Research

Life Sciences & Biotechnology

Title :	Posttranscriptional regulation by PUF Proteins- FBF-1 and FBF-2 regulate cAMP and cGMP signaling in Caenorhabditis elegans
Area of research :	Life Sciences & Biotechnology
Principal Investigator :	Dr. Kavita Babu, Indian Institute Of Science, Bangalore, Karnataka
Timeline Start Year :	2024
Timeline End Year :	2027

Details

Executive Summary :

Secondary signaling involving cAMP and cGMP are crucial for spatiotemporal regulation of signaling mechanisms that are required for normal development of an organism, memory formation and synaptic plasticity among multiple other functions across most multicellular animals. Hence, these pathways must be tightly regulated at multiple levels. RNA binding proteins (RBPs) are a class of proteins that post-transcriptionally regulate gene expression by modulating the stability of the mRNA to which they bind. In this work, we explore the RBP-mediated post-transcriptional regulation of Phosphodiesterase 2 (PDE-2) and Protein Kinase (KIN-2), key molecules in the secondary signaling pathways through cAMP and cGMP across phyla. Using yeast-three-hybrid and luminescence assays we show that two RBP proteins belonging to the PUF protein family (FBF-1 and FBF-2) bind directly to the 3 UTR of PDE-2 and KIN-2. Furthermore, mutating the binding sites at the 3 UTR of PDE-2 and KIN-2, prevents PUF binding to the 3UTR of PDE-2 and KIN-2. We have gone on to characterize the FBF-1/2 binding domains in PDE-2 and show that mutating this domain using CRISPR technology leads to developmental defects in C. elegans. We also observe an increase in PDE-2 expressing through qPCR on studying the mutant lines that prevent FBF-1/2 binding. Taking this preliminary work forward, this grant aims to study in detail the interactions between FBF-1/2 and their targets. Further, we propose to understand the physiological relevance of these interactions using the free-living nematode, Caenorhabditis elegans, as a model organism. Our work will build upon the crucial role played by RBPs in regulating important signaling cascades and go on to implicate the interaction between PUF proteins, FBF-1 and FBF-2 and the 3 UTRs of their targets in multiple processes including development of the organism and normal functioning of neurons in the animal. This work will also allow us to study how FBF-1/2 binding to the 3 UTR of targets affects target gene expression (PDE-2 and KIN-2 in this case) and how inhibiting this binding using just three point-mutations at the FBF-1/2 binding site could give rise to defects in C. elegans development.