PUF partner interactions at a conserved interface shape the RNA binding landscape and cell fate

Protein-RNA regulatory networks underpin much of biology. C. elegans FBF-2, a PUF RNA-binding protein, binds over 1000 RNAs to govern stem cells and differentiation. FBF-2 interacts with multiple protein partners via a key tyrosine, Y479. Here we investigate the in vivo significance of partnerships using a Y479A mutant. Y479A occupancy increases or decreases at specific sites across the transcriptome, varying with the RNA, and shifts from 3'UTRs to other regions. Germline development also changes in a very specific fashion: Y479A abolishes one FBF-2 function – the sperm to oocyte cell fate switch. Y479A changes to regulation of one mRNA, gld-1, are critical to this fate change, though other network changes are also important. FBF-2 switches from repression to activation of gld-1 RNA, likely by distinct FBF-2 partnerships. The role of partnerships in governing RNA regulatory networks will extend broadly, including for human health and disease, as protein partnerships pervade RNA control. Overall design: Enhanced crosslinking and immunoprecipitation (eCLIP) followed by sequencing was used to determine the occupancy of wild-type FBF-2 and FBF-2(Y479A) on RNA targets in the germline of C. elegans. 4 biological replicates of each genotype were obtained (IP samples) as well as their corresponding size-matched input control (SMI).