAR2Δ375 without the N-terminal signal sequences were inserted into pDisplay (Invitrogen).…”

Section: Methodsmentioning

confidence: 99%

STAT2 is an essential adaptor in USP18-mediated suppression of type I interferon signaling

Arimoto

Löchte

Stoner

et al. 2017

Nat Struct Mol Biol

Self Cite

161

164

View full text Add to dashboard Cite

Type I interferons (IFNs) are multifunctional cytokines that regulate immune responses and cellular functions but also can have detrimental effects on human health. A tight regulatory network therefore controls IFN signaling, which in turn interferes with medical interventions. The JAK-STAT signaling pathway transmits the IFN extracellular signal to the nucleus for alterations of gene expression. STAT2 is a well-known essential and specific positive effector of type I IFN signaling. Here, we report that STAT2 is also a previously unrecognized crucial component of the USP18-mediated negative feedback control in both, human and murine cells. We found that STAT2 recruits USP18 to the type I IFN receptor subunit IFNAR2 via its constitutive membrane-distal STAT2 binding site. This mechanistic coupling of effector and negative feedback functions of STAT2 provides novel strategies in treatment of IFN signaling related human diseases.

show abstract

Section: Methodsmentioning

confidence: 99%

STAT2 is an essential adaptor in USP18-mediated suppression of type I interferon signaling

Arimoto

Löchte

Stoner

et al. 2017

Nat Struct Mol Biol

Self Cite

161

164

View full text Add to dashboard Cite

show abstract

“…By using a patterning of polymers with either a saturated or an unsaturated lipid, the binding of particular lipids to the surface was altered leading to the recruitment of certain membrane components. 27,28 Alternatively, Morigaki and coworkers [29][30][31] used polymers that were partially or fully removed in confined areas to control membrane adhesion to the surface. They found that the l o -phase is localized in areas without polymer coating and the l d -phase in areas with partial polymer coating.…”

Section: Introductionmentioning

confidence: 99%

Phase separation in pore-spanning membranes induced by differences in surface adhesion

Sibold

Tewaag

Vagedes

et al. 2020

Phys. Chem. Chem. Phys.

View full text Add to dashboard Cite

Lipid domains in plasma membranes act as molecular sorting platforms for e.g., signalling processes. In model membranes, such as freestanding or supported bilayers, some lipid domains with defined chemical composition, lipid packing and physical behaviour can be reproduced. However, in vivo, the plasma membrane experiences a proteinaceous scaffold underneath, which can sort, compartmentalize and recruit components within the membrane. The influence of such scaffolds on the phase behaviour of lipid membranes has been barely studied. Here, we investigated the partial attachment of a membrane to a support and its influence on the phase behaviour using pore-spanning membranes (PSMs). PSMs were prepared on SiO x=1-2 functionalized silicon substrates with 1.2 mm-sized pores by spreading giant unilamellar vesicles (GUVs) composed of DOPC/sphingomyelin (1 : 1) with different cholesterol concentrations. Using two different fluorophores, PSMs were visualized by fluorescence microscopy allowing us to distinguish between different membrane phases, a gel (l b ), a liquid ordered (l o ), and a liquid disordered (l d ) phase. At low cholesterol concentrations, coexistence of l b and l d was found, while at higher cholesterol concentrations, coexistence of l o and l d was predominant. Below the mixing temperature, determined by temperature scans, the more ordered phase was always found in the freestanding PSMs, whereas the l d -phase was present in the supported PSMs. We attribute this lipid sorting to a stronger adhesion of the l d -phase lipids to the underlying scaffold. The difference in adhesion alters the phase behaviour from a nominal DOPC/sphingomyelin (1 : 1) mixture to a DOPC/ sphingomyelin (1 : 2-1 : 4) mixture compared to phase diagrams obtained from GUVs highlighting the importance of differential adhesive surfaces on lipid domain formation.

show abstract

“…The L o /L d partitioning of urokinase receptors and integrin has been evaluated in SPBs (19,20). Micropatterning techniques can also be applied to SPBs for generating a patterned array of L o and L d domains (21,22). SPBs with spatially well-defined membrane domains should provide a robust and scalable platform for systematically and quantitatively studying the raftophilicity of a wide variety of membrane proteins.…”

Section: Introductionmentioning

confidence: 99%

Evaluating the Raftophilicity of Rhodopsin Photoreceptor in a Patterned Model Membrane

Tanimoto

Okada

Hayashi

et al. 2015

Biophysical Journal

View full text Add to dashboard Cite

Lipid rafts in the cell membrane are believed to affect various membrane functions, including the signaling by G-protein coupled receptors (GPCRs). However, the regulatory roles of lipid rafts on GPCRs' functions are still poorly understood, partially owing to the lack of the methods to quantitatively evaluate the affinity of membrane proteins to lipid raft (raftophilicity). Here, we describe a methodology to gauge the raftophilicity of a representative GPCR in vertebrate photoreceptor, i.e., rhodopsin (Rh), and its cognate G protein transducin (Gt) by using a patterned model membrane. We generated a substrate-supported planar lipid bilayer that has patterned regions of liquid-ordered (Lo) and liquid-disordered (Ld) membrane domains. We reconstituted Rh and Gt into the patterned membrane and observed their lateral distribution and diffusion. Mobile and functional Rh molecules could be reconstituted through the rapid dilution of solubilized Rh, by optimizing the reconstitution conditions including the chamber design, protein/detergent concentrations, and solution mixing. We determined the partition and diffusion coefficients of Rh and Gt in the Lo-rich and Ld-rich regions. Both Rh and Gt were predominantly localized in the Ld phase, suggesting their low affinity to lipid rafts. Patterned model membrane offers a robust and scalable platform for systematically and quantitatively studying the functional roles of lipid rafts in biological membranes including retinal disk membranes.

show abstract

Spatial Organization of Lipid Phases in Micropatterned Polymer-Supported Membranes

Cited by 32 publications

References 37 publications

STAT2 is an essential adaptor in USP18-mediated suppression of type I interferon signaling

STAT2 is an essential adaptor in USP18-mediated suppression of type I interferon signaling

Phase separation in pore-spanning membranes induced by differences in surface adhesion

Evaluating the Raftophilicity of Rhodopsin Photoreceptor in a Patterned Model Membrane

Contact Info

Product

Resources

About