CDK-dependent nuclear localization of B-Cyclin Clb1 promotes FEAR activation during meiosis I in budding yeast

Cyclin-dependent kinases (CDK) are master regulators of the cell cycle in eukaryotes. CDK activity is regulated by the presence, post-translational modification and spatial localization of its regulatory subunit cyclin. In budding yeast, the B-cyclin Clb1 is phosphorylated and localizes to the nucleus during meiosis I. However the functional significance of Clb1's phosphorylation and nuclear localization and their mutual dependency is unknown. In this paper, we demonstrate that meiosis-specific phosphorylation of Clb1 requires its import to the nucleus but not vice versa. While Clb1 phosphorylation is dependent on activity of both CDK and polo-like kinase Cdc5, its nuclear localization requires CDK but not Cdc5 activity. Furthermore we show that increased nuclear localization of Clb1 during meiosis enhances activation of FEAR (Cdc Fourteen Early Anaphase Release) pathway. We discuss the significance of our results in relation to regulation of exit from meiosis I

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

Cellular Radiosensitivity: How much better do we understand it?

Purpose: Ionizing radiation exposure gives rise to a variety of lesions in DNA that result in genetic instability and potentially tumorigenesis or cell death. Radiation extends its effects on DNA by direct interaction or by radiolysis of H2O that generates free radicals or aqueous electrons capable of interacting with and causing indirect damage to DNA. While the various lesions arising in DNA after radiation exposure can contribute to the mutagenising effects of this agent, the potentially most damaging lesion is the DNA double strand break (DSB) that contributes to genome instability and/or cell death. Thus in many cases failure to recognise and/or repair this lesion determines the radiosensitivity status of the cell. DNA repair mechanisms including homologous recombination (HR) and non-homologous end-joining (NHEJ) have evolved to protect cells against DNA DSB. Mutations in proteins that constitute these repair pathways are characterised by radiosensitivity and genome instability. Defects in a number of these proteins also give rise to genetic disorders that feature not only genetic instability but also immunodeficiency, cancer predisposition, neurodegeneration and other pathologies. Conclusions: In the past fifty years our understanding of the cellular response to radiation damage has advanced enormously with insight being gained from a wide range of approaches extending from more basic early studies to the sophisticated approaches used today. In this review we discuss our current understanding of the impact of radiation on the cell and the organism gained from the array of past and present studies and attempt to provide an explanation for what it is that determines the response to radiation

The Ionizing Radiation-Induced Bystander Effect: Evidence, Mechanism, and Significance

It has long been considered that the important biological effects of ionizing radiation are a direct consequence of unrepaired or misrepaired DNA damage occurring in the irradiated cells. It was presumed that no effect would occur in cells in the population that receive no direct radiation exposure. However, in vitro evidence generated over the past two decades has indicated that non-targeted cells in irradiated cell cultures also experience significant biochemical and phenotypic changes that are often similar to those observed in the targeted cells. Further, nontargeted tissues in partial body-irradiated rodents also experienced stressful effects, including oxidative and oncogenic effects. This phenomenon, termed the “bystander response,” has been postulated to impact both the estimation of health risks of exposure to low doses/low fluences of ionizing radiation and the induction of second primary cancers following radiotherapy. Several mechanisms involving secreted soluble factors, oxidative metabolism, gap-junction intercellular communication, and DNA repair, have been proposed to regulate radiation-induced bystander effects. The latter mechanisms are major mediators of the system responses to ionizing radiation exposure, and our knowledge of the biochemical and molecular events involved in these processes is reviewed in this chapter

HIV-1 Promotes Intake of Leishmania Parasites by Enhancing Phosphatidylserine-Mediated, CD91/LRP-1-Dependent Phagocytosis in Human Macrophages

Over the past decade, the number of reported human immunodeficiency virus type-1 (HIV-1)/Leishmania co-infections has risen dramatically, particularly in regions where both diseases are endemic. Although it is known that HIV-1 infection leads to an increase in susceptibility to Leishmania infection and leishmaniasis relapse, little remains known on how HIV-1 contributes to Leishmania parasitaemia. Both pathogens infect human macrophages, and the intracellular growth of Leishmania is increased by HIV-1 in co-infected cultures. We now report that uninfected bystander cells, not macrophages productively infected with HIV-1, account for enhanced phagocytosis and higher multiplication of Leishmania parasites. This effect can be driven by HIV-1 Tat protein and transforming growth factor-beta (TGF-β). Furthermore, we show for the first time that HIV-1 infection increases surface expression of phosphatidylserine receptor CD91/LRP-1 on human macrophages, thereby leading to a Leishmania uptake by uninfected bystander cells in HIV-1-infected macrophage populations. The more important internalization of parasites is due to interactions between the scavenger receptor CD91/LRP-1 and phosphatidylserine residues exposed at the surface of Leishmania. We determined also that enhanced CD91/LRP-1 surface expression occurs rapidly following HIV-1 infection, and is triggered by the activation of extracellular TGF-β. Thus, these results establish an intricate link between HIV-1 infection, Tat, surface CD91/LRP-1, TGF-β, and enhanced Leishmania phosphatidylserine-mediated phagocytosis

TCR signal strength controls thymic differentiation of discrete proinflammatory gamma delta T cell subsets

The mouse thymus produces discrete gd T cell subsets that make either interferon-g (IFN-g) or interleukin 17 (IL-17), but the role of the T cell antigen receptor (TCR) in this developmental process remains controversial. Here we show that Cd3g+/− Cd3d+/− (CD3 double-haploinsufficient (CD3DH)) mice have reduced TCR expression and signaling strength on gd T cells. CD3DH mice had normal numbers and phenotypes of ab thymocyte subsets, but impaired differentiation of fetal Vg6+ (but not Vg4+) IL-17- producing gd T cells and a marked depletion of IFN-g-producing CD122+ NK1.1+ gd T cells throughout ontogeny. Adult CD3DH mice showed reduced peripheral IFN-g+ gd T cells and were resistant to experimental cerebral malaria. Thus, TCR signal strength within specific thymic developmental windows is a major determinant of the generation of proinflammatory gd T cell subsets and their impact on pathophysiology

The pathogenic mechanism of the Mycobacterium ulcerans virulence factor, mycolactone, depends on blockade of protein translocation into the ER.

Infection with Mycobacterium ulcerans is characterised by tissue necrosis and immunosuppression due to mycolactone, the necessary and sufficient virulence factor for Buruli ulcer disease pathology. Many of its effects are known to involve down-regulation of specific proteins implicated in important cellular processes, such as immune responses and cell adhesion. We have previously shown mycolactone completely blocks the production of LPS-dependent proinflammatory mediators post-transcriptionally. Using polysome profiling we now demonstrate conclusively that mycolactone does not prevent translation of TNF, IL-6 and Cox-2 mRNAs in macrophages. Instead, it inhibits the production of these, along with nearly all other (induced and constitutive) proteins that transit through the ER. This is due to a blockade of protein translocation and subsequent degradation of aberrantly located protein. Several lines of evidence support this transformative explanation of mycolactone function. First, cellular TNF and Cox-2 can be once more detected if the action of the 26S proteasome is inhibited concurrently. Second, restored protein is found in the cytosol, indicating an inability to translocate. Third, in vitro translation assays show mycolactone prevents the translocation of TNF and other proteins into the ER. This is specific as the insertion of tail-anchored proteins into the ER is unaffected showing that the ER remains structurally intact. Fourth, metabolic labelling reveals a near-complete loss of glycosylated and secreted proteins from treated cells, whereas cytosolic proteins are unaffected. Notably, the profound lack of glycosylated and secreted protein production is apparent in a range of different disease-relevant cell types. These studies provide a new mechanism underlying mycolactone's observed pathological activities both in vitro and in vivo. Mycolactone-dependent inhibition of protein translocation into the ER not only explains the deficit of innate cytokines, but also the loss of membrane receptors, adhesion molecules and T-cell cytokines that drive the aetiology of Buruli ulcer

H2AX phosphorylation at the sites of DNA double-strand breaks in cultivated mammalian cells and tissues

A sequence variant of histone H2A called H2AX is one of the key components of chromatin involved in DNA damage response induced by different genotoxic stresses. Phosphorylated H2AX (γH2AX) is rapidly concentrated in chromatin domains around DNA double-strand breaks (DSBs) after the action of ionizing radiation or chemical agents and at stalled replication forks during replication stress. γH2AX foci could be easily detected in cell nuclei using immunofluorescence microscopy that allows to use γH2AX as a quantitative marker of DSBs in various applications. H2AX is phosphorylated in situ by ATM, ATR, and DNA-PK kinases that have distinct roles in different pathways of DSB repair. The γH2AX serves as a docking site for the accumulation of DNA repair proteins, and after rejoining of DSBs, it is released from chromatin. The molecular mechanism of γH2AX dephosphorylation is not clear. It is complicated and requires the activity of different proteins including phosphatases and chromatin-remodeling complexes. In this review, we summarize recently published data concerning the mechanisms and kinetics of γH2AX loss in normal cells and tissues as well as in those deficient in ATM, DNA-PK, and DSB repair proteins activity. The results of the latest scientific research of the low-dose irradiation phenomenon are presented including the bystander effect and the adaptive response estimated by γH2AX detection in cells and tissues