Paper: Specific but interdependent functions for Arabidopsis AGO4 and AGO6 in RNA-directed DNA methylation by Duan et al.
The function of AGO6 has been considered redundant with AGO4 previously. This paper, however, shows that the redundancy of AGO4 and AGO6 in mediating RdDM is much smaller than we would expect. AGO4 and AGO6 dependent methylation is profiled by genome-wide bisulfite sequencing. Interestingly, DNA methylation in only a small subset of loci is redundantly regulated by AGO4 and AGO6. In more than half of the hypomethylation loci, DNA methylation is similarly reduced in either ago4-6 or ago6-2, and no significant reduction is observed in double mutant. This result indicates that AGO4 and AGO6 have related yet specific function in RdDM.
The authors also want to show the distinct function of AGO4 and AGO6 by studying their subcellular localization. The conclusion of the paper is that AGO4 and AGO6 show different co-localization patterns with DNA dependent RNA polymerases. However, I am not quite convinced by these immuno-staining figures. The localization of AGO4 and AGO6 are scattered in the nucleus and the co-localization signal with Pol IV or Pol V is not obvious. Even though the co-localization data is not convincing to me, I do agree with the authors that studying the localization of AGO4 and AGO6, especially the co-localization pattern with Pol IV or Pol V, is very important.
The other thing I am interested in is that the authors studied the accumulation of Pol V transcripts as well as Pol V occupancy in ago4 and ago6 mutant. A very interesting result is that Pol V occupancy at most tested IGN loci obviously decreases in ago6 mutant. The accumulation of most tested Pol V transcripts decreases in ago6 mutant but increases in ago4 mutant. These results indicate that AGO6 is required for Pol V recruitment. It is very intriguing that AGO4 and AGO6 show such distinct effect on Pol V occupancy. In my own study, I am trying to pull down AGO4 associated Pol V transcripts. It might be interesting to see if Pol V transcripts could also be pull down by AGO6. We have to notice that only a small number of Pol V transcripts are studied here. It remains unclear that whether this small subset can represent the real pattern.
Last thing to mention, another paper from the Slotkin lab (McCue et al. 2015) shows that AGO6 can load 21-22nt siRNAs and establish RdDM, which is also distinct from AGO4. In conclusion, these two proteins may have more specific functions than we expect.