Costa Dos Santos, S. J. | Search bioRxiv

bioRxiv2024

The PUF RNA-binding protein, FBF-2, maintains stem cells without binding to RNA

Like all canonical PUF proteins, C. elegans FBF-2 binds to specific RNAs via tripartite recognition motifs (TRMs). Here we report that an FBF-2 mutant protein that cannot bind to RNA, is nonetheless biologically active and maintains stem cells. This unexpected result challenges the conventional wisdom that RBPs must bind to RNAs to achieve biological activity. Also unexpectedly, FBF-2 interactions with partner proteins can compensate for loss of RNA-binding. FBF-2 only loses biological activity when its RNA-binding and partner interactions are both defective. These findings highlight the complementary contributions of RNA-binding and protein partner interactions to activity of an RNA-binding protein.

developmental biology↗

bioRxiv2023

Functional significance of PUF partnerships in C. elegans germline stem cells

PUF RNA-binding proteins are conserved stem cell regulators. Four PUF proteins govern self-renewal of C. elegans germline stem cells together with two intrinsically disordered proteins, LST-1 and SYGL-1. Based on yeast two-hybrid results, we proposed a composite self-renewal hub in the stem cell regulatory network, with eight PUF partnerships and extensive redundancy. Here, we investigate LST-1-PUF and SYGL-1-PUF partnerships and their molecular activities in their natural context - nematode stem cells. We confirm LST-1-PUF partnerships and their specificity to self-renewal PUFs by co-immunoprecipitation and show that an LST-1(AmBm) mutant defective for PUF-interacting motifs does not complex with PUFs in nematodes. LST-1(AmBm) is used to explore the functional significance of the LST-1-PUF partnership. Tethered LST-1 requires the partnership to repress expression of a reporter RNA, and LST-1 requires the partnership to co-immunoprecipitate with NTL-1/Not1 of the CCR4-NOT complex. We suggest that the partnership provides multiple molecular interactions that work together to form an effector complex on PUF target RNAs. Comparison of PUF-LST-1 and Pumilio-Nanos reveals fundamental molecular differences, making PUF-LST-1 a distinct paradigm for PUF partnerships. Summary statementPartnerships between PUF RNA-binding proteins and intrinsically disordered proteins are essential for stem cell maintenance and RNA repression.

developmental biology↗