Cholesterol and phosphatidylserine are engaged in adenoviral dodecahedron endocytosis

https://doi.org/10.1016/j.bbamem.2018.09.002Get rights and content
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Highlights

  • Adenoviral dodecahedron (Dd) is an efficient delivery vector.

  • Cell type specificity of Dd is not well understood.

  • Dd displays specific affinity to phosphatidylserine.

  • Cholesterol is indirectly involved in Dd endocytosis.

Abstract

Adenoviral dodecahedron is a virus-like particle composed of twelve penton base proteins, derived from the capsid of human adenovirus type 3. Due to the high cell penetration capacity, it was used as a vector for protein, peptide and drug delivery. Two receptors are known to be involved in the endocytic dodecahedron uptake, namely αv integrins and heparan sulfate proteoglycans. Since it has been observed, that dodecahedron efficiently penetrates a wide range of cancer cells, it suggests that other cellular compounds may play a role in the particle endocytosis. To shed some light onto the interactions with membrane lipids and their potential role in dodecahedron entry, we performed a series of experiments including biochemical assays, fluorescence confocal imaging of giant unilamellar vesicles and surface plasmon resonance, which indicated specific preference of the particle to anionic phosphatidylserine. Experiments performed on cholesterol-depleted epithelial cells showed that cholesterol is essential in the endocytic uptake, however a direct interaction was not observed. We believe that the results will allow to better understand the role of lipids in dodecahedron entry and to design more specific dodecahedron-based vectors for drug delivery to cancer cells.

Abbreviations

BSA
bovine serum albumin
CL
cardiolipin
Dd
dodecahedron
ECL
enhanced chemiluminescence
FBS
fetal bovine serum
GUV
giant unilamellar vesicle
HAd3
human adenovirus serotype 3
HRP
horseradish peroxidase
HSPG
heparan sulphate proteoglycan
LUV
large unilamellar vesicles
mβCD
methyl-β-cyclodextrin
NHS
N-hydroxysulfoxuccinimide
PA
phosphatic acid
Pb
penton base protein
PBS
phosphate buffered saline
PC
phosphatidylcholine
PE
phosphatidylethanolamine
PG
phosphatidylglycerol
PI
phosphatidylinositol (PtdIns4)
PIP2
PtdIns(4,5)P
PIP3
PtdIns(3,4,5)P
PIP4
PtdIns(4)P
PS
phosphatidylserine
RU
response units
SA
3-sulfogalactosylceramide (sulfatide)
SDS-PAGE
sodium dodecyl sulfate polyacrylamide gel electrophoresis
SPR
surface plasmon resonance
TMB
3,3′,5,5′-tetramethylbenzidine
VLP
virus-like particle

Keywords

Virus-like particles
Protein-lipid interactions
Giant unilamellar vesicles
Artificial model membranes
Fluorescence confocal microscopy
Phosphatidylserine

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