Lysine acetylation of the Mycobacterium tuberculosis HU protein modulates its DNA binding and genome organization

Nucleoid‐associated protein HU, a conserved protein across eubacteria is necessary for maintaining the nucleoid organization and global regulation of gene expression. Mycobacterium tuberculosis HU (MtHU) is distinct from the other orthologues having 114 amino acid long carboxyl terminal extensions with a high degree of sequence similarity to eukaryotic histones. In this study, we demonstrate that the DNA binding property of MtHU is regulated by posttranslational modifications akin to eukaryotic histones. MtHU purified from M. tuberculosis cells is found to be acetylated on multiple lysine residues unlike the E. coli expressed recombinant protein. Using coimmunoprecipitation assay, we identified Eis as one of the acetyl transferases that interacts with MtHU and modifies it. Although Eis is known to acetylate aminoglycosides, the kinetics of acetylation showed that its protein acetylation activity on MtHU is robust. In vitro Eis modified MtHU at various lysine residues, primarily those located at the carboxyl terminal domain. Acetylation of MtHU caused reduced DNA interaction and alteration in DNA compaction ability of the NAP. Over‐expression of the Eis leads to hyperacetylation of HU and decompaction of genome. These results provide first insights into the modulation of the nucleoid structure by lysine acetylation in bacteria.     

The Plasmodium translocon of exported proteins component EXP2 is critical for establishing a patent malaria infection in mice

Export of most malaria proteins into the erythrocyte cytosol requires the Plasmodium translocon of exported proteins (PTEX) and a cleavable Plasmodium export element (PEXEL). In contrast, the contribution of PTEX in the liver stages and export of liver stage proteins is unknown. Here, using the FLP/FRT conditional mutatagenesis system, we generate transgenic Plasmodium berghei parasites deficient in EXP2, the putative pore‐forming component of PTEX. Our data reveal that EXP2 is important for parasite growth in the liver and critical for parasite transition to the blood, with parasites impaired in their ability to generate a patent blood‐stage infection. Surprisingly, whilst parasites expressing a functional PTEX machinery can efficiently export a PEXEL‐bearing GFP reporter into the erythrocyte cytosol during a blood stage infection, this same reporter aggregates in large accumulations within the confines of the parasitophorous vacuole membrane during hepatocyte growth. Notably HSP101, the putative molecular motor of PTEX, could not be detected during the early liver stages of infection, which may explain why direct protein translocation of this soluble PEXEL‐bearing reporter or indeed native PEXEL proteins into the hepatocyte cytosol has not been observed. This suggests that PTEX function may not be conserved between the blood and liver stages of malaria infection.     

Conservation of the C-type lectin fold for accommodating massive sequence variation in archaeal diversity-generating retroelements

Background

Diversity-generating retroelements (DGRs) provide organisms with a unique means for adaptation to a dynamic environment through massive protein sequence variation. The potential scope of this variation exceeds that of the vertebrate adaptive immune system. DGRs were known to exist only in viruses and bacteria until their recent discovery in archaea belonging to the ‘microbial dark matter’, specifically in organisms closely related to Nanoarchaeota. However, Nanoarchaeota DGR variable proteins were unassignable to known protein folds and apparently unrelated to characterized DGR variable proteins.

Results

To address the issue of how Nanoarchaeota DGR variable proteins accommodate massive sequence variation, we determined the 2.52 Å resolution limit crystal structure of one such protein, AvpA, which revealed a C-type lectin (CLec)-fold that organizes a putative ligand-binding site that is capable of accommodating 10¹³ sequences. This fold is surprisingly reminiscent of the CLec-folds of viral and bacterial DGR variable protein, but differs sufficiently to define a new CLec-fold subclass, which is consistent with early divergence between bacterial and archaeal DGRs. The structure also enabled identification of a group of AvpA-like proteins in multiple putative DGRs from uncultivated archaea. These variable proteins may aid Nanoarchaeota and these uncultivated archaea in symbiotic relationships.

Conclusions

Our results have uncovered the widespread conservation of the CLec-fold in viruses, bacteria, and archaea for accommodating massive sequence variation. In addition, to our knowledge, this is the first report of an archaeal CLec-fold protein.

     

Functional characterization of recombinant human granulocyte colony stimulating factor (hGMCSF) immobilized onto silica nanoparticles

Objectives

Granulocyte macrophage colony stimulating factor (GMCSF), an important therapeutic cytokine, was immobilized onto silica nanoparticles. Maintenance of structural integrity and biological performance in immobilized cytokine was assessed to augment its applicability in possible biomedical implications.

Results

Following its cloning and expression in E. coli, the recombinant human GMCSF (hGMCSF) was purified as a GST-tagged protein corresponding to a 42 kDa band on SDS-PAGE. The purified cytokine was immobilized onto biocompatible silica nanoparticles (~129.4 nm) by adsorption and the binding was confirmed by dynamic light scattering and infrared spectroscopy. Maximum binding of hGMCSF was at 6.4 µg mg^?1 silica nanoparticles. Efficient release of the cytokine from the nanoparticles with its structural integrity intact was deduced from circular dichroism spectroscopy. hGMCSF-immobilized silica nanoparticles efficiently increased the proliferation of RAW 264.7 macrophage cells with 50 % increase in proliferation at 600 ng hGMCSF µg^?1 silica nanoparticles.

Conclusions

Silica nanoparticles successfully immobilized hGMCSF maintaining its structural integrity. The release of the immobilized cytokine from silica nanoparticles resulted in the increased proliferation of macrophages indicating the potential of the system in future applications.

     

Naringin and Sertraline Ameliorate Doxorubicin-Induced Behavioral Deficits Through Modulation of Serotonin Level and Mitochondrial Complexes Protection Pathway in Rat Hippocampus

The present study was designed to investigate the neuroprotective effect of naringin (NR) alone as well as its combination with sertraline (SRT) against doxorubicin (DOX)-induced neurobehavioral and neurochemical anomalies. DOX (15 mg/kg; i.p.) administration caused behavioral alterations, oxidative stress, neuroinflammation, mitochondrial dysfunction and monoamines alteration in male Wistar rats. NR (50 and 100 mg/kg; i.p.) and SRT (5 mg/kg; i.p.) treatment significantly attenuated DOX-induced anxiety and depressive-like behavior as evident from elevated plus maze (EPM) and modified forced swimming test (mFST), respectively. NR treatment significantly attenuated DOX-induced raised plasma corticosterone (CORT), tumor necrosis factor-alpha (TNF-α) and interleukin-1 beta (IL-1β) levels in the hippocampus (HC). Furthermore, we found that combination of NR and SRT regimen ameliorated DOX-induced behavioral anomalies through modulation of the 5-HT level and mitochondrial complexes protection pathway along with alleviation of oxidative stress in the HC region. Therefore, NR treatment alone or in combination with SRT could be beneficial against DOX-induced neurotoxicity.     

Glycation of human serum albumin alters its binding efficacy towards the dietary polyphenols: a comparative approach

Diabetes is a major problem in the world. The proteins became modified during glycation after reacting with the reducing sugars (e.g. D-glucose) via non-enzymatic pathways. The glycated analogue of human serum albumin (HSA) has been characterized with the help of multi-spectroscopic methods. It has been observed that six glucose molecules can bind covalently to HSA under experimental condition. The binding affinity of the modified HSA towards the dietary polyphenols has been estimated using UV–vis and fluorescence spectroscopic techniques. The binding constant values of the ligands were found to decrease after the modification of HSA.     

A tale of two paralogs: human Transformer2 proteins with differential RNA-binding affinities

The Transformer2 (Tra2) proteins in humans are homologues of the Drosophila Tra2 protein. One of the two RNA-binding paralogs, Tra2β, has been very well-studied over the past decade, but not much is known about Tra2α. It was very recently shown that the two proteins demonstrate the phenomenon of paralog compensation. Here, we provide a structural basis for this genetic backup circuit, using molecular modelling and dynamics studies. We show that the two proteins display similar binding specificities, but differential affinities to a short GAA-rich RNA stretch. Starting from the 6-nucleotide RNA in the solution structure, close to 4000 virtual mutations were modelled on RNA and the domain–RNA interactions were studied after energy minimisation to convergence. Separately, another known 13-nucleotide stretch was docked and the domain–RNA interactions were observed through a 100-ns dynamics trajectory. We have also demonstrated the ‘compensatory’ mechanism at the level of domains in one of the domain repeat-containing RNA-binding proteins.     

Arsenic mitigation in soil-plant system through zinc application in West Bengal soils

The study area of the experiment was selected on the basis of a preliminary field survey and laboratory detection of accumulation, accompanied by biomagnification, of arsenic in the water-soil-plant continuum. A laboratory incubation study was conducted with soil samples from the selected areas of West Bengal with arsenic-contaminated groundwater, and from a low arsenic zone. These soils were treated with different concentrations of zinc and arsenic and incubated for 10 and 25 days. The findings from the incubation study indicate the decrement of arsenic release in the soil solution of the given soils on application of zinc, which tends to bind arsenic electrostatically in the soil matrix, thereby helping to mitigate the toxicity of arsenic to some extent in the soil-plant system. The arsenic-zinc interactions were further revalidated in the arsenic-affected fields where a similar type of moderating effect (as in the incubation study) was observed in rice-rice cropping sequence. Arsenic-zinc interactions were critically analyzed through a number of accumulation, translocation, and mobility parameters, considering soil to plant uptake in different plant parts.     

Exosomes in carcinogenesis: molecular palkis carry signals for the regulation of cancer progression and metastasis

Exosomes, which act as biological cargo vessels, are cell-released, phospholipid-enclosed vesicles. In eukaryotic cells, exosomes carry and exchange biological materials or signals for the benefit or detriment to the cells. Thereby, we consider exosomes to be molecular Palkis (carriers). Although exosomes are currently one of the most popularly researched cellular entities, they have remained largely enigmatic and warrant continued investigation into their structure and functions. These membraned vesicles are between 30 and 150 nm in diameter and are actively secreted by all cell types. While initially considered cellular “trash bags,” recent years have revealed exosomes to be dynamic and multi-functional vesicles that may play a crucial role in cancer development, progression and metastasis. Thereby, they have the potential to be used in development of therapeutic modalities for cancer and other diseases. As more research studies emerge, it’s becoming evident that exosomes are released by cells with a purpose and are representatives of certain cell types and disease conditions. Hence, they may also be used as biomarkers for the detection of cancer initiation, progression and organotropic metastatic growth of cancer cells. This review will focus on the recent developments achieved in identifying the role of exosomes in cancer development and progression as well as therapeutic implications. The review will also discuss the pitfalls of methodologies used for the extraction of exosomes.