Fig. 6

MAS mediates the recruitment of WDR5a to MAF4. a Upper panel, schematic representation of MAF4 locus, the positions of primers (R1 to R5) used for ChIP- and ChIRP-qPCR are indicated. Lower panel, detection of H3K4me3 levels in Col-0 and amiR-MAS-1/2 lines after 20 d of cold exposure by ChIP-qPCR. b Detection of MAS and GAPDH in the whole cell, cytoplasmic and nuclear fractions by RT-PCR. c Detection of MAS and GAPDH in histone 3 (H3) immunoprecipitates by RT-PCR. d ChIRP-qPCR analyses of MAS association with MAF4 locus after 20 d of cold exposure. Left panel, ChIRP enrichment of MAS transcript, but not Actin transcript in both odd and even probe pools. Right panel, qPCR detection of different regions of MAF4 locus in immunoprecipitated DNA. Probes targeting LacZ mRNA were used as negative controls. e Association between MAS and WDR5a detected by RIP with anti-FLAG Magnetic Beads in control and transgenic (FLAG-WDR5a and FLAG-WDR5a^F250A) plants after 20 d of cold exposure. Purification of WDR5a and WDR5a^F250A was validated by western blot (upper panel). The levels of MAS and MAF4 in the immunoprecipitates were determined by RT-qPCR (lower panel). f Detection of WDR5a at MAF4 locus by ChIP-qPCR with an antibody against WDR5 in Col-0 and amiR-MAS-1/2 lines after 20 d of cold exposure. g Detection of WDR5a and WDR5a^F250A at MAF4 locus by ChIP-qPCR in Col-0 and the transgenic plants after 20 d of cold exposure. In (a), d–g error bars represent s.e.m (n = 3–4), asterisks indicate a significant difference between the indicated groups (t-test, P-value < 0.05). h A model for MAS-mediated activation of MAF4 gene expression during vernalization. Source data are provided as a Source Data file