Glutathione, an unusual tripeptide, is the most abundant small molecular weight thiol compound of living cells. Discovered more than 125 years ago, it plays many vital roles that includes its function as the cells redox buffer. The degradation of glutathione was first described in 1952 with the discovery of the g-glutamyl- transpeptidase enzyme. For more than 5 decades, this enzyme, which acts on non-cytosolic pools of glutathione was thought to be the only enzyme involved in glutathione degradation in living cells. Recent studies from our lab have thrown fresh light on glutathione degradation with the discovery of alternate enzymes for glutathione degradation. This has also led to a better understanding on the roles of glutathione degradation. One role that will be discussed in detail here is the role in calcium signalling. Yeasts have two channels that pump calcium into the cytoplasm, Cch1p and Yvc1p. Yvc1p is a yeast vacuolar TRP channel, while Cch1p is a plasma membrane L-type channel. Yvc1p and Cch1p werefound to be activated by redox through glutathionylation. The uncovering of the mechanisms by which these signalling pathways are regulated by redox has led to new insights on the role of glutathione degradation in cell signalling.
Feb 20, 2017
Prof. Dipankar Chatterjee, MBU, IISc., Bangalore
Venue & Time: C V Raman Hall at 3-30 PM to 5-00 PM
Second Messengers (p)ppGppand Cyclic di-GMP mediated regulation of Cell Shape, Cell Division and Antibiotic Sensitivity in Mycobacterium smegmatis
Quorum Sensing, a cell to cell communication phenomenon, is involved in modulating the social behavior of bacteria. Nucleotide based second messengers like (p)ppGpp and c-di-GMP are known to regulate such communication in mycobacteria. (p)ppGpp is synthesized by bacteria to face any kind of stress; while the signalling nucleotide c-di-GMP is synthesized principally to switch from motile (planktonic) to sessile (biofilm) life style. We investigated the effect of disrupting (p)ppGpp and c-di-GMP signalling on the antibiotic sensitivity in M. smegmatis. Using Phenotype Microarray (PM) technology, the growth of Δreland ΔdcpAknock out strains was compared to those of the wild-type and respective complemented strains in 240 different antimicrobials. It was found that the knockout mutants displayed enhanced survival in the presence of multiple antibiotics. The PM data was corroborated by the independent determination of minimum inhibitory concentrations of seven different antibiotics. Microscopy analyses revealed that the Δrel and ΔdcpA strains are elongated, multinucleate and multiseptate in M. smegmatis. The higher levels of (p)ppGpp and c-di-GMP caused M. Smegmatisassume coccoid morphology. The overproduction of (p)ppGpp and c-di-GMP, achieved through overexpression of Rel and DcpA proteins, encased the overexpression strains relOE and dcpAOE in a biofilm like matrix.
Molecular Insights into Meiotic Chromosome Synapsis from Single Molecule Analysis
In humans, defects in meiotic chromosome synapsis and segregation results in aneuploidy leading to miscarriages and genetic disorders. Aneuploidy occurs with much higher probability for the female counterpart, and increases dramatically with age. An evolutionarily conserved meiosis-specific proteinaceous structure, the synaptonemal complex (SC), is required for normal synapsis of meiotic chromosomes. However, very little is known about the biochemical properties of SC components or the mechanisms underlying their roles in meiotic chromosome synapsis and recombination. Structural and functional analysis of Saccharomyces cerevisiae Hop1, a key structural component of SC, has begun to reveal important insights into its function in the synapsis of meiotic chromosomes. Toward this end, we showed that Hop1 is a structure-specific DNA-binding protein, displays higher binding affinity for G-quadruplex DNA and the Holliday junction, and causes structural distortion of the latter at the core of the junction. Using atomic force microscopy and magnetic tweezers techniques, we discovered that Hop1 exhibits the ability to bridge non-contiguous DNA segments into intramolecular stem-loop structures in which the DNA segments are fully synapsed within the filamentous protein stems. Additional evidence suggested that Hop1 folds DNA into rigid protein-DNA filaments and higher-order nucleoprotein structures. Importantly, Hop1 promotes robust intra- and intermolecular synapsis between double-stranded DNA molecules, suggesting that juxtaposition of DNA sequences may assist in strand exchange between homologs by recombination-associated proteins. Furthermore, evidence from in vitro and in vivo studies disclosed the existence of G-quadruplex and i-motif structures at meiosis-specific recombination hot spots. Taken together, these studies support the notion that understanding of meiotic chromosome synapsis and recombination must consider protein binding interactions in conjunction with DNA structural motifs at meiosis-specific recombination hotspots.
Feb 6, 2017
Prof. B J Rao, TIFR, Mumbai
Venue & Time: C V Raman Hall, 6th Feb, 2017 at 4:00 PM
Local biochemical problems and non-local solutions in biological designs: some of our recent take-homes
It is a recurring theme that local biochemical reactions often require large-scale systemic non-local changes in cells and tissues, the design of which is not easily intuitive. We explore this theme using DNA Damage Response (DDR) paradigm in normal human cells and Drosophila. We uncover that damaged chromosomes traverse large distances reversibly to repair damaged DNA. Damaged replication forks collapse if the repair remains persistently active where the repair has inbuilt non-local design to attenuate itself. Interestingly, un-repaired dying cells seem to trigger changes across large distances in the tissue-field to provoke additional cellular proliferation, “compensating for death and thereby renewing life”. I illustrate these with examples and argue that most of these biological regulations are based on the “system as a whole” rather than “the parts thereof” based regulation.