Geranylgeranoic acid (GGA) originally was identified in some animals and has been developed as an agent for preventing second primary hepatoma. We previously have also identified GGA as an acyclic diterpenoid in some medicinal herbs. Recently, we reported that in human hepatoma-derived HuH-7 cells, GGA is metabolically labeled from ¹³C-mevalonate. Several cell-free experiments have demonstrated that GGA is synthesized through geranylgeranial by oxygen-dependent oxidation of geranylgeraniol (GGOH), but the exact biochemical events giving rise to GGA in hepatoma cells remain unclear. Monoamine oxidase B (MOAB) has been suggested to be involved in GGOH oxidation. Here, using two human hepatoma cell lines, we investigated whether MAOB contributes to GGA biosynthesis. Using either HuH-7 cell lysates or recombinant human MAOB, we found that: 1) the MAO inhibitor tranylcypromine dose-dependently downregulates endogenous GGA levels in HuH-7 cells; and 2) siRNA-mediated MAOB silencing reduces intracellular GGA levels in HuH-7 and Hep3B cells. Unexpectedly, however, CRISPR/Cas9-generated MAOB-KO human hepatoma Hep3B cells had GGA levels similar to those in MAOB-WT cells. A sensitivity of GGA levels to siRNA-mediated MAOB downregulation was recovered when the MAOB-KO cells were transfected with a MAOB-expression plasmid, suggesting that MAOB is the enzyme primarily responsible for GGOH oxidation and that some other latent metabolic pathways may maintain endogenous GGA levels in the MAOB-KO hepatoma cells. Along with the previous findings, these results provide critical insights into the biological roles of human MAOB and provide evidence that hepatic MAOB is involved in endogenous GGA biosynthesis via GGOH oxidation.

Many clinical studies and epidemiological investigations have clearly demonstrated that women are twice as likely to develop cholesterol gallstones as men, and oral contraceptives and other estrogen therapies dramatically increase that risk. Further, animal studies have revealed that estrogen promotes cholesterol gallstone formation through the estrogen receptor (ER) α, but not ERβ, pathway. More importantly, some genetic and pathophysiological studies have found that G protein-coupled estrogen receptor (GPER) 1 is a new gallstone gene, Lith18, on chromosome 5 in mice and produces additional lithogenic actions, working independently of ERα, to markedly increase cholelithogenesis in female mice. Based on computational modeling of GPER, a novel series of GPER-selective antagonists were designed, synthesized, and subsequently assessed for their therapeutic effects via calcium mobilization, cAMP, and ERα and ERβ fluorescence polarization binding assays. From this series of compounds, one new compound, 2-cyclohexyl-4-isopropyl-N-(4-methoxybenzyl)aniline (CIMBA), exhibits superior antagonism and selectivity exclusively for GPER. Furthermore, CIMBA reduces the formation of 17β-estradiol-induced gallstones in a dose-dependent manner in ovariectomized mice fed a lithogenic diet for 8 weeks. At 32 μg/day/kg CIMBA, no gallstones are found, even in ovariectomized ERα (^–/–) mice treated with 6 μg/day 17β-estradiol and fed the lithogenic diet for 8 weeks. In conclusion, CIMBA treatment protects against the formation of estrogen-induced cholesterol gallstones by inhibiting the GPER signaling pathway in female mice. CIMBA may thus be a new agent for effectively treating cholesterol gallstone disease in women.­

The formation and properties of liquid-ordered (Lo) lipid domains (rafts) in the plasma membrane are still poorly understood. This limits our ability to manipulate ordered lipid domain-dependent biological functions. Giant plasma membrane vesicles (GPMVs) undergo large-scale phase separations into coexisting Lo and liquid-disordered lipid domains. However, large-scale phase separation in GPMVs detected by light microscopy is observed only at low temperatures. Comparing Förster resonance energy transfer-detected versus light microscopy-detected domain formation, we found that nanodomains, domains of nanometer size, persist at temperatures up to 20°C higher than large-scale phases, up to physiologic temperature. The persistence of nanodomains at higher temperatures is consistent with previously reported theoretical calculations. To investigate the sensitivity of nanodomains to lipid composition, GPMVs were prepared from mammalian cells in which sterol, phospholipid, or sphingolipid composition in the plasma membrane outer leaflet had been altered by cyclodextrin-catalyzed lipid exchange. Lipid substitutions that stabilize or destabilize ordered domain formation in artificial lipid vesicles had a similar effect on the thermal stability of nanodomains and large-scale phase separation in GPMVs, with nanodomains persisting at higher temperatures than large-scale phases for a wide range of lipid compositions. This indicates that it is likely that plasma membrane nanodomains can form under physiologic conditions more readily than large-scale phase separation. We also conclude that membrane lipid substitutions carried out in intact cells are able to modulate the propensity of plasma membranes to form ordered domains. This implies lipid substitutions can be used to alter biological processes dependent upon ordered domains.

The autosomal dominant disorder Schnyder corneal dystrophy (SCD) is caused by mutations in UbiA prenyltransferase domain-containing protein-1 (UBIAD1), which uses geranylgeranyl pyrophosphate (GGpp) to synthesize the vitamin K₂ subtype menaquinone-4 (MK-4). SCD is characterized by opacification of the cornea, owing to aberrant build-up of cholesterol in the tissue. We previously discovered that sterols stimulate association of UBIAD1 with ER-localized HMG-CoA reductase, which catalyzes a rate-limiting step in the synthesis of cholesterol and nonsterol isoprenoids, including GGpp. Binding to UBIAD1 inhibits sterol-accelerated ER-associated degradation (ERAD) of reductase and permits continued synthesis of GGpp in cholesterol-replete cells. GGpp disrupts UBIAD1-reductase binding and thereby allows for maximal ERAD of reductase as well as ER-to-Golgi translocation of UBIAD1. SCD-associated UBIAD1 is refractory to GGpp-mediated dissociation from reductase and remains sequestered in the ER to inhibit ERAD. Here, we report development of a biochemical assay for UBIAD1-mediated synthesis of MK-4 in isolated membranes and intact cells. Using this assay, we compared enzymatic activity of WT UBIAD1 with that of SCD-associated variants. Our studies revealed that SCD-associated UBIAD1 exhibited reduced MK-4 synthetic activity, which may result from its reduced affinity for GGpp. Sequestration in the ER protects SCD-associated UBIAD1 from autophagy and allows intracellular accumulation of the mutant protein, which amplifies the inhibitory effect on reductase ERAD. These findings have important implications not only for the understanding of SCD etiology but also for the efficacy of cholesterol-lowering statin therapy, which becomes limited, in part, because of UBIAD1-mediated inhibition of reductase ERAD.

Activation of microglia and astrocytes secondary to inflammatory processes contributes to the development and perpetuation of pain with a neuropathic phenotype. This pain state presents as a chronic debilitating condition and affects a large population of patients with conditions like rheumatoid arthritis and diabetes, or after surgery, trauma, or chemotherapy. Here, we review the regulation of lipid rafts in glial cells and the role they play as a key component of neuroinflammatory sensitization of central pain signaling pathways. In this context, we introduce the concept of an inflammaraft (i-raft), enlarged lipid rafts harboring activated receptors and adaptor molecules and serving as an organizing platform to initiate inflammatory signaling and the cellular response. Characteristics of the inflammaraft include increased relative abundance of lipid rafts in inflammatory cells, increased content of cholesterol per raft, and increased levels of inflammatory receptors, such as toll-like receptor (TLR)4, adaptor molecules, ion channels, and enzymes in lipid rafts. This inflammaraft motif serves an important role in the membrane assembly of protein complexes, for example, TLR4 dimerization. Operating within this framework, we demonstrate the involvement of inflammatory receptors, redox molecules, and ion channels in the inflammaraft formation and the regulation of cholesterol and sphingolipid metabolism in the inflammaraft maintenance and disruption. Strategies for targeting inflammarafts, without affecting the integrity of lipid rafts in noninflammatory cells, may lead to developing novel therapies for neuropathic pain states and other neuroinflammatory conditions.

Lipid rafts are small, dynamic membrane areas characterized by the clustering of selected membrane lipids as the result of the spontaneous separation of glycolipids, sphingolipids, and cholesterol in a liquid-ordered phase. The exact dynamics underlying phase separation of membrane lipids in the complex biological membranes are still not fully understood. Nevertheless, alterations in the membrane lipid composition affect the lateral organization of molecules belonging to lipid rafts. Neural lipid rafts are found in brain cells, including neurons, astrocytes, and microglia, and are characterized by a high enrichment of specific lipids depending on the cell type. These lipid rafts seem to organize and determine the function of multiprotein complexes involved in several aspects of signal transduction, thus regulating the homeostasis of the brain. The progressive decline of brain performance along with physiological aging is at least in part associated with alterations in the composition and structure of neural lipid rafts. In addition, neurodegenerative conditions, such as lysosomal storage disorders, multiple sclerosis, and Parkinson’s, Huntington’s, and Alzheimer’s diseases, are frequently characterized by dysregulated lipid metabolism, which in turn affects the structure of lipid rafts. Several events underlying the pathogenesis of these diseases appear to depend on the altered composition of lipid rafts. Thus, the structure and function of lipid rafts play a central role in the pathogenesis of many common neurodegenerative diseases.

Cholesterol/sphingolipid-rich membrane domains, known as lipid rafts or membrane rafts, play a critical role in the compartmentalization of signaling pathways. Physical segregation of proteins in lipid rafts may modulate the accessibility of proteins to regulatory or effector molecules. Thus, lipid rafts serve as sorting platforms and hubs for signal transduction proteins. Cancer cells contain higher levels of intracellular cholesterol and lipid rafts than their normal non-tumorigenic counterparts. Many signal transduction processes involved in cancer development (insulin-like growth factor system and phosphatidylinositol 3-kinase-AKT) and metastasis [cluster of differentiation (CD)44] are dependent on or modulated by lipid rafts. Additional proteins playing an important role in several malignant cancers (e.g., transmembrane glycoprotein mucin 1) are also being detected in association with lipid rafts, suggesting a major role of lipid rafts in tumor progression. Conversely, lipid rafts also serve as scaffolds for the recruitment and clustering of Fas/CD95 death receptors and downstream signaling molecules leading to cell death-promoting raft platforms. The partition of death receptors and downstream signaling molecules in aggregated lipid rafts has led to the formation of the so-called cluster of apoptotic signaling molecule-enriched rafts, or CASMER, which leads to apoptosis amplification and can be pharmacologically modulated. These death-promoting rafts can be viewed as a linchpin from which apoptotic signals are launched. In this review, we discuss the involvement of lipid rafts in major signaling processes in cancer cells, including cell survival, cell death, and metastasis, and we consider the potential of lipid raft modulation as a promising target in cancer therapy.

Expert

Expert

Expert

Expert

Expert

Expert

Expert

Expert

Expert

Expert

Expert

Expert

Expert

Expert

Expert

Expert

Expert

Expert

Expert

Expert

Cytochrome c oxidase (CcO) reduces O2 to water, coupled with a proton-pumping process. The structure of the O2-reduction site of CcO contains two reducing equivalents, Fea32+ and CuB1+, and suggests that a peroxide-bound state (Fea33+–O−–O−–CuB2+) rather than an O2-bound state (Fea32+–O2) is the initial catalytic intermediate. Unexpectedly, however, resonance Raman spectroscopy results have shown that the initial intermediate is Fea32+–O2, whereas Fea33+–O−–O−–CuB2+ is undetectable. Based on X-ray structures of static noncatalytic CcO forms and mutation analyses for bovine CcO, a proton-pumping mechanism has been proposed. It involves a proton-conducting pathway (the H-pathway) comprising a tandem hydrogen-bond network and a water channel located between the N- and P-side surfaces. However, a system for unidirectional proton-transport has not been experimentally identified. Here, an essentially identical X-ray structure for the two catalytic intermediates (P and F) of bovine CcO was determined at 1.8 Å resolution. A 1.70 Å Fe–O distance of the ferryl center could best be described as Fea34+ = O2−, not as Fea34+–OH−. The distance suggests an ∼800-cm−1 Raman stretching band. We found an interstitial water molecule that could trigger a rapid proton-coupled electron transfer from tyrosine-OH to the slowly forming Fea33+–O−–O−–CuB2+ state, preventing its detection, consistent with the unexpected Raman results. The H-pathway structures of both intermediates indicated that during proton-pumping from the hydrogen-bond network to the P-side, a transmembrane helix closes the water channel connecting the N-side with the hydrogen-bond network, facilitating unidirectional proton-pumping during the P-to-F transition.

Optic atrophy 1 (OPA1) is a dynamin protein that mediates mitochondrial fusion at the inner membrane. OPA1 is also necessary for maintaining the cristae and thus essential for supporting cellular energetics. OPA1 exists as membrane-anchored long form (L-OPA1) and short form (S-OPA1) that lacks the transmembrane region and is generated by cleavage of L-OPA1. Mitochondrial dysfunction and cellular stresses activate the inner membrane–associated zinc metallopeptidase OMA1 that cleaves L-OPA1, causing S-OPA1 accumulation. The prevailing notion has been that L-OPA1 is the functional form, whereas S-OPA1 is an inactive cleavage product in mammals, and that stress-induced OPA1 cleavage causes mitochondrial fragmentation and sensitizes cells to death. However, S-OPA1 contains all functional domains of dynamin proteins, suggesting that it has a physiological role. Indeed, we recently demonstrated that S-OPA1 can maintain cristae and energetics through its GTPase activity, despite lacking fusion activity. Here, applying oxidant insult that induces OPA1 cleavage, we show that cells unable to generate S-OPA1 are more sensitive to this stress under obligatory respiratory conditions, leading to necrotic death. These findings indicate that L-OPA1 and S-OPA1 differ in maintaining mitochondrial function. Mechanistically, we found that cells that exclusively express L-OPA1 generate more superoxide and are more sensitive to Ca2+-induced mitochondrial permeability transition, suggesting that S-OPA1, and not L-OPA1, protects against cellular stress. Importantly, silencing of OMA1 expression increased oxidant-induced cell death, indicating that stress-induced OPA1 cleavage supports cell survival. Our findings suggest that S-OPA1 generation by OPA1 cleavage is a survival mechanism in stressed cells.

Site-specific arrest of RNA polymerases (RNAPs) is fundamental to several technologies that assess RNA structure and function. Current in vitro transcription “roadblocking” approaches inhibit transcription elongation by blocking RNAP with a protein bound to the DNA template. One limitation of protein-mediated transcription roadblocking is that it requires inclusion of a protein factor extrinsic to the minimal in vitro transcription reaction. In this work, we developed a chemical approach for halting transcription by Escherichia coli RNAP. We first established a sequence-independent method for site-specific incorporation of chemical lesions into dsDNA templates by sequential PCR and translesion synthesis. We then show that interrupting the transcribed DNA strand with an internal desthiobiotin-triethylene glycol modification or 1,N6-etheno-2'-deoxyadenosine base efficiently and stably halts Escherichia coli RNAP transcription. By encoding an intrinsic stall site within the template DNA, our chemical transcription roadblocking approach enables display of nascent RNA molecules from RNAP in a minimal in vitro transcription reaction.

The arrangement of functionally-related genes in operons is a fundamental element of how genetic information is organized in prokaryotes. This organization ensures coordinated gene expression by co-transcription. Often, however, alternative genetic responses to specific stress conditions demand the discoordination of operon expression. During cold temperature stress, accumulation of the gene encoding the sole Asp–Glu–Ala–Asp (DEAD)-box RNA helicase in Synechocystis sp. PCC 6803, crhR (slr0083), increases 15-fold. Here, we show that crhR is expressed from a dicistronic operon with the methylthiotransferase rimO/miaB (slr0082) gene, followed by rapid processing of the operon transcript into two monocistronic mRNAs. This cleavage event is required for and results in destabilization of the rimO transcript. Results from secondary structure modeling and analysis of RNase E cleavage of the rimO–crhR transcript in vitro suggested that CrhR plays a role in enhancing the rate of the processing in an auto-regulatory manner. Moreover, two putative small RNAs are generated from additional processing, degradation, or both of the rimO transcript. These results suggest a role for the bacterial RNA helicase CrhR in RNase E-dependent mRNA processing in Synechocystis and expand the known range of organisms possessing small RNAs derived from processing of mRNA transcripts.

K. D. Cherednichenko, Yu. Yu. Ershova, A. V. Kiselev and S. N. Naboko
Trans. Moscow Math. Soc. 80 (2020), 251-294.
Abstract, references and article information

A. M. Savchuk and I. V. Sadovnichaya
Trans. Moscow Math. Soc. 80 (2020), 235-250.
Abstract, references and article information

I. A. Sheipak and T. A. Garmanova
Trans. Moscow Math. Soc. 80 (2020), 189-210.
Abstract, references and article information

V. V. Vlasov and N. A. Rautian
Trans. Moscow Math. Soc. 80 (2020), 169-188.
Abstract, references and article information

N. F. Valeev, Ya. T. Sultanaev and É. A. Nazirova
Trans. Moscow Math. Soc. 80 (2020), 153-167.
Abstract, references and article information

K. A. Mirzoev and T. A. Safonova
Trans. Moscow Math. Soc. 80 (2020), 133-151.
Abstract, references and article information

V. A. Sadovnichii, Ya. T. Sultanaev and A. M. Akhtyamov
Trans. Moscow Math. Soc. 80 (2020), 123-131.
Abstract, references and article information

Trans. Moscow Math. Soc. 80 (2020), 113-122.
Abstract, references and article information

A. G. Sergeev and Kh. A. Khachatryan
Trans. Moscow Math. Soc. 80 (2020), 95-111.
Abstract, references and article information

S. A. Kashchenko
Trans. Moscow Math. Soc. 80 (2020), 53-71.
Abstract, references and article information

S. A. Nazarov
Trans. Moscow Math. Soc. 80 (2020), 1-51.
Abstract, references and article information

Exame poderá detectar a doença na fase inicial. Pesquisadores da Faculdade de Medicina Osteopática da Universidade de Rowan, nos Estados Unidos, afirmam que estão perto de desenvolver um exame de sangue para detectar Alzheimer com precisão, o que dar...

The post Teste de sangue para detectar Alzheimer está próximo da realidade appeared first on Saúde Próspera.

Com que roupa eu vou? Todos os dias nós, mulheres, enfrentamos o dilema de decidir o que vestir. Veja algumas dicas para lhe ajudar a escolher qual roupa usar… A…

The post Com que roupa eu vou? Dicas de qual roupa usar appeared first on Saúde Próspera.

A. Knightly and C. Li
Memoirs of the AMS 224 (2012).
Abstract, references and article information

Publication Date: Apr 10 2020 Yes. According to the SEC, once an ETF becomes eligible to operate under Rule 6c-11 and elects to list on Nasdaq under Nasdaq Rule 5704, the existing order related to that fund (and all other funds under that exemptive order) will be rescinded. Once a fund is listed under Nasdaq Rule 5704, it will not be able to relist under Nasdaq Rule 5705(b) (Index Fund Shares) or Nasdaq Rule 5735 (Managed Fund Shares) unless a new exemptive relief order is obtained from the SEC....

Publication Date: May 4 2020 On May 1, 2020, Nasdaq adopted Rule 5636T, operative through, and including, June 30, 2020, to provide listed companies with a temporary exception from certain shareholder approval requirements. A Company must submit an application to Nasdaq’s Listing Qualifications Department demonstrating that the transaction satisfies the requirements in Rule 5636T and must provide the Notification Form: Listing of Additional Shares (“LAS Form”) required by...

Web hosting behemoth GoDaddy just filed a data breach notification with the US state of California.