Origin of RNA Polymerase II pause in eumetazoans: Insights from Hydra

Multicellular organisms have evolved sophisticated mechanisms for responding to various developmental,environmental and physical stimuli by regulating transcription. The correlation of distribution of RNAPolymerase II (RNA Pol II) with transcription is well established in higher metazoans, however genome-wideinformation about its distribution in early metazoans, such as Hydra, is virtually absent. To gain insights intoRNA Pol II-mediated transcription and chromatin organization in Hydra, we performed chromatin immunoprecipitation (ChIP)-coupled high-throughput sequencing (ChIP-seq) for RNA Pol II and Histone H3. Strikingly, we found that Hydra RNA Pol II is uniformly distributed across the entire gene body, as opposed to itscounterparts in bilaterians such as human and mouse. Furthermore, correlation with transcriptome datarevealed that the levels of RNA Pol II correlate with the magnitude of gene expression. Strikingly, thecharacteristic peak of RNA Pol II pause typically observed in bilaterians at the transcription start sites (TSSs)was not observed in Hydra. The RNA Pol II traversing ratio in Hydra was found to be intermediate to yeastand bilaterians. The search for factors involved in RNA Pol II pause revealed that RNA Pol II pausingmachinery was most likely acquired first in Cnidaria. However, only a small subset of genes exhibited thepromoter proximal RNP Pol II pause. Interestingly, the nucleosome occupancy is highest over the subset ofpaused genes as compared to total Hydra genes, which is another indication of paused RNA Pol II at thesegenes. Thus, this study provides evidence for the molecular basis of RNA Pol II pause early during theevolution of multicellular organisms.

Comparative sequence analyses of genome and transcriptome reveal novel transcripts and variants in the Asian elephant Elephas maximus.

The Asian elephant Elephas maximus and the African elephant Loxodonta africana that diverged 5–7 million years ago exhibit differences in their physiology, behaviour and morphology. A comparative genomics approach would be useful and necessary for evolutionary and functional genetic studies of elephants. We performed sequencing of E. maximus and map to L. africana at ~15X coverage. Through comparative sequence analyses, we have identified Asian elephant specific homozygous, non-synonymous single nucleotide variants (SNVs) that map to 1514 protein coding genes, many of which are involved in olfaction. We also present the first report of a high-coverage transcriptome sequence in E. maximus from peripheral blood lymphocytes. We have identified 103 novel protein coding transcripts and 66-long non-coding (lnc)RNAs. We also report the presence of 181 protein domains unique to elephants when compared to other Afrotheria species. Each of these findings can be further investigated to gain a better understanding of functional differences unique to elephant species, as well as those unique to elephantids in comparison with other mammals. This work therefore provides a valuable resource to explore the immense research potential of comparative analyses of transcriptome and genome sequences in the Asian elephant.

N-terminal PDZ-like domain of chromatin organizer SATB1 contributes towards its function as transcription regulator.

The special AT-rich DNA-binding protein 1 (SATB1) is a matrix attachment region (MAR)-binding protein that acts as a global repressor via recruitment of CtBP1:HDAC1-containing co-repressors to its binding targets. The N-terminal PSD95/Dlg-A/ZO-1 (PDZ)-like domain of SATB1 mediates interactions with several chromatin proteins. In the present study, we set out to address whether the PDZ-domain-mediated interactions of SATB1 are critical for its in vivo function as a global repressor. We reasoned that since the N-terminal PDZ-like domain (amino acid residues 1–204) lacks DNA binding activity, it would fail to recruit the interacting partners of SATB1 to its genomic binding sites and hence would not repress the SATB1-regulated genes. Indeed, in vivo MAR-linked luciferase reporter assay revealed that overexpression of the PDZ-like domain resulted in de-repression, indicating that the PDZ-like domain exerts a dominant negative effect on genes regulated by SATB1. Next, we developed a stable dominant negative model in human embryonic kidney (HEK) 293T cells that conditionally expressed the N-terminal 1–204 region harbouring the PDZ-like domain of SATB1. To monitor the effect of sequestration of the interaction partners on the global gene regulation by SATB1, transcripts from the induced and uninduced clones were subjected to gene expression profiling. Clustering of expression data revealed that 600 out of 19000 genes analysed were significantly upregulated upon overexpression of the PDZ-like domain. Induced genes were found to be involved in important signalling cascades and cellular functions. These studies clearly demonstrated the role of PDZ domain of SATB1 in global gene regulation presumably through its interaction with other cellular proteins.