Antisense oligonucleotides (ASOs) modified with phosphorothioate (PS) linkages and different 2' modifications can be used either as drugs (e.g., to treat homozygous familial hypercholesterolemia and spinal muscular atrophy) or as research tools to alter gene expression. PS-ASOs can enter cells without additional modification or formulation and can be designed to mediate sequence-specific cleavage of different types of RNA (including mRNA and non-coding RNA) targeted by endogenous RNase H1. Although PS-ASOs function in both the cytoplasm and nucleus, localization to different subcellular regions can affect their therapeutic potency. Cellular uptake and intracellular distribution of PS ASOs are mediated by protein interactions. The main proteins involved in these processes have been identified, and intracellular sites in which PS ASOs are active, or inactive, cataloged.

Histone lysines can be mono-, di-, or trimethylated, providing an ample magnitude of epigenetic information for transcription regulation. In fungi, SET2 is the sole methyltransferase responsible for mono-, di-, and trimethylation of H3K36. Here we show that in Arabidopsis thaliana, the degree of H3K36 methylation is regulated by distinct methyltransferases. The SET2 homologs SDG8 and SDG26 each can methylate oligonucleosomes in vitro, and both proteins are localized in the nucleus. While the previously reported loss-offunction sdg8 mutants have an early-flowering phenotype, the loss-of-function sdg26 mutants show a lateflowering phenotype. Consistently, several MADS-box flowering repressors are down-regulated by sdg8 but up-regulated by sdg26. The sdg8 but not the sdg26 mutant plants show a dramatically reduced level of both diand trimethyl-H3K36 and an increased level of monomethyl-H3K36. SDG8 is thus specifically required for diand trimethylation of H3K36. Our results further establish that H3K36 di-and tri-but not monomethylation correlates with transcription activation. Finally, we show that SDG8 and VIP4, which encodes a component of the PAF1 complex, act independently and synergistically in transcription regulation. Together our results reveal that the deposition of H3K36 methylation is finely regulated, possibly to cope with the complex regulation of growth and development in higher eukaryotes.During the past few years, histone lysine (K) methylation has been viewed to play widespread roles in transcriptional regulation, DNA repair, and epigenetic inheritance (15, 32). It occurs on histone H3K4, H3K9, H3K27, H3K36, and H4K20 in several studied eukaryotes. In general, H3K4 and H3K36 methylation is associated with actively transcribed genes, whereas H3K9, H3K27, and H4K20 methylation is associated with transcriptional repression and silenced chromatin regions. Furthermore, K residues can be mono-, di-or trimethylated, and the degree of methylation on H3K4, H3K9, H3K27, and H4K20 has considerable influence on transcriptional activation or repression (15,43,55,63). In comparison, methylation on H3K36 is less extensively characterized. In fungi, such as Saccharomyces cerevisiae, Schizosaccharomyces pombe, and Neurospora crassa, a sole histone-lysine-methyltransferase (HKMT), SET2, is responsible for mono-, di-, and trimethylation of H3K36 (1, 35, 49). In mammals, both the Sotos syndrome and leukemia-associated protein NSD1 and the Huntington disease protein HYPB can methylate H3K36 in vitro (42,50). In Arabidopsis thaliana, the loss-of-function sdg8 (also named efs) mutants show a dramatically reduced level of H3K36 dimethylation and an early-flowering phenotype (25,65). Because of these phenotype-associated crucial functions, unraveling the mechanism of deposition of H3K36 methylation in mammals and in plants is of particular importance.Proper timing of flowering is pivotal for the reproductive success of plants and thus is controlled by complex genetic networks, which involve histone modifications and chroma...

Flowering represents a crucial transition from a vegetative to a reproductive phase of the plant life cycle. Despite extensive studies, the molecular mechanisms controlling flowering remain elusive. Although the enzymes involved are unknown, methylation of histone H3 K9 and K27 correlates with repression of FLOWERING LOCUS C (FLC), an essential transcriptional repressor involved in flowering time control in Arabidopsis thaliana; in contrast, methylation of H3K4 correlates with FLC activation. Here we show that loss-of-function of SET DOMAIN GROUP 8 (SDG 8), which encodes a homologue of the yeast SET2 histone methyltransferase, results in reduced dimethylation of histone H3K36, particularly in chromatin associated with the FLC promoter and the first intron, regions that contain essential cis-elements for transcription. sdg8 mutants display reduced FLC expression and flower early, establishing SDG8-mediated H3K36 methylation as a novel epigenetic memory code required for FLC expression in preventing early flowering. This is the first demonstrated role of H3K36 methylation in eukaryote development.