We investigated the possibility that H3 provides a platform to integrate information relevant to whether the associated DNA should be methylated. P16 caused partial or total loss of DNA methylationin vivo. Only residues R8-G12 were required for DIM-5 activityin vitro. DIM-5 activity was inhibited by dimethylation of H3K4 and by phosphorylation of H3S10, but not by acetylation of H3K14. We conclude that this H3 tail acts as an integrating platform for signals that influence DNA methylation, in part through methylation of H3K9. == Author Summary == DNA methylation is usually a common feature of eukaryotic genomes. Methylation is typically associated with silenced chromosomal domains and is essential for development of plants and animals. Even though control of DNA methylation is not well understood, recent findings with model organisms, including the fungusNeurospora crassa, revealed FANCD connections between modifications of histones and DNA. DNA methylation is usually dispensable in Neurospora, facilitating genetic studies. Isolation of mutants defective in DNA methylation revealed that a histone H3 methyltransferase, DIM-5, is required for DNA methylation. DIM-5 trimethylates H3K9, which is then recognized by Heterochromatin Protein 1 (HP1), which recruits the DNA methyltransferase DIM-2. We investigated the possibility that H3 provides a platform to integrate information relevant to whether the associated DNA should be methylated. Indeed, we found that DIM-5 is usually sensitive to methylation of H3K4 and phosphorylation of H3S10. Our analyses further revealed that H3K14 is usually criticalin vivo, but not because acetyl-K14 inhibits DIM-5. We also found that H3R2 is essential for DNA methylationin vivobut not important for DIM-5 activity. Interestingly, we found H3 mutants that show recessive defects in DNA methylation as well as others with dominant effects. We also defined a set of H3 mutations that are lethal. == Introduction == The primary structures of histones, the small basic proteins that are complexed with DNA to form chromatin in eukaryotes, are highly conserved but not invariant[1],[2]. For example, comparisons between a sea urchin and the filamentous fungusNeurospora crassareveal that the two most highly conserved histones, H3 and H4, have 16/135 and 9/102 amino acid differences, respectively[3],[4]. Pioneering studies with the yeastsSaccharomyces cerevisiaeandSchizosaccharomyces pomberevealed Tenofovir alafenamide fumarate that mutation, or even deletion, of some histone residues is not lethal, allowing for genetic studies of structure/function relationships of these prominent proteins[5][9]. Histones are subject to a variety of posttranslational Tenofovir alafenamide fumarate modifications including methylation, acetylation, phosphorylation, ubiquitylation, sumoylation, as well as ADP-ribosylation, deimination and proline isomerization[1],[10]. An explosion of findings has implicated these modifications in various fundamental cellular processes, including transcription, option splicing, replication, chromosome condensation, recombination, DNA repair and DNA methylation[1],[10],[11]. One approach to investigate potential involvement of modifications of a particular histone residue is usually to test for effects of substitutions that prevent the modification or mimic the modified state. For example, because acetylation of lysines reduces the positive charge of this residue, it can be informative to test the effect of substituting lysines with non-modifyable residues that are either neutral (e.g. glutamine) or positively charged (e.g. Tenofovir alafenamide fumarate arginine)[12]. To gain insight into residues involved Tenofovir alafenamide fumarate in DNA methylation and other cellular processes, we carried out a systematic analysis of the heavily altered N-terminus ofN. crassahistone H3[13]. Neurospora is an excellent organism to study effects of histone H3 mutations in part because the wild-type genome contains a single H3 gene,hH3[3],[4]. A direct link between H3 modifications and DNA methylation was originally exhibited in.