Heavy metals and metalloids such as cadmium [Cd(II)] and arsenic [As(III)] are widespread environmental toxicants responsible for multiple adverse health effects in humans. However, the molecular mechanisms underlying metal-induced cytotoxicity and carcinogenesis, as well as the detoxification and tolerance pathways, are incompletely understood. Here, we use global fitness profiling by barcode sequencing to quantitatively survey the Schizosaccharomyces pombe haploid deletome for genes that confer tolerance of cadmium or arsenic. We identified 106 genes required for cadmium resistance and 110 genes required for arsenic resistance, with a highly significant overlap of 36 genes. A subset of these 36 genes account for almost all proteins required for incorporating sulfur into the cysteine-rich glutathione and phytochelatin peptides that chelate cadmium and arsenic. A requirement for Mms19 is explained by its role in directing iron–sulfur cluster assembly into sulfite reductase as opposed to promoting DNA repair, as DNA damage response genes were not enriched among those required for cadmium or arsenic tolerance. Ubiquinone, siroheme, and pyridoxal 5′-phosphate biosynthesis were also identified as critical for Cd/As tolerance. Arsenic-specific pathways included prefoldin-mediated assembly of unfolded proteins and protein targeting to the peroxisome, whereas cadmium-specific pathways included plasma membrane and vacuolar transporters, as well as Spt–Ada–Gcn5-acetyltransferase (SAGA) transcriptional coactivator that controls expression of key genes required for cadmium tolerance. Notable differences are apparent with corresponding screens in the budding yeast Saccharomyces cerevisiae, underscoring the utility of analyzing toxic metal defense mechanisms in both organisms.

Alzheimer’s disease (AD) is a major cause of age-related dementia and is characterized by progressive brain damage that gradually destroys memory and the ability to learn, which ultimately leads to the decline of a patient’s ability to perform daily activities. Although some of the pharmacological treatments of AD are available for symptomatic relief, they are not able to limit the progression of AD and have several side effects. Mesenchymal stem/stromal cells (MSCs) could be a potential therapeutic option for treating AD due to their immunomodulatory, anti-inflammatory, regenerative, antioxidant, anti-apoptotic, and neuroprotective effects. MSCs not only secret neuroprotective and anti-inflammatory factors to promote the survival of neurons, but they also transfer functional mitochondria and miRNAs to boost their bioenergetic profile as well as improve microglial clearance of accumulated protein aggregates. This review focuses on different clinical and preclinical studies using MSC as a therapy for treating AD, their outcomes, limitations and the strategies to potentiate their clinical translation.

Transcription is the most fundamental step in gene expression in any living organism. Various environmental cues help in the maturation of core RNA polymerase (RNAP; α(2)ββ'ω) with different σ-factors, leading to the directed recruitment of RNAP to different promoter DNA sequences. Thus it is essential to determine the σ-factors that affect the preferential partitioning of core RNAP among various σ-actors, and the role of σ-switching in transcriptional gene regulation. Further, the macromolecular assembly of holo RNAP takes place in an extremely crowded environment within a cell, and thus far the kinetics and thermodynamics of this molecular recognition process have not been well addressed. In this study we used a site-directed bioaffinity immobilization method to evaluate the relative binding affinities of three different Escherichia coli σ-factors to the same core RNAP with variations in temperature and ionic strength while emulating the crowded cellular milieu. Our data indicate that the interaction of core RNAP-σ is susceptible to changes in external stimuli such as osmolytic and thermal stress, and the degree of susceptibility varies among different σ-factors. This allows for a reversible σ-switching from housekeeping factors to alternate σ-factors when the organism senses a change in its physiological conditions.

Sequence specific interaction between DNA and protein molecules has been a subject of active investigation for decades now. Here, we have chosen single promoter containing bacteriophage DeltaD(III) T7 DNA and Escherichia coli RNA polymerase and followed their recognition at the air-water interface by using the surface plasmon resonance (SPR) technique, where the movement of one of the reacting species is restricted by way of arraying them on an immobilized support. For the Langmuir monolayer studies, we used a RNA polymerase with a histidine tag attached to one of its subunits, thus making it an excellent substrate for Ni(II) ions, while the SPR studies were done using biotin-labeled DNA immobilized on a streptavidin-coated chip. Detailed analysis of the thermodynamic parameters as a function of concentration and temperature revealed that the interaction of RNA polymerase with T7 DNA is largely entropy driven (83 (+/-12) kcal mol(-1)) with a positive enthalpy of 13.6 (+/-3.6) kcal mol(-1). The free energy of reaction determined by SPR and Langmuir-Blodgett technique was -11 (+/-2) and -15.6 kcal mol(-1), respectively. The ability of these methods to retain the specificity of the recognition process was also established.

At the heart of understanding cellular processes lies our ability to explore the specific nature of communication between sequential information carrying biopolymers. However, the data extracted from conventional solution phase studies may not reflect the dynamics of communication between recognized partners as they occur in the crowded cellular milieu. We use the principle of immobilization of histidine-tagged biopolymers at a Ni(II)-encoded Langmuir monolayer to study sequence-specific protein-protein interactions in an artificially crowded environment. The advantage of this technique lies in increasing the surface density of one of the interacting partners that allows us to study macromolecular interactions in a controlled crowded environment, but without compromising the speed of the reactions. We have taken advantage of this technique to follow the sequential assembly process of the multiprotein complex Escherichia coli RNA polymerase at the interface and also deciphered the role of one of the proteins, omega (ω), in the assembly pathway. Our reconstitution studies indicate that in the absence of molecular chaperones or other cofactors, omega (ω) plays a decisive role in refolding the largest protein beta prime (β') and its recruitment into the multimeric assembly to reconstitute an active RNA polymerase. It was also observed that the monolayer had the ability to distinguish between sequence-specific and -nonspecific interactions despite the immobilization of one of the biomacromolecules. The technique provides a universal two-dimensional template for studying protein-ligand interactions while mimicking molecular crowding.