The HIV capsid mimics karyopherin engagement of FG-nucleoporins.

HIV can infect non-dividing cells because the viral capsid can overcome the selective barrier of the nuclear pore complex and deliver the genome directly into the nucleus1,2. Remarkably, the intact HIV capsid is more than 1,000 times larger than the size limit prescribed by the diffusion barrier of the nuclear pore3. This barrier in the central channel of the nuclear pore is composed of intrinsically disordered nucleoporin domains enriched in phenylalanine-glycine (FG) dipeptides. Through multivalent FG interactions, cellular karyopherins and their bound cargoes solubilize in this phase to drive nucleocytoplasmic transport4. By performing an in vitro dissection of the nuclear pore complex, we show that a pocket on the surface of the HIV capsid similarly interacts with FG motifs from multiple nucleoporins and that this interaction licences capsids to penetrate FG-nucleoporin condensates. This karyopherin mimicry model addresses a key conceptual challenge for the role of the HIV capsid in nuclear entry and offers an explanation as to how an exogenous entity much larger than any known cellular cargo may be able to non-destructively breach the nuclear envelope.

Electrical characterizations of biomimetic molecular layers on gold and silicon substrates.

Electrical impedance technology was used to characterize DNA recognition in a monolayer containing single-stranded DNA probes immobilized on a gold substrate using thiol self-assembly chemistry. Recognition of targeted complementary DNA was principally correlated with an eight-fold increase in the conductance of the monolayer and attributed to electron conduction through double helices formed upon the binding of the DNA targets to the probes. The high recognitive sensitivity was possible without the use of the redox labels or large bias voltages required for recognition using cyclic and Osteryoung square wave voltammetry. The impedance technology also provided atomic resolution of a hybrid bimolecular lipid membrane formed by deposition of a phospholipid:cholesterol monolayer onto a hydrophobic alkyl monolayer covalently attached to a silicon substrate via silicon-carbon bonds. Atomic resolution was achieved through preparation of membranes on surfaces approaching atomic flatness and the performance of impedance measurements over precisely defined areas of the surface in contact with solutions. Principally capacitive properties distinguished between the immobilized (octadecyl) and more fluidic (lipid:cholesterol) leaflets of the hybrid membrane. The lipid:cholesterol leaflets were structurally similar to those leaflets in free-standing bimolecular lipid membranes. The hybrid membrane therefore provides a highly stable and physiologically relevant surface for studying biomolecular interactions with membrane surfaces.

Effects of singlet oxygen on membrane sterols in the yeast Saccharomyces cerevisiae.

Photodynamic treatment of the yeast Saccharomyces cerevisiae with the singlet oxygen sensitizer toluidine blue and visible light leads to rapid oxidation of ergosterol and accumulation of oxidized ergosterol derivatives in the plasma membrane. The predominant oxidation product accumulated was identified as 5alpha, 6alpha-epoxy-(22E)-ergosta-8,22-dien-3beta,7a lpha-diol (8-DED). 9(11)-dehydroergosterol (DHE) was identified as a minor oxidation product. In heat inactivated cells ergosterol is photooxidized to ergosterol epidioxide (EEP) and DHE. Disrupted cell preparations of S. cerevisiae convert EEP to 8-DED, and this activity is abolished in a boiled control indicating the presence of a membrane associated enzyme with an EEP isomerase activity. Yeast selectively mobilizes ergosterol from the intracellular sterol ester pool to replenish the level of free ergosterol in the plasma membrane during singlet oxygen oxidation. The following reaction pathway is proposed: singlet oxygen-mediated oxidation of ergosterol leads to mainly the formation of EEP, which is enzymatically rearranged to 8-DED. Ergosterol 7-hydroperoxide, a known minor product of the reaction of singlet oxygen with ergosterol, is formed at a much lower rate and decomposes to give DHE. Changes of physical properties of the plasma membrane are induced by depletion of ergosterol and accumulation of polar derivatives. Subsequent permeation of photosensitizer through the plasma membrane into the cell leads to events including impairment of mitochondrial function and cell inactivation.