Biology subjects

Chakrabarti, K.

Publications and source records attributed to Chakrabarti, K..

4 recordsLinked to original sources

bioRxiv2025

Nucleotide-resolution Mapping of RNA N6-Methyladenosine (m6A) modifications and comprehensive analysis of global polyadenylation events in mRNA 3' end processing in malaria pathogen Plasmodium falciparum

Plasmodium falciparum is an obligate human parasite of the phylum Apicomplexa and is the causative agent of the most lethal form of human malaria. Although N6-methyladenosine modification is thought to be one of the major post-transcriptional regulatory mechanisms for stage-specific gene expression in apicomplexan parasites, the precise base position of m6A in mRNAs or noncoding RNAs in these parasites remains unknown. Here, we report global nucleotide-resolution mapping of m6A residues in P. falciparum using DART-seq technology, which quantitatively displayed a stage-specific, dynamic distribution pattern with enrichment near mRNA 3 ends. In this process we identified 894, 788, and 1,762 m6A-modified genes in Ring, Trophozoite and Schizont stages respectively, with an average of 5-7 m6A sites per-transcript at the individual gene level. Notably, several genes involved in malaria pathophysiology, such as KAHRP, ETRAMPs, SERA and stress response genes, such as members of Heat Shock Protein (HSP) family are highly enriched in m6A and therefore could be regulated by this RNA modification. Since we observed preferential methylation at the 3 ends of P. falciparum transcripts and because malaria polyadenylation specificity factor PfCPSF30 harbors an m6A reader YTH domain, we reasoned that m6A might play an important role in 3-end processing of malaria mRNAs. To investigate this, we used two complementary high-throughput RNA 3-end mapping approaches, which provided an initial framework to explore potential roles of m6A in the regulation of alternative polyadenylation (APA) during malaria development in human hosts.

genomics↗

bioRxiv2023

Bacterial deamidases modulate ubiquitin structure and dynamics to dysregulate ubiquitin signaling

The deamidases secreted by Burkholderia pseudomallei and Enteropathogenic Ecoli modify the residue 40 in ubiquitin from a Glutamine (Q) to Glutamate (E), triggering several downstream processes to cause cell cycle arrest and activate immune responses. Deamidation hampers the activity of ubiquitin and its interaction with ubiquitin chain receptors by an unknown mechanism. Here, we study the effect of deamidation on ubiquitin structure and dynamics. We report the crystal structure of the deamidated ubiquitin, supported by NMR and molecular dynamics simulations. The structure reveals a new intra-molecular salt bridge between the deamidated region and the C-terminal tail of ubiquitin. The salt bridge perturbs the dynamics of the ubiquitin tail to reduce affinity for ubiquitin receptors like the p62 ubiquitin-associated domain. The salt bridge disrupts the transition to catalytically active E2~Ub closed conformation. Consequently, RING E3s fail to interact with E2~Ub, reducing ubiquitination activity. Our findings reveal that deamidation-induced intramolecular salt bridges in ubiquitin modulate conformational ensembles to inactivate ubiquitination.

biochemistry↗

bioRxiv2022

Proteomic Analysis Defines the Interactome of Telomerase in the Protozoan Parasite, Trypanosoma brucei

Telomerase is a ribonucleoprotein enzyme responsible for maintaining the telomeric end of the chromosome. The telomerase enzyme requires two main components to function: the telomerase reverse transcriptase (TERT) and the telomerase RNA (TR), which provides the template for telomeric DNA synthesis. TR is a long noncoding RNA, which forms the basis of a large structural scaffold upon which many accessory proteins can bind and form the complete telomerase holoenzyme. These accessory protein interactions are required for telomerase activity and regulation inside cells. The interacting partners of TERT have been well studied in yeast, human, and Tetrahymena models, but not in lower eukaryotes, including clinically relevant human parasites. Here, using the protozoan parasite, Trypanosoma brucei (T. brucei) as a model, we have identified the interactome of T. brucei TERT (TbTERT) using a mass spectrometry-based approach. We identified previously known and unknown interacting factors of TbTERT, highlighting unique features of T. brucei telomerase biology. These unique interactions with TbTERT, suggest mechanistic differences in telomere maintenance between T. brucei and other eukaryotes.

molecular biology↗

bioRxiv2020

The RNA Structurome in the Asexual Blood Stages of Malaria Pathogen Plasmodium falciparum

RNA as an effector of biological functions often adopts secondary and tertiary structural folds. Plasmodium falciparum is a deadly human pathogen responsible for the devastating disease called malaria. In this study, we measured the differential accumulation of RNA secondary structures in coding and noncoding transcripts from the asexual developmental cycle in P. falciparum in human red blood cells. Our comprehensive analysis, combining high-throughput nuclease mapping of RNA structures by duplex RNA-seq, immunoaffinity purification and RNA analysis, collectively measured differentially base-paired RNA regions during the parasite development. Our mapping data not only aligned to a diverse pool of RNAs with known structures but also enabled us to identify new structural RNA regions in the malaria genome. On average, ~71% of the genes with secondary structures are found to be protein coding mRNAs. Mapping pattern of these base-paired RNAs corresponded to all parts of protein-coding mRNAs, including 5 UTR, CDS and 3 UTR. In addition to histone family genes which are known to form secondary structures in their mRNAs, transcripts from genes which are important for transcriptional and post-transcriptional control, such as unique plant-like transcription factor family, ApiAP2, DNA/RNA binding protein family, Alba, ribosomal proteins and eukaryotic initiation factors involved in translational control and the ones important for RBC invasion and cytoadherence also show strong accumulation of duplex RNA reads in various asexual stages. Intriguingly, our study determined a positive relationship between mRNA structural contents and translation efficiency in P. falciparum asexual blood stages, suggesting an essential role of RNA structural changes in malaria gene expression programs.

molecular biology↗