In higher eukaryotes, heterochromatin is mainly composed of transposable elements (TEs) silenced by epigenetic mechanisms. But, the silencing of certain heterochromatin-associated TEs is disrupted by heat stress. By comparing genome-wide high-resolution chromatin packing patterns under normal or heat conditions obtained through Hi-C analysis, we show here that heat stress causes global rearrangement of the 3D genome in Arabidopsis thaliana. Contacts between pericentromeric regions and distal chromosome arms, as well as proximal intra-chromosomal interactions along the chromosomes, are enhanced. However, interactions within pericentromeres and those between distal intra-chromosomal regions are decreased. Many inter-chromosomal interactions, including those within the KNOT, are also reduced. Furthermore, heat activation of TEs exhibits a high correlation with the reduction of chromosomal interactions involving pericentromeres, the KNOT, the knob, and the upstream and downstream flanking regions of the activated TEs. Together, our results provide insights into the relationship between TE activation and 3D genome reorganization.

In Arabidopsis, an RNA-directed DNA methylation pathway (RdDM) is responsible for de novo establishment of DNA methylation and contributes to transcriptional gene silencing. Recently, the microrchidia (MORC)-type ATPases were shown to play essential roles in enforcing transcriptional gene silencing of a subset of genes and transposons by regulating the formation of higher-order chromatin architecture. However, how MORC proteins cooperate with the RdDM pathway components to regulate gene expression remains largely unclear. In this study, SUVH9 and MORC6 were identified from a screening of suppressors of idm1, which is a mutant defective in active DNA demethylation. SUVH9 and MORC6 are required for silencing of two reporter genes and some endogenous genes without enhancing DNA methylation levels. SUVH9, one of SU(VAR)3-9 homologs involved in RdDM, directly interacts with MORC6 and its two close homologs, MORC1 and MORC2. Similar to MORC6, SUVH9 and its homolog SUVH2 are required for heterochromatin condensation and formation of 3D chromatin architecture at SDC and Solo-LTR loci. We propose that SUVH2 and SUVH9 bind to the methylated DNA and facilitate the recruitment of a chromatin-remodeling complex to the target loci in association with MORC proteins.

Emerging evidence indicates that tRNA-derived small RNAs (tsRNAs) are involved in fine-tuning gene expression and become dysregulated in various cancers. We recently showed that the 22nt LeuCAG tsRNA from the 3´ end of tRNALeu is required for efficient translation of a ribosomal protein mRNA and ribosome biogenesis. Inactivation of this 3´tsRNA induced apoptosis in rapidly dividing cells and suppressed the growth of a patient-derived orthotopic hepatocellular carcinoma in mice. The mechanism involved in the generation of the 3´tsRNAs remains elusive and it is unclear if the 3´-ends of 3´tsRNAs are aminoacylated. Here we report an enzymatic method utilizing exonuclease T to determine the 3´charging status of tRNAs and tsRNAs. Our results showed that the LeuCAG 3´tsRNA, and two other 3´tsRNAs are fully aminoacylated. When the leucyl-tRNA synthetase (LARS1) was inhibited, there was no change in the total tRNALeu concentration but a reduction in both the charged tRNALeu and LeuCAG 3´tsRNA, suggesting the 3´tsRNAs are fully charged and originated solely from the charged mature tRNA. Altering LARS1 expression or the expression of various tRNALeu mutants were also shown to affect the generation of the LeuCAG 3´tsRNA further suggesting they are created in a highly regulated process. The fact that the 3´tsRNAs are aminoacylated and their production is regulated provides additional insights into their importance in post-transcriptional gene regulation that includes coordinating the production of the protein synthetic machinery.