Exploring diffuse radio emission in galaxy clusters and groups with the uGMRT and the SKA

Diffuse radio emission has been detected in a considerable number of galaxy clusters and groups, revealing the presence of pervasive cosmic magnetic fields, and of relativistic particles in the large-scale structure (LSS) of the Universe. Since cluster radio emission is faint and steep spectrum, its observations are largely limited by the instrument sensitivity and frequency of observation, leading to a dearth of information, more so for lower-mass systems. The unprecedented sensitivity of recently commissioned low-frequency radio telescope arrays, aided by the development of advanced calibration and imaging techniques, have helped in achieving unparalleled image quality. At the same time, the development of sophisticated numerical simulations and the availability of supercomputing facilities have paved the way for high-resolution numerical modeling of radio emission, and the structure of the cosmic magnetic fields in LSS, leading to predictions matching the capabilities of observational facilities. In view of these rapidly-evolving scenerio in modeling and observations, in this review, we summarise the role of the new telescope arrays and the development of advanced imaging techniques and discuss the detections of various kinds of cluster radio sources. In particular, we discuss observations of the cosmic web in the form of supercluster filaments, studies of emission in poor clusters and groups of galaxies, and of ultra-steep spectrum sources. We also review the current theoretical understanding of various diffuse cluster radio sources and the associated magnetic field and polarization. As the statistics of detections improve along with our theoretical understanding, we update the source classification schemes based on their intrinsic properties. We conclude by summarising the role of the upgraded GMRT and our expectations from the upcoming Square Kilometre Array (SKA) observatories.Comment: 32 pages, 10 figures, accepted for publication in the Journal of Astrophysics and Astronomy (JoAA) (to appear in the special issue on "Indian participation in the SKA"

The type III secretion system effector EspO of enterohaemorrhagic Escherichia coli inhibits apoptosis through an interaction with HAX-1

Many enteric pathogens employ a type III secretion system (T3SS) to translocate effector proteins directly into the host cell cytoplasm, where they subvert signalling pathways of the intestinal epithelium. Here, we report that the anti-apoptotic regulator HS1-associated protein X1 (HAX-1) is an interaction partner of the T3SS effectors EspO of enterohaemorrhagic Escherichia coli (EHEC) and Citrobacter rodentium, OspE of Shigella flexneri and Osp1STYM of Salmonella enterica serovar Typhimurium. EspO, OspE and Osp1STYM have previously been reported to interact with the focal adhesions protein integrin linked kinase (ILK). We found that EspO localizes both to the focal adhesions (ILK localisation) and mitochondria (HAX-1 localisation), and that increased expression of HAX-1 leads to enhanced mitochondrial localisation of EspO. Ectopic expression of EspO, OspE and Osp1STYM protects cells from apoptosis induced by staurosporine and tunicamycin. Depleting cells of HAX-1 indicates that the anti-apoptotic activity of EspO is HAX-1 dependent. Both HAX-1 and ILK were further confirmed as EspO1-interacting proteins during infection using T3SS-delivered EspO1. Using cell detachment as a proxy for cell death we confirmed that T3SS-delivered EspO1 could inhibit cell death induced during EPEC infection, to a similar extent as the anti-apoptotic effector NleH, or treatment with the pan caspase inhibitor z-VAD. In contrast, in cells lacking HAX-1, EspO1 was no longer able to protect against cell detachment, while NleH1 and z-VAD maintained their protective activity. Therefore, during both infection and ectopic expression EspO protects cells from cell death by interacting with HAX-1. These results suggest that despite the differences between EHEC, C. rodentium, Shigella and S. typhimurium infections, hijacking HAX-1 anti-apoptotic signalling is a common strategy to maintain the viability of infected cells

Heat shock protein 90 localizes to the surface and augments virulence factors of Cryptococcus neoformans

Background Thermotolerance is an essential attribute for pathogenesis of Cryptococcus as exemplified by the fact that only two species in the genus, which can grow at 37 degrees C, are human pathogens. Species which have other virulence factors including capsule formation and melanisation, but lack the ability to propagate at 37 degrees C are not pathogenic. In another related fungal pathogen, Candida albicans, heat shock protein 90 has been implicated to be a central player in commanding pathogenicity by governing yeast to hyphal transition and drug resistance. Exploring Hsp90 biology in Cryptococcus in context of thermotolerance may thus highlight important regulatory principles of virulence and open new therapeutic avenues. Methodology/Principal findings Hsp90 is involved in regulating thermotolerance in Cryptococcus as indicated by growth hypersensitivity at 37 degrees C upon mild compromise of Hsp90 function relative to 25 E C. Biochemical studies revealed a more potent inhibition of ATPase activity by pharmacological inhibitor 17-AAG at 37 degrees C as compared to 25 E C. Catalytic efficiency of the protein at 37 E C was found to be 6.39x10(-5)M(-1). Furthermore, indirect immunofluorescence analysis using a specific antibody revealed cell surface localization of Hsp90 via ER Golgi classical secretory pathway. Hsp90 was found to be induced under capsule inducing conditions and Hsp90 inhibition led to decrease in capsular volume. Finally compromising Hsp90 function improved anidulafungin tolerance in Cryptococcus. Conclusions/Significance Our findings highlight that Hsp90 regulates pathogenicity of the fungus by myriad ways. Firstly, it is involved in mediating thermotolerance which implies targeting Hsp90 can abrogate thermotolerance and hence growth of the fungus. Secondly, this study provides the first report of biochemical properties of Hsp90 of a pathogenic fungus. Finally, since Hsp90 is localised at the cell wall, targeting cell surface Hsp90 can represent a novel strategy to combat this lethal infection

Heat shock protein 90 localizes to the surface and augments virulence factors of <i>Cryptococcus neoformans</i>

<div><p>Background</p><p>Thermotolerance is an essential attribute for pathogenesis of <i>Cryptococcus</i> as exemplified by the fact that only two species in the genus, which can grow at 37°C, are human pathogens. Species which have other virulence factors including capsule formation and melanisation, but lack the ability to propagate at 37°C are not pathogenic. In another related fungal pathogen, <i>Candida albicans</i>, heat shock protein 90 has been implicated to be a central player in commanding pathogenicity by governing yeast to hyphal transition and drug resistance. Exploring Hsp90 biology in <i>Cryptococcus</i> in context of thermotolerance may thus highlight important regulatory principles of virulence and open new therapeutic avenues.</p><p>Methodology/Principal findings</p><p>Hsp90 is involved in regulating thermotolerance in <i>Cryptococcus</i> as indicated by growth hypersensitivity at 37°C upon mild compromise of Hsp90 function relative to 25°C. Biochemical studies revealed a more potent inhibition of ATPase activity by pharmacological inhibitor 17-AAG at 37°C as compared to 25°C. Catalytic efficiency of the protein at 37°C was found to be 6.39×10⁻⁵μM^-1. Furthermore, indirect immunofluorescence analysis using a specific antibody revealed cell surface localization of Hsp90 via ER Golgi classical secretory pathway. Hsp90 was found to be induced under capsule inducing conditions and Hsp90 inhibition led to decrease in capsular volume. Finally compromising Hsp90 function improved anidulafungin tolerance in <i>Cryptococcus</i>.</p><p>Conclusions/Significance</p><p>Our findings highlight that Hsp90 regulates pathogenicity of the fungus by myriad ways. Firstly, it is involved in mediating thermotolerance which implies targeting Hsp90 can abrogate thermotolerance and hence growth of the fungus. Secondly, this study provides the first report of biochemical properties of Hsp90 of a pathogenic fungus. Finally, since Hsp90 is localised at the cell wall, targeting cell surface Hsp90 can represent a novel strategy to combat this lethal infection.</p></div

Binding of Hsp90 inhibitor 17-AAG and its effect on CnHsp90 ATPase activity.

<p>(a) Binding affinity of competitive inhibitor 17-AAG to CnHsp90 using tryptophan fluorescence was determined. Change in intrinsic fluorescence intensity upon ligand binding was plotted against ligand concentration. Dissociation constant, K_d, for 17-AAG binding was calculated to be 12.92 μM. (b) IC₅₀ for inhibition of ATPase activity of CnHsp90 was determined by incubating the pure protein with fixed, saturating concentration of ATP and varying concentrations of 17-AAG. The reaction was carried out at 25°C and 37°C. The percent activity remaining was plotted against concentration of 17-AAG in logarithmic scale to obtain the inhibition curve. 17-AAG mediates inhibition of CnHsp90 activity at both temperatures tested. (c) IC₅₀ values obtained at 25°C and 37°C for CnHsp90 inhibition by 17-AAG was found to be 26.89 μM and 117.15 μM respectively. Therefore, 17-AAG inhibits CnHsp90 more potently at 37°C.</p

Hsp90 is involved in induction and maintenance of capsule in <i>C</i>. <i>neoformans</i>.

<p>(a)Western blot analysis to probe levels of Hsp90 at 25°C and 37°C indicate similar expression levels at both temperatures tested. (b) Hsp90 is significantly upregulated under capsule inducing conditions. (c) Representative microscopic images showing compromise of Hsp90 function during or after capsule induction leads to drastic compromise in capsular size. (d) Hsp90 is involved in capsule assembly as well as maintenance around the cell wall as pre-and post-treatment with RAD leads to approximately 60% decrease in capsular volume relative to no drug and vehicle control.</p

Comparison of biochemical properties of Hsp90 from <i>C</i>. <i>neoformans</i> and comparison with Hsp90 from other organisms [3,6,24,29].

<p>Comparison of biochemical properties of Hsp90 from <i>C</i>. <i>neoformans</i> and comparison with Hsp90 from other organisms [<a href="http://www.plosntds.org/article/info:doi/10.1371/journal.pntd.0005836#pntd.0005836.ref003" target="_blank">3</a>,<a href="http://www.plosntds.org/article/info:doi/10.1371/journal.pntd.0005836#pntd.0005836.ref006" target="_blank">6</a>,<a href="http://www.plosntds.org/article/info:doi/10.1371/journal.pntd.0005836#pntd.0005836.ref024" target="_blank">24</a>,<a href="http://www.plosntds.org/article/info:doi/10.1371/journal.pntd.0005836#pntd.0005836.ref029" target="_blank">29</a>].</p

Hsp90 is associated with the fungal cell wall.

<p>(a) Exponential phase cells were grown at 25°C and 37°C and subjected to indirect immunofluorescence without permeabilization using specific antibody raised against CnHsp90. Hsp90 was found to be localized at the cell surface. Colocalization of CFW (chitin staining) and FITC signal (Hsp90 staining) is also observed indicating cell wall localization of CnHsp90. Antibody to cytosolic protein PGK and pre-immune sera served as negative controls for the experiments. b) Association of Hsp90 at the cell surface was also probed by whole cell based ELISA wherein plates were incubated with intact yeast cells. Significant binding of CnHsp90 Ab to cell surface Hsp90 was seen at both 25°C and 37°C. Anti PGK Ab was used as a negative control. c) Immunoblot analysis of <i>C</i>. <i>neoformans</i> βME cell surface extract also detected a band corresponding to Hsp90 further confirming the presence of Hsp90 in the cell wall fraction.</p

Hsp90 governs thermotolerance in <i>C</i>. <i>neoformans</i>.

<p>(a) Cells were grown in the presence of Hsp90 inhibitor RAD at various concentrations ranging from 5nM to 100 μM at the temperatures indicated above. Comparison of MIC₅₀ values for RAD at 25°C and 37°C clearly indicates that <i>C</i>. <i>neoformans</i> growth is hypersensitive to Hsp90 inhibition at 37°C. (b) Spot agar assay without inhibitors indicate that Hsp90 inhibition is fungicidal at 37°C. Growth is not seen when cells grown in presence of RAD at 37°C were spotted and incubated at 25°C and 37°C. (c) Comparison of MIC₅₀ values for antifungal drug Amphotericin B shows similar susceptibilities at both temperatures ruling out lower MIC value due to enhanced drug uptake at higher temperature. (d) <i>C</i>. <i>neoformans</i> clinical isolate shows 12.4-fold hypersensitivity to Hsp90 inhibition at 37°C. (e) <i>C</i>. <i>neoformans</i> clinical isolate show similar susceptibility to AmB at both temperatures tested ruling out nonspecific effect. (f) Comparison of MIC₅₀ values for RAD in <i>Candida albicans</i> at 25°C and 37°C clearly indicates that <i>C</i>. <i>albicans</i> growth is not hypersensitive to Hsp90 inhibition at 37°C.</p

Cell surface association of Hsp90 depends on ER Golgi classical secretory pathway.

<p>Indirect immunofluorescence was carried out using Anti CnHsp90 Ab after treatment with different inhibitors. In each case first panel corresponds to cells grown at 25°C and second panel represents cells grown at 37°C. (a) Indirect Immunofluorescence using specific antibody show surface localization of Hsp90 at both temperatures. (b) Cells were treated with vesicle trafficking inhibitor BFA and immunoflourescence was carried out as above. Complete loss of surface signal for Hsp90 was seen in the treated cells both at 25°C (first panel) and 37°C (second panel). (c) Disulfide blocker NEM also abrogated Hsp90 localisation at cell surface. At 25°C diffused signal is seen whereas at 37°C there is no signal thereby implicating the involvement of ER Golgi classical secretion pathway in the process. (d) Hsp90 may be involved in chaperoning a client protein which gets localised to the cell wall. To test this hypothesis, RAD was used to abrogate client association with Hsp90. Loss of cell surface localization was seen in case of RAD treated cells indicating pharmacological inhibitor can also interfere with the process of Hsp90 transport to cell surface.</p