Most but not all arabidopsis YTHDF proteins can replace ECT2/3/4-function.

(A) Diagram showing the relative length of the N-terminal IDRs of Arabidopsis thaliana (Ath) YTHDF proteins (ECT1-ECT11) together with Marchantia polymorpha (Mpo) DFE, Homo sapiens (Hs) YTHDF2, and Drosophila melanogaster (Dm) YTHDF. Numbers indicate the length of the proteins in amino acids (aa). (B) Strategy followed for the functional assay. US7Yp:cECT(X)-mCherry constructs are introduced in ect2-1/ect3-1/ect4-2 (te234) plants, and complementation rates are estimated by the percentage of primary transformants (T1) whose first true leaves have a size (s) exceeding 0.5 mm after 10 days of growth. The construct US7Yp:mCherry is used as negative control. Examples of T1 seedlings expressing control and ECT2 constructs are shown. Dashed outlines are magnified below to show mCherry fluorescence in emerging leaves. (C) Weighed averages of the complementation percentages observed for each US7Yp:cECT(X)-mCherry construct in 2–5 independent transformations (I.T.). Letters between parenthesis indicate the DF- clade assigned to each protein. Colouring highlights ECT2 (reference) and proteins with low or negligible complementation capacity (ECT1/9/11) for fast identification along the manuscript. Low transformation efficiency (T.E.) for ECT9 can be inferred by the low number of transformants (n) highlighted in red, but detailed T.E. and raw complementation rates in each transformation can be found in S9 and S7A Figs respectively. Additional data regarding this assay is shown in S8 Fig. (D) Same as C on a single transformation using the rdr6-12/te234 background. (E-H) Rosette phenotype of 32-day-old T1s bearing US7Yp:cECT(X)-mCherry constructs in the rdr6-12/te234 background, sorted according to their degree of complementation in three panels: F (high), G (residual) and H (non-complementation and enhancement). Different genetic backgrounds and control transformants (mCherry) are shown in E as a reference. The scalebar (5 cm) applies to all plants in the four panels.