Biology subjects

Vyas, P.

Publications and source records attributed to Vyas, P..

23 records · Page 2Linked to original sources

bioRxiv2020

Leptospira surface protein LigA plays a multifaceted role in modulating the host innate immune response

Leptospira, a zoonotic pathogen, is known to infect various hosts and can establish persistent infection. This remarkable ability of bacteria is attributed to its potential to modulate the host immune response by exploiting its surface proteins. We have identified and characterized the domain of the variable region of Leptospira immunoglobulin-like protein A (LAV) involved in immune modulation. The 11th domain (A11) of the variable region of LigA (LAV) induces a strong TLR4 dependent innate response leading to subsequent induction of humoral and cellular immune responses in mice. A11 is also involved in acquiring complement regulator FH and binds to host protease Plasminogen (PLG), there by mediating functional activity to escape from complement-mediated killing. The deletion of A11 domain significantly impaired TLR4 signaling and subsequent reduction in the innate and adaptive immune response. It also inhibited the binding of FH and PLG thereby mediating killing of bacteria. Our study discovered an unprecedented role of LAV as a nuclease capable of degrading Neutrophil Extracellular Traps (NETs). This nuclease activity was primarily mediated by A11. These results highlighted the moonlighting function of LigA and demonstrated that a single domain of a surface protein is involved in evading a myriad of host innate immune defenses, which might allow the persistence of Leptospira in different hosts for a long term without clearance.

immunology↗

bioRxiv2020

Helicase-Like Functions in Phosphate Loop Containing Beta-Alpha Polypeptides

The P-loop Walker A motif underlies hundreds of essential enzyme families that bind nucleotide triphosphates (NTPs) and mediate phosphoryl transfer (P-loop NTPases), including the earliest DNA/RNA helicases, translocases and recombinases. What were the primordial precursors of these enzymes? Could these large and complex proteins emerge from simple polypeptides? Previously, we showed that P-loops embedded in simple {beta} repeat proteins bind NTPs, but also, unexpectedly so, ssDNA and RNA. Here, we extend beyond the purely biophysical function of ligand binding to demonstrate rudimentary helicase-like activities. We further constructed simple 40-residue polypeptides comprising just one {beta}-(P-loop)- element. Despite their simplicity, these P-loop prototypes confer functions such as strand separation and exchange. Foremost, these polypeptides unwind dsDNA, and upon addition of NTPs, or inorganic polyphosphates, release the bound ssDNA strands to allow reformation of dsDNA. Binding kinetics and low-resolution structural analyses indicate that activity is mediated by oligomeric forms spanning from dimers to high-order assemblies. The latter are reminiscent of extant P-loop recombinases such as RecA. Overall, these P-loop prototypes comprise a plausible description of the sequence, structure and function of the earliest P-loop NTPases. They also indicate that multifunctionality and dynamic assembly were key in endowing short polypeptides with elaborate, evolutionarily relevant functions. Significance statementIt is widely assumed that todays large and complex proteins emerged from much shorter and simpler polypeptides. Yet the nature of these early precursors remains enigmatic. We describe polypeptides that contain one of the earliest protein motifs, a phosphate-binding loop, or P-loop, embedded in a single beta-alpha element. These P-loop prototypes show intriguing characteristics of a primordial world comprised of nucleic acids and peptides. They are generalists capable of binding different phospho-ligands, including inorganic polyphosphates and single-stranded DNA. Nonetheless, in promoting double-stranded DNA unwinding and strand-exchange they resemble modern P-loop helicases and recombinases. Our study describes a missing link in the evolution of complex proteins - simple polypeptides that tangibly relate to contemporary P-loop enzymes in sequence, structure and function.

molecular biology↗

bioRxiv2020

On the Emergence of P-Loop NTPase and Rossmann Enzymes from a Beta-Alpha-Beta Ancestral Fragment

Dating back to the last universal common ancestor (LUCA), the P-loop NTPases and Rossmanns now comprise the most ubiquitous and diverse enzyme lineages. Intriguing similarities in their overall architecture and phosphate binding motifs suggest common ancestry; however, due to a lack of sequence identity and some fundamental structural differences, these families are considered independent emergences. To address this longstanding dichotomy, we systematically searched for bridge proteins with structure and sequence elements shared by both lineages. We detected homologous segments that span the first {beta}{beta} segment of both lineages and include two key functional motifs: (i) a phosphate binding loop - the Walker A motif of P-loop NTPases or the Rossmann equivalent, both residing at the N-terminus of 1; and (ii) an Asp at the tip of {beta}2. The latter comprises the Walker B aspartate that chelates the catalytic metal in P-loop NTPases, or the canonical Rossmann {beta}2-Asp that binds the cofactors ribose moiety. Tubulin, a Rossmann GTPase, demonstrates the potential of the {beta}2-Asp to take either one of these two roles. We conclude that common P-loops/Rossmann ancestry is plausible, although convergence cannot be completely ruled out. Regardless, both lineages most likely emerged from a polypeptide comprising a {beta}{beta} segment carrying the above two functional motifs, a segment that comprises the core of both enzyme families to this very day.

evolutionary biology↗

bioRxiv2019

Oncogenic Gata1 causes stage-specific megakaryocyte differentiation delay

The megakaryocyte/erythroid Transient Myeloproliferative Disorder (TMD) in newborns with Down Syndrome (DS) occurs when N-terminal truncating mutations of the hemopoietic transcription factor GATA1, that produce GATA1short protein (GATA1s), are acquired early in development. Prior work has shown that murine GATA1s, by itself, causes a transient yolk sac myeloproliferative disorder. However, it is unclear where in the hemopoietic cellular hierarchy GATA1s exerts its effects to produce this myeloproliferative state. Here, through a detailed examination of hemopoiesis from murine GATA1s ES cells and GATA1s embryos we define defects in erythroid and megakaryocytic differentiation that occur relatively in hemopoiesis. GATA1s causes an arrest late in erythroid differentiation in vivo, and even more profoundly in ES-cell derived cultures, with a marked reduction of Ter-119 cells and reduced erythroid gene expression. In megakaryopoiesis, GATA1s causes a differentiation delay at a specific stage, with accumulation of immature, kit-expressing CD41hi megakaryocytic cells. In this specific megakaryocytic compartment, there are increased numbers of GATA1s cells in S-phase of cell cycle and reduced number of apoptotic cells compared to GATA1 cells in the same cell compartment. There is also a delay in maturation of these immature GATA1s megakaryocytic lineage cells compared to GATA1 cells at the same stage of differentiation. Finally, even when GATA1s megakaryocytic cells mature, they mature aberrantly with altered megakaryocyte-specific gene expression and activity of the mature megakaryocyte enzyme, acetylcholinesterase. These studies pinpoint the hemopoietic compartment where GATA1s megakaryocyte myeloproliferation occurs, defining where molecular studies should now be focussed to understand the oncogenic action of GATA1s.\n\nScientific CategoryHaematopoiesis and Stem Cells\n\nKey PointsGATA1s-induced stage-specific differentiation delay increases immature megakaryocytes in vivo and in vitro, during development.\n\nDifferentiation delay is associated with increased numbers of cells in S-phase and reduced apoptosis.

cell biology↗

bioRxiv2018

DOT1L inhibition reveals a distinct class ofenhancers dependent on H3K79 methylation

Enhancer elements are a key regulatory feature of many important genes. Several general features including the presence of specific histone modifications are used to identify and subcategorize enhancers. Here we identify a distinct subset of enhancers in leukemia cells that are functionally dependent upon H3K79me3. Using the DOT1L inhibitor, EPZ-5676, we show that loss of H3K79me3 at these H3K79me3 enhancer elements (KEEs) leads to reduced chromatin accessibility, histone acetylation and transcription factor binding. We then use Capture-C, a high-resolution chromosome conformation capture technique, to show that H3K79me3 is required for KEE interactions with the promoter as well as transcription of the associated genes. Together these data implicate H3K79me3 in having a functional role at a subset of active enhancers where it helps maintain histone acetylation and chromatin accessibility, potentially by promoting phase-separated condensates.

molecular biology↗