Within the realm of gene regulation, the spotlight has increasingly turned to covalent nucleotide modifications such as N6-methyladenosine (m6A). m6A is the predominant internal mRNA modification in eukaryotes and it adds a crucial layer of post-transcriptional gene regulation in plants as well as in animals. m6A is intricately connected to growth and development in eukaryotes through the regulation of mRNA fate. The main functional impact of m6A is mediated through the recruitment of m6A readers, of which the most prominent group features a YT521-homology (YTH) domain for the specific recognition of m6A. In conjunction with their YTH-domain, most YTH-domain proteins exhibit a long intrinsically disordered region (IDR) at the N-terminus. Despite evidence from animals pointing to the IDR as the primary effector region, the exact mechanisms governing its regulatory functions in plants remain elusive. The model flowering plant Arabidopsis thaliana features 13 YTHdomain proteins known as EVOLUTIONARILY CONSERVED C-TERMINAL REGION (ECT). Among them, only ECT2, ECT3 and ECT4 have been studied; however, their molecular functions remain poorly understood.
During my PhD thesis, I have functionally dissected the N-terminal IDR of ECT2 through a series of deletion mutants, thereby identifying short linear motifs required for ECT2 function in vivo. More specifically, I found that a tyrosine-rich motif directly interacts with cytoplasmic poly(A)-binding proteins (PABPs) to stimulate primed stem cell proliferation. The results from this project led us to propose that, under conditions of limited PABP availability, ECT2-PABP interaction is necessary for efficient PABP-binding of mRNAs. This mechanism is particularly important for a subset of ECT2/ECT3/ECT4 target mRNAs essential for leaf formation. I also found that ECTs cooperate with another class of RNA-binding proteins, ACETYLATION LOWERS BINDING AFFINITY (ALBAs), to bind m6A-mRNAs and stimulate growth. On the side, one of the ECT2 deletion mutants that I engineered was used by a collaborating group in Spain to demonstrate a role of YTH-domain proteins as direct effectors of immunity of a plant positive-strand RNA virus. Thanks to this project, we have provided evidence of a novel branch of basal antiviral immunity operating in plants. Finally, we performed a systematic comparison of YTH-domain proteins in plants, to show that most of them can carry out redundant molecular functions and, remarkably, this molecular activity has been present across more than 450 million years of land plant evolution.