Thyroid hormone (T3) is important for proper development and normal physiology of many adult organs/tissues in vertebrates [1, 2]. Sever T3 deficiency during human development leads to the formation of human cretins, who are short in stature and severely mentally retarded . The most critical period of T3 action appears to be the several months around birth, the so-called postembryonic period, when T3 levels are high . It has been difficult to study how T3 affects mammalian postembryonic development due to the lack of easily manipulatable models.
Metamorphosis in anurans such as Xenopus laevis or tropicalis mimics mammalian postembryonic development [1, 4]. This process involves distinct changes in different organs and tissues [4, 5]. The larval specific organs, such as the tail and gills, are totally resorbed during metamorphosis, while the adult specific ones, such as the limbs, develop de novo. Most of the organs/tissues are present in both tadpoles and frogs but are drastically remodeled during metamorphosis. For example, the animal intestine involves apoptotic degeneration of larval epithelial cells and concurrent de novo formation of adult stem cells to eventually develop the adult epithelium resembling that in mammals [6, 7]. Interestingly, all such diverse changes during amphibian metamorphosis are totally dependent on T3 . T3 can regulate transcription through T3 receptor (TR). TRs can form heterodimers with 9-cis retinoic acid receptors (RXRs) and repress or activate T3-inducible genes in the absence or presence of T3, respectively [2, 8–10]. Recent studies have shown that TR appears to be both necessary and sufficient to mediate the metamorphic effects of T3 by regulating the transcription of T3 target gene [11–23]. In premetamorphic tadpoles, T3 levels are low and unliganded TR represses T3-inducible genes by recruiting histone deacetylase-containing corepressor complexes [16, 24–26]. This helps to ensure proper premetamorphic growth and prevent premature metamorphosis [16, 26]. When T3 becomes available, liganded TR recruits histone modifying coactivator complexes to these target genes, leading to histone modifications, chromatin remodeling and gene activation [18, 19, 22, 27–31]. This results in metamorphic transformations of different organs.
TR is believed to regulate overlapping but distinct target genes in different organs to effect organ-specific metamorphosis. Toward identifying such target genes, we carried out a ChIP (chromatin immunoprecipitation)-on-chip analysis of the intestine by using a set of microarray chips covering a 8 kb region flanking each putative promoter of 17000 Xenopus tropicalis genes to look for genes bound by TR (unpublished observation). While the ChIP-on-chip data was very preliminary, it was of interest that one of the putative target genes thus identified corresponded to the Xenopus tropicalis homolog of the mammalian Dot1L gene.
Dot1L (Dot1-Like) is the homolog of yeast Dot1 gene, originally identified as a disruptor of telomeric silencing in Saccharomyces cerevisiae. It belongs to the family of lysine methyltransferases (KMTs) [33–36]. Essentially all KMTs contain a SET (Su(var)3-9, Enhancer of Zeste (E(Z)), and Trithorax (trx)) domain. To date, Dot1L is the only known non-SET domain-containing KMT and is the only known KMT that possesses histone methyltransferase activity toward histone H3 lysine (K) 79 [33, 34, 37] Consistently, knockout of Dot1L in mice leads to a complete loss of H3K79 methylation .
H3K79 methylation is one of a large number of posttranslational modifications that occurs at the histone tails in eukaryotic cells . In addition to K79, a number of other K and arginine (R) residues in both histone H3 and H4 can be methylated. These and other histone modifications are distributed in distinct patterns in the genome and variably associated with gene expression levels [22, 40–49]. Among the well-characterized lysine methylation residues include K4, K9, K27, and K79 of histone H3. In general, H3K4 and H3K79 methylations correlate with high levels of transcription, while H3K9 and H3K27 methylations correlate with transcriptional repression. Of particular interest is the recent observation that H3K79 methylation levels are increased at TR target genes upon activation in Xenopus tropicalis intestine during natural and T3-induced metamorphosis . This raises an interesting possibility that Dot1L may be transcriptionally activated by liganded TR and it in turn enhances chromatin remodeling and gene activation by liganded TR during development. Here we provide in vitro and in vivo evidence to show direct transcriptional activation of the Dot1L gene via the binding of TR to a T3 response element (TRE) in the Dot1L promoter in both the intestine and tail. We further show that Dot1L expression correlates with metamorphic changes in both organs, supporting a positive feedback role of Dot1L during metamorphosis.