Role and significance of asprosin in feeding behaviour and metabolism

This article presents a review of available information on asprosin — a hormone of white adipose tissue discovered in 2016. The history of its discovery, as well as its action mechanisms and main targets are examined. Changes in the plasma level of asprosin under some pathological conditions are analysed. The importance of studying asprosin is determined by its functions: asprosin regulates physiological processes during fasting and plays an important role in the development of metabolic disorders, such as insulin resistance. There are relatively few studies concerned with asprosin; however, this hormone can already be considered as a diagnostic marker and a potential target in the treatment of certain metabolic disorders, e.g. diabetes mellitus and obesity

Complex I-mediated reactive oxygen species generation: modulation by cytochrome c and NAD(P)+ oxidation-reduction state.

Several lines of evidence indicate that mitochondrial reactive oxygen species (ROS) generation is the major source of oxidative stress in the cell. It has been shown that ROS production accompanies cytochrome c release in different apoptotic paradigms, but the site(s) of ROS production remain obscure. In the current study, we demonstrate that loss of cytochrome c by mitochondria oxidizing NAD(+)-linked substrates results in a dramatic increase of ROS production and respiratory inhibition. This increased ROS production can be mimicked by rotenone, a complex I inhibitor, as well as other chemical inhibitors of electron flow that act further downstream in the electron transport chain. The effects of cytochrome c depletion from mitoplasts on ROS production and respiration are reversible upon addition of exogenous cytochrome c. Thus in these models of mitochondrial injury, a primary site of ROS generation in both brain and heart mitochondria is proximal to the rotenone inhibitory site, rather than in complex III. ROS production at complex I is critically dependent upon a highly reduced state of the mitochondrial NAD(P)(+) pool and is achieved upon nearly complete inhibition of the respiratory chain. Redox clamp experiments using the acetoacetate/L-beta-hydroxybutyrate couple in the presence of a maximally inhibitory rotenone concentration suggest that the site is approx. 50 mV more electronegative than the NADH/NAD(+) couple. In the absence of inhibitors, this highly reduced state of mitochondria can be induced by reverse electron flow from succinate to NAD(+), accounting for profound ROS production in the presence of succinate. These results lead us to propose a model of thermodynamic control of mitochondrial ROS production which suggests that the ROS-generating site of complex I is the Fe-S centre N-1a

Disruption of mitochondrial quality control genes promotes caspase-resistant cell survival following apoptotic stimuli

In cells undergoing cell-intrinsic apoptosis, mitochondrial outer membrane permeabilization (MOMP) typically marks an irreversible step in the cell death process. However, in some cases, a subpopulation of treated cells can exhibit a sublethal response, termed “minority MOMP.” In this phenomenon, the affected cells survive, despite a low level of caspase activation and subsequent limited activation of the endonuclease caspase-activated DNase (DNA fragmentation factor subunit beta). Consequently, these cells can experience DNA damage, increasing the probability of oncogenesis. However, little is known about the minority MOMP response. To discover genes that affect the MOMP response in individual cells, we conducted an imaging-based phenotypic siRNA screen. We identified multiple candidate genes whose downregulation increased the heterogeneity of MOMP within single cells, among which were genes related to mitochondrial dynamics and mitophagy that participate in the mitochondrial quality control (MQC) system. Furthermore, to test the hypothesis that functional MQC is important for reducing the frequency of minority MOMP, we developed an assay to measure the clonogenic survival of caspase-engaged cells. We found that cells deficient in various MQC genes were indeed prone to aberrant post-MOMP survival. Our data highlight the important role of proteins involved in mitochondrial dynamics and mitophagy in preventing apoptotic dysregulation and oncogenesis.publishe

HIV-1 Fusion with CD4+ T cells Is Promoted by Proteins Involved in Endocytosis and Intracellular Membrane Trafficking

The HIV-1 entry pathway into permissive cells has been a subject of debate. Accumulating evidence, including our previous single virus tracking results, suggests that HIV-1 can enter different cell types via endocytosis and CD4/coreceptor-dependent fusion with endosomes. However, recent studies that employed indirect techniques to infer the sites of HIV-1 entry into CD4+ T cells have concluded that endocytosis does not contribute to infection. To assess whether HIV-1 enters these cells via endocytosis, we probed the role of intracellular trafficking in HIV-1 entry/fusion by a targeted shRNA screen in a CD4+ T cell line. We performed a screen utilizing a direct virus-cell fusion assay as readout and identified several host proteins involved in endosomal trafficking/maturation, including Rab5A and sorting nexins, as factors regulating HIV-1 fusion and infection. Knockdown of these proteins inhibited HIV-1 fusion irrespective of coreceptor tropism, without altering the CD4 or coreceptor expression, or compromising the virus’ ability to mediate fusion of two adjacent cells initiated by virus-plasma membrane fusion. Ectopic expression of Rab5A in non-permissive cells harboring Rab5A shRNAs partially restored the HIV-cell fusion. Together, these results implicate endocytic machinery in productive HIV-1 entry into CD4+ T cells

Bax Activation Initiates the Assembly of a Multimeric Catalyst that Facilitates Bax Pore Formation in Mitochondrial Outer Membranes

<div><p>Bax/Bak-mediated mitochondrial outer membrane permeabilization (MOMP) is essential for “intrinsic” apoptotic cell death. Published studies used synthetic liposomes to reveal an intrinsic pore-forming activity of Bax, but it is unclear how other mitochondrial outer membrane (MOM) proteins might facilitate this function. We carefully analyzed the kinetics of Bax-mediated pore formation in isolated MOMs, with some unexpected results. Native MOMs were more sensitive than liposomes to added Bax, and MOMs displayed a lag phase not observed with liposomes. Heat-labile MOM proteins were required for this enhanced response. A two-tiered mathematical model closely fit the kinetic data: first, Bax activation promotes the assembly of a multimeric complex, which then catalyzes the second reaction, Bax-dependent pore formation. Bax insertion occurred immediately upon Bax addition, prior to the end of the lag phase. Permeabilization kinetics were affected in a reciprocal manner by [cBid] and [Bax], confirming the “hit-and-run” hypothesis of cBid-induced direct Bax activation. Surprisingly, MOMP rate constants were linearly related to [Bax], implying that Bax acts non-cooperatively. Thus, the oligomeric catalyst is distinct from Bax. Moreover, contrary to common assumption, pore formation kinetics depend on Bax monomers, not oligomers. Catalyst formation exhibited a sharp transition in activation energy at ∼28°C, suggesting a role for membrane lipid packing. Furthermore, catalyst formation was strongly inhibited by chemical antagonists of the yeast mitochondrial fission protein, Dnm1. However, the mammalian ortholog, Drp1, was undetectable in mitochondrial outer membranes. Moreover, ATP and GTP were dispensable for MOMP. Thus, the data argue that oligomerization of a catalyst protein, distinct from Bax and Drp1, facilitates MOMP, possibly through a membrane-remodeling event.</p> </div

Bax-induced pore formation requires heat-labile protein(s) and is inhibited by mdivi-1 analogs.

<p>For heat-induced protein inactivation, OMVs were preincubated at 68°C for 10 min and equilibrated at room temperature. Dextran-fluorescein release induced by 100 nM Bax in the presence of 40 nM cBid was inhibited in heat-treated OMVs (A), but restored at higher (400–800 nM) Bax concentrations (B). OMVs were incubated with mdivi-1 analogs B and H for 5 min prior to the addition of Bax (C–E). Note that at 25°C, compound H (25–50 µM) completely inhibits dextran release (D); at 45°C, however, this compound produces merely a long lag phase without much effect on the rate of pore formation (E). Black lines, control OMVs; red lines, heat-treated OMVs; purple lines, OMVs treated with the mdivi-1 analogs. Arrows indicate additions of Bax; cBid was added 2–3 min prior to Bax. Data shown are representative of at least three independent experiments (see also <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.1001394#pbio.1001394.s006" target="_blank">Figure S6</a>).</p

cBid/Bax–dependent membrane permeabilization: native mitochondrial outer membranes and protein-free liposomes display different permeabilization kinetics, and native membranes respond to lower concentrations of Bax.

<p>Dextran release was measured continuously in rat liver OMVs (A–C) and defined liposomes (D). OMVs were loaded with either 10 kDa dextran-cascade blue (A,C) or 70 kDa dextran-fluorescein (B). Bax concentration was 25 nM (A,B) or as indicated (C,D). Arrows indicate additions of Bax and 0.05% Triton X-100. When present, cBid (40 nM) was added 2–3 min prior to Bax. Bcl-xL (1 µM) was added 2 min before Bax or at later points indicated by arrows (B; blue lines). (E) Respiration-based continuous measurements of Bax-induced MOMP in isolated rat liver mitochondria. Mitochondria were incubated in a KCl-based respiration buffer containing 0.3 µM FCCP, 2 µM myxothiazol (Myxo), 2 mM NADH, and 80 µM cytochrome c (CytC). Under these conditions, the rate of NADH-dependent respiration depends on the rate of cytochrome c permeation through the MOM and therefore reflects the kinetics of MOMP. Arrows indicate additions of myxothiazol, NADH, CytC, and Bax (60 nM) mixed with cBid (40 nM). Control trace (blue line): no cBid/Bax added. Data are plotted as the first derivative of oxygen concentration (i.e., respiration rate) versus time. The insert shows the original oxygen consumption curve, in the presence of cBid and Bax. Note that the kinetics of MOMP in whole mitochondria and OMVs are similarly biphasic. Data shown are representative of at least three independent experiments. See also <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.1001394#pbio.1001394.s001" target="_blank">Figure S1</a>.</p