Phospholipid scramblases and Tubby-like proteins belong to a new superfamily of membrane tethered transcription factors

Motivation: Phospholipid scramblases (PLSCRs) constitute a family of cytoplasmic membrane-associated proteins that were identified based upon their capacity to mediate a Ca2+-dependent bidirectional movement of phospholipids across membrane bilayers, thereby collapsing the normally asymmetric distribution of such lipids in cell membranes. The exact function and mechanism(s) of these proteins nevertheless remains obscure: data from several laboratories now suggest that in addition to their putative role in mediating transbilayer flip/flop of membrane lipids, the PLSCRs may also function to regulate diverse processes including signaling, apoptosis, cell proliferation and transcription. A major impediment to deducing the molecular details underlying the seemingly disparate biology of these proteins is the current absence of any representative molecular structures to provide guidance to the experimental investigation of their function

Pb(II) Induces Scramblase Activation and Ceramide-Domain Generation in Red Blood Cells

The mechanisms of Pb(II) toxicity have been studied in human red blood cells using confocal microscopy, immunolabeling, fluorescence-activated cell sorting and atomic force microscopy. The process follows a sequence of events, starting with calcium entry, followed by potassium release, morphological change, generation of ceramide, lipid flip-flop and finally cell lysis. Clotrimazole blocks potassium channels and the whole process is inhibited. Immunolabeling reveals the generation of ceramide-enriched domains linked to a cell morphological change, while the use of a neutral sphingomyelinase inhibitor greatly delays the process after the morphological change, and lipid flip-flop is significantly reduced. These facts point to three major checkpoints in the process: first the upstream exchange of calcium and potassium, then ceramide domain formation, and finally the downstream scramblase activation necessary for cell lysis. In addition, partial non-cytotoxic cholesterol depletion of red blood cells accelerates the process as the morphological change occurs faster. Cholesterol could have a role in modulating the properties of the ceramide-enriched domains. This work is relevant in the context of cell death, heavy metal toxicity and sphingolipid signaling.AGA was a predoctoral student supported by the Basque Government and later by the University of the Basque Country (UPV/EHU). This work was also supported in part by grants from the Spanish Government (FEDER/MINECO BFU 2015-66306-P to F.M.G. and A.A.) and the Basque Government (IT849-13 to F.M.G. and IT838-13 to A.A.), and by the Swiss National Science Foundation

Membrane lipid interactions in intestinal ischemia/reperfusion-induced Injury

Ischemia, lack of blood flow, and reperfusion, return of blood flow, is a common phenomenon affecting millions of Americans each year. Roughly 30,000 Americans per year experience intestinal ischemia-reperfusion (IR), which is associated with a high mortality rate. Previous studies of the intestine established a role for neutrophils, eicosanoids, the complement system and naturally occurring antibodies in IR-induced pathology. Furthermore, data indicate involvement of a lipid or lipid-like moiety in mediating IR-induced damage. It has been proposed that exposure of neo-antigens are recognized by antibodies, triggering action of the complement cascade. While it is evident that the pathophysiology of IR-induced injury is complex and multi-factorial, we focus this review on the involvement of eicosanoids, phospholipids and neo-antigens in the early pathogenesis. Lipid changes occurring in response to IR, neo-antigens exposed and the role of a phospholipid transporter, phospholipid scramblase 1 will be discussed

The excited-state structure, vibrations, lifetimes, and nonradiative dynamics of jet-cooled 1-methylcytosine

We have investigated the S0 → S1 UV vibronic spectrum and time-resolved S1 state dynamics of jet-cooled amino-keto 1-methylcytosine (1MCyt) using two-color resonant two-photon ionization, UV/UV holeburning and depletion spectroscopies, as well as nanosecond and picosecond timeresolved pump/delayed ionization measurements. The experimental study is complemented with spin-component-scaled second-order coupled-cluster and multistate complete active space second order perturbation ab initio calculations. Above the weak electronic origin of 1MCyt at 31 852 cm−1 about 20 intense vibronic bands are observed. These are interpreted as methyl group torsional transitions coupled to out-of-plane ring vibrations, in agreement with the methyl group rotation and out-of-plane distortions upon 1ππ∗ excitation predicted by the calculations. The methyl torsion and ν′1 (butterfly) vibrations are strongly coupled, in the S1 state. The S0 → S1 vibronic spectrum breaks off at a vibrational excess energy Eexc ∼ 500 cm−1, indicating that a barrier in front of the ethylene-type S1 S0 conical intersection is exceeded, which is calculated to lie at Eexc = 366 cm−1. The S1 S0 internal conversion rate constant increases from kIC = 2 · 109 s−1 near the S1(v = 0) level to 1 · 1011 s−1 at Eexc = 516 cm−1. The 1ππ∗ state of 1MCyt also relaxes into the lower-lying triplet T1 (3ππ∗) state by intersystem crossing (ISC); the calculated spin-orbit coupling (SOC) value is 2.4 cm−1. The ISC rate constant is 10–100 times lower than kIC; it increases from kISC = 2 · 108 s−1 near S1(v = 0) to kISC = 2 · 109 s−1 at Eexc = 516 cm−1. The T1 state energy is determined from the onset of the time-delayed photoionization efficiency curve as 25 600 ± 500 cm−1. The T2 (3nπ∗) state lies >1500 cm−1 above S1(v = 0), so S1 T2 ISC cannot occur, despite the large SOC parameter of 10.6 cm−1. An upper limit to the adiabatic ionization energy of 1MCyt is determined as 8.41 ± 0.02 eV. Compared to cytosine, methyl substitution at N1 lowers the adiabatic ionization energy by ≥0.32 eV and leads to a much higher density of vibronic bands in the S0 → S1 spectrum. The effect of methylation on the radiationless decay to S0 and ISC to T1 is small, as shown by the similar break-off of the spectrum and the similar computed mechanismsThis research has been supported by the Schweiz. Nationalfonds (Grant Nos. 121993 and 132540), the Agència de Gestió d’Ajuts Universitaris i de Recerca (AGAUR) from Catalonia (Spain) (Grant No. 2014SGR1202), the Ministerio de Economía y Competividad (MINECO) from Spain (Grant No. CTQ2015-69363-P), and the National Natural Science Foundation of China (Grant No. 21303007

Bsx, a novel hypothalamic factor linking feeding with locomotor activity, is regulated by energy availability

Es un estudio en modelos de rata acerca del papel de BSX, la actividad y la alimentación.We recently reported that the hypothalamic homeobox domain transcription factor Bsx plays an essential role in the central nervous system control of spontaneous physical activity and the generation of hyperphagic responses. Moreover, we found Bsx to be a master regulator for the hypothalamic expression of key orexigenic neuropeptide Y and agouti gene-related protein. We now hypothesized that Bsx, which is expressed in the dorsomedial and arcuate nucleus (ARC) of the hypothalamus, is regulated by afferent signals in response to peripheral energy balance. Bsx expression was analyzed using in situ hybridization in fed vs. fasted (24 h) and ghrelin vs. leptin-treated rats, as well as in mice deficient for leptin or the ghrelin signaling. Ghrelin administration increased, whereas ghrelin receptor antagonist decreased ARC Bsx expression. Leptin injection attenuated the fasting-induced increase in ARC Bsx levels but had no effect in fed rats. Dorsomedial hypothalamic nucleus Bsx expression was unaffected by pharmacological modifications of leptin or ghrelin signaling. Obese leptin-deficient (ob/ob) mice, but not obese melanocortin 4 receptor-knockout mice, showed higher expression of Bsx, consistent with dependency from afferent leptin rather than increased adiposity per se. Interestingly, exposure to a high-fat diet triggered Bsx expression, consistent with the concept that decreased leptin signaling due to a highfat diet induced leptin resistance. Our data indicate that ARC Bsx expression is specifically regulated by afferent energy balance signals, including input from leptin and ghrelin. Future studies will be necessary to test if Bsx may be involved in the pathogenesis of leptin resistance

Ghrelin-induced hypothermia: A physiological basis but no clinical risk

Ghrelin increases food intake and decreases energy expenditure, promoting a positive energy balance. We observed a single case of serious hypothermia during sustained ghrelin treatment in a male subject, suggesting that ghrelin may play a role in the regulation of body temperature. We therefore investigated the effect of ghrelin treatment on body temperature in rodents and humans under controlled conditions. Intriguingly, we could demonstrate ghrelin binding in axon terminals of the medial preoptic area of the hypothalamus located in the vicinity of cold-sensitive neurons. This localization of ghrelin receptors provides a potential anatomical basis for the regulation of body temperature by ghrelin. However, our follow-up studies also indicated that neither a chronic i.c.v. application of ghrelin in rats, nor a single s.c. injection under cold exposure in mice resulted in a relevant decrease in body core temperature. In addition, a four-hour intravenous ghrelin infusion did not decrease body surface temperature in healthy humans. We concluded that while there is a theoretical molecular basis for ghrelin to modify body temperature in mammals, its magnitude is irrelevant under physiologic circumstances. Hypothermia is not likely to represent a serious risk associated with this agent and pathway

An integrated genomic approach to dissect the genetic landscape regulating the cell-to-cell transfer of α-synuclein

Neuropathological and experimental evidence suggests that the cell-to-cell transfer of α-synuclein has an important role in the pathogenesis of Parkinson's disease (PD). However, the mechanism underlying this phenomenon is not fully understood. We undertook a small interfering RNA (siRNA), genome-wide screen to identify genes regulating the cell-to-cell transfer of α-synuclein. A genetically encoded reporter, GFP-2A-αSynuclein-RFP, suitable for separating donor and recipient cells, was transiently transfected into HEK cells stably overexpressing α-synuclein. We find that 38 genes regulate the transfer of α-synuclein-RFP, one of which is ITGA8, a candidate gene identified through a recent PD genome-wide association study (GWAS). Weighted gene co-expression network analysis (WGCNA) and weighted protein-protein network interaction analysis (WPPNIA) show that those hits cluster in networks that include known PD genes more frequently than expected by random chance. The findings expand our understanding of the mechanism of α-synuclein spread

The Phospholipid Scramblases 1 and 4 Are Cellular Receptors for the Secretory Leukocyte Protease Inhibitor and Interact with CD4 at the Plasma Membrane

Secretory leukocyte protease inhibitor (SLPI) is secreted by epithelial cells in all the mucosal fluids such as saliva, cervical mucus, as well in the seminal liquid. At the physiological concentrations found in saliva, SLPI has a specific antiviral activity against HIV-1 that is related to the perturbation of the virus entry process at a stage posterior to the interaction of the viral surface glycoprotein with the CD4 receptor. Here, we confirm that recombinant SLPI is able to inhibit HIV-1 infection of primary T lymphocytes, and show that SLPI can also inhibit the transfer of HIV-1 virions from primary monocyte-derived dendritic cells to autologous T lymphocytes. At the molecular level, we show that SLPI is a ligand for the phospholipid scramblase 1 (PLSCR1) and PLSCR4, membrane proteins that are involved in the regulation of the movements of phospholipids between the inner and outer leaflets of the plasma membrane. Interestingly, we reveal that PLSCR1 and PLSCR4 also interact directly with the CD4 receptor at the cell surface of T lymphocytes. We find that the same region of the cytoplasmic domain of PLSCR1 is involved in the binding to CD4 and SLPI. Since SLPI was able to disrupt the association between PLSCR1 and CD4, our data suggest that SLPI inhibits HIV-1 infection by modulating the interaction of the CD4 receptor with PLSCRs. These interactions may constitute new targets for antiviral intervention

Interactions between selected bile salts and Triton X-100 or sodium lauryl ether sulfate

<p>Abstract</p> <p>Background</p> <p>In order to develop colloidal drug carriers with desired properties, it is important to determine physico-chemical characteristics of these systems. Bile salt mixed micelles are extensively studied as novel drug delivery systems. The objective of the present investigation is to develop and characterize mixed micelles of nonionic (Triton X-100) or anionic (sodium lauryl ether sulfate) surfactant having oxyethylene groups in the polar head and following bile salts: cholate, deoxycholate and 7-oxodeoxycholate.</p> <p>Results</p> <p>The micellization behaviour of binary anionic-nonionic and anionic-anionic surfactant mixtures was investigated by conductivity and surface tension measurements. The results of the study have been analyzed using Clint's, Rubingh's, and Motomura's theories for mixed binary systems. The negative values of the interaction parameter indicate synergism between micelle building units. It was noticed that Triton X-100 and sodium lauryl ether sulfate generate the weakest synergistic interactions with sodium deoxycholate, while 7-oxodeoxycholate creates the strongest attractive interaction with investigated co-surfactants.</p> <p>Conclusion</p> <p>It was concluded that increased synergistic interactions can be attributed to the larger number of hydrophilic groups at α side of the bile salts. Additionally, 7-oxo group of 7-oxodeoxycholate enhance attractive interactions with selected co-surfactants more than 7-hydroxyl group of sodium cholate.</p