Willett, C. G. | bioRxiv Science

bioRxiv2023

Differences in transcription initiation directionality underlie distinctions between plants and animalsin chromatin modification patterns at genes and cis-regulatory elements

Transcriptional initiation is among the first regulated steps controlling eukaryotic gene expression. High-throughput profiling of fungal and animal genomes has revealed that RNA Polymerase II (Pol II) often initiates transcription in both directions at the promoter transcription start site (TSS), but generally only elongates productively into the gene body. Additionally, Pol II can initiate transcription in both directions at cis-regulatory elements (CREs) such as enhancers. These bidirectional Pol II initiation events can be observed directly with methods that capture nascent transcripts, and they are also revealed indirectly by the presence of transcription-associated histone modifications on both sides of the TSS or CRE. Previous studies have shown that nascent RNAs and transcription-associated histone modifications in the model plant Arabidopsis thaliana accumulate mainly in the gene body, suggesting that transcription does not initiate widely in the upstream direction from genes in this plant. We compared transcription-associated histone modifications and nascent transcripts at both TSSs and CREs in Arabidopsis thaliana, Drosophila melanogaster, and Homo sapiens. Our results provide evidence for mostly unidirectional Pol II initiation at both promoters and gene-proximal CREs of Arabidopsis thaliana, whereas bidirectional transcription initiation is observed widely at promoters in both Drosophila melanogaster and Homo sapiens, as well as CREs in Drosophila. Furthermore, the distribution of transcription-associated histone modifications around TSSs in the Oryza sativa (rice) and Glycine max (soybean) genomes suggests that unidirectional transcription initiation is the norm in these genomes as well. These results suggest that there are fundamental differences in transcriptional initiation directionality between flowering plant and metazoan genomes, which are manifested as distinct patterns of chromatin modifications around RNA polymerase initiation sites.

genomics↗

bioRxiv2023

Replacement of Arabidopsis H2A.Z with human H2A.Z orthologs reveals extensive functional conservation and limited importance of the N-terminal tail sequence for Arabidopsis development

The incorporation of histone variants, distinct paralogs of core histones, into chromatin affects all DNA-templated processes in the cell, including the regulation of transcription. In recent years, much research has been focused on H2A.Z, an evolutionarily conserved H2A variant found in all eukaryotes. In order to investigate the functional conservation of H2A.Z histones during eukaryotic evolution we transformed h2a.z deficient plants with each of the three human H2A.Z variants to assess their ability to rescue the mutant defects. We discovered that human H2A.Z.1 and H2A.Z.2.1 fully complement the phenotypic abnormalities of h2a.z plants despite significant divergence in the N-terminal tail sequences of Arabidopsis and human H2A.Zs. In contrast, the brain-specific splice variant H2A.Z.2.2 has a dominant-negative effect in wild-type plants, mimicking an H2A.Z deficiency phenotype. Furthermore, H2A.Z.1 almost completely re-establishes normal H2A.Z chromatin occupancy in h2a.z plants and restores the expression of more than 84% of misexpressed genes. Finally, we used a series of N-terminal tail truncations of Arabidopsis HTA11 to reveal that the N-terminal tail of Arabidopsis H2A.Z is not necessary for normal plant development but does play an important role in mounting proper environmental stress responses. Article SummaryH2A.Z is a histone variant conserved across the eukaryotes that plays important roles in transcription. Despite high overall conservation of H2A.Z protein sequence between organisms, sequence variation is seen in the N-terminal region, a site for many posttranslational modifications implicated in H2A.Z function. We replaced Arabidopsis thaliana H2A.Z with each of the human H2A.Z orthologs and found that both human H2A.Z isoforms rescue the severe phenotypes of plant H2A.Z null mutants, despite divergent N-terminal sequences. We further analyzed N-terminal sequence requirements by progressively deleting the endogenous plant H2A.Z N-terminus. Plants with a tailless version of H2A.Z developed normally but were impaired in stress responsiveness. Our results indicate that human H2A.Zs are broadly functional in plants and that the N-terminus of H2A.Z is not essential for normal development but is implicated in stress responsiveness.

genetics↗