Research

Life Sciences & Biotechnology

Title :	Elucidating the role of Cdk-related kinases (Crk7, Crk9, and Crk11) mediating transcript maturation in the asexual developmental stages of Toxoplasma gondii.
Area of research :	Life Sciences & Biotechnology
Principal Investigator :	Dr. Abhijit Subhashrao Deshmukh, National Institute Of Animal Biotechnology
Timeline Start Year :	2024
Timeline End Year :	2027

Details

Executive Summary :

Toxoplasma gondii is a protozoan parasite of humans and animals, causing life-threatening disease in the immunocompromised and fetal abnormalities. Central to the pathogenicity of this parasite is its ability to differentiate between rapidly replicating tachyzoites (acute stage) and the encysted bradyzoites (chronic stage) during asexual developmental stages. Transcriptional reprogramming plays a significant role in the transition from tachyzoite to bradyzoite, with hundreds of genes differentially expressed between the stages. Eukaryotic transcription is a functionally coupled process to ensure pre-mRNA maturation into mRNA. The production of mRNAs involves multiple catalytic activities such as transcription, capping, splicing, polyadenylation, cleavage, and export. These processes are primarily controlled by RNA Pol II (RNAP II)-associated complex. Phosphorylation of RNAP II carboxyl-terminal domain (RNAPII-CTD) plays a central role in regulating these activities. Protein kinases belonging to the CDK family called transcriptional CDKs (tCDKs 7-13, 18-20) phosphorylate RNAPII-CTD and coordinate multiple cotranscriptional events important for gene expression. Unlike humans (10 tCDKs), T. gondii has a limited number of transcriptional CDK-related kinases (3 Crks). In humans, tCDKs play largely non-redundant roles in coordinating transcription and RNA processing. Given that only three tCrks (Crk7, Crk9, and Crk11) are present in Toxoplasma, overlapping roles of these kinases may not be ruled out. Previously, we showed that Toxoplasma tCrks (Crk7 and Crk9) phosphorylate RNAP II CTD in synchrony with the transcription initiation and elongation cycle. These experiments used broadly selective kinase inhibitors and were limited to a few tachyzoite-specific genes. In addition, we observed that compared to mammalian RNAP II CTD, which contains 52 identical YSPTSPS heptad repeats, the Toxoplasma RNAP II CTD tail displays wide variation in length and composition. A limited number of tCrks and diverse RNAP II CTD, indicate Toxoplasma-specific transcription regulatory events. However, specific inhibitions of tCrks may help determine these kinases global role in transcript maturation. Moreover, the role of Crk11 in Toxoplasma is yet to be identified. The study objectives are i) To study the expression and localization of tCkrs (Crk7, -9, and -11) in the bradyzoite stage of Toxoplasma. ii) To examine the transcription-associated regulatory roles of Crk7 and Crk9 kinases in the asexual stages of Toxoplasma. iii) To study the role of Crk11 kinase in transcription stages and RNA processing events of Toxoplasma. iv) To investigate the regulatory roles of tCrks in transcription-linked RNA processing of Toxoplasma. This project will significantly add to the understanding of Toxoplasma tCrk-mediated regulation of transcript maturation required to achieve stage-specific expression of genes in a need-based and time-dependent fashion.