Patterns of chromatin accessibility along the anterior-posterior axis in the early Drosophila embryo.

As the Drosophila embryo transitions from the use of maternal RNAs to zygotic transcription, domains of open chromatin, with relatively low nucleosome density and specific histone marks, are established at promoters and enhancers involved in patterned embryonic transcription. However it remains unclear how regions of activity are established during early embryogenesis, and if they are the product of spatially restricted or ubiquitous processes. To shed light on this question, we probed chromatin accessibility across the anterior-posterior axis (A-P) of early Drosophila melanogaster embryos by applying a transposon based assay for chromatin accessibility (ATAC-seq) to anterior and posterior halves of hand-dissected, cellular blastoderm embryos. We find that genome-wide chromatin accessibility is highly similar between the two halves, with regions that manifest significant accessibility in one half of the embryo almost always accessible in the other half, even for promoters that are active in exclusively one half of the embryo. These data support previous studies that show that chromatin accessibility is not a direct result of activity, and point to a role for ubiquitous factors or processes in establishing chromatin accessibility at promoters in the early embryo. However, in concordance with similar works, we find that at enhancers active exclusively in one half of the embryo, we observe a significant skew towards greater accessibility in the region of their activity, highlighting the role of patterning factors such as Bicoid in this process.

Convergence of topological domain boundaries, insulators, and polytene interbands revealed by high-resolution mapping of chromatin contacts in the early Drosophila melanogaster embryo.

High-throughput assays of three-dimensional interactions of chromosomes have shed considerable light on the structure of animal chromatin. Despite this progress, the precise physical nature of observed structures and the forces that govern their establishment remain poorly understood. Here we present high resolution Hi-C data from early Drosophila embryos. We demonstrate that boundaries between topological domains of various sizes map to DNA elements that resemble classical insulator elements: short genomic regions sensitive to DNase digestion that are strongly bound by known insulator proteins and are frequently located between divergent promoters. Further, we show a striking correspondence between these elements and the locations of mapped polytene interband regions. We believe it is likely this relationship between insulators, topological boundaries, and polytene interbands extends across the genome, and we therefore propose a model in which decompaction of boundary-insulator-interband regions drives the organization of interphase chromosomes by creating stable physical separation between adjacent domains.

Dense Bicoid hubs accentuate binding along the morphogen gradient.

Morphogen gradients direct the spatial patterning of developing embryos; however, the mechanisms by which these gradients are interpreted remain elusive. Here we used lattice light-sheet microscopy to perform in vivo single-molecule imaging in early Drosophila melanogaster embryos of the transcription factor Bicoid that forms a gradient and initiates patterning along the anteroposterior axis. In contrast to canonical models, we observed that Bicoid binds to DNA with a rapid off rate throughout the embryo such that its average occupancy at target loci is on-rate-dependent. We further observed Bicoid forming transient "hubs" of locally high density that facilitate binding as factor levels drop, including in the posterior, where we observed Bicoid binding despite vanishingly low protein levels. We propose that localized modulation of transcription factor on rates via clustering provides a general mechanism to facilitate binding to low-affinity targets and that this may be a prevalent feature of other developmental transcription factors.

Local regulation of gene expression by lncRNA promoters, transcription and splicing.

Mammalian genomes are pervasively transcribed to produce thousands of long non-coding RNAs (lncRNAs). A few of these lncRNAs have been shown to recruit regulatory complexes through RNA-protein interactions to influence the expression of nearby genes, and it has been suggested that many other lncRNAs can also act as local regulators. Such local functions could explain the observation that lncRNA expression is often correlated with the expression of nearby genes. However, these correlations have been challenging to dissect and could alternatively result from processes that are not mediated by the lncRNA transcripts themselves. For example, some gene promoters have been proposed to have dual functions as enhancers, and the process of transcription itself may contribute to gene regulation by recruiting activating factors or remodelling nucleosomes. Here we use genetic manipulation in mouse cell lines to dissect 12 genomic loci that produce lncRNAs and find that 5 of these loci influence the expression of a neighbouring gene in cis. Notably, none of these effects requires the specific lncRNA transcripts themselves and instead involves general processes associated with their production, including enhancer-like activity of gene promoters, the process of transcription, and the splicing of the transcript. Furthermore, such effects are not limited to lncRNA loci: we find that four out of six protein-coding loci also influence the expression of a neighbour. These results demonstrate that cross-talk among neighbouring genes is a prevalent phenomenon that can involve multiple mechanisms and cis-regulatory signals, including a role for RNA splice sites. These mechanisms may explain the function and evolution of some genomic loci that produce lncRNAs and broadly contribute to the regulation of both coding and non-coding genes.

ZBTB7A Suppresses Melanoma Metastasis by Transcriptionally Repressing MCAM.

UNLABELLED: The excessive metastatic propensity of melanoma makes it the most deadly form of skin cancer, yet the underlying mechanism of metastasis remains elusive. Here, mining of cancer genome datasets discovered a frequent loss of chromosome 19p13.3 and associated downregulation of the zinc finger transcription factor ZBTB7A in metastatic melanoma. Functional assessment of ZBTB7A-regulated genes identified MCAM, which encodes an adhesion protein key to melanoma metastasis. Using an integrated approach, it is demonstrated that ZBTB7A directly binds to the promoter and transcriptionally represses the expression of MCAM, establishing ZBTB7A as a bona fide transcriptional repressor of MCAM. Consistently, downregulation of ZBTB7A results in marked upregulation of MCAM and enhanced melanoma cell invasion and metastasis. An inverse correlation of ZBTB7A and MCAM expression in association with melanoma metastasis is further validated with data from analysis of human melanoma specimens. IMPLICATIONS: Together, these results uncover a previously unrecognized role of ZBTB7A in negative regulation of melanoma metastasis and have important clinical implications.

ZBTB7A acts as a tumor suppressor through the transcriptional repression of glycolysis.

Elevated glycolysis is a common metabolic trait of cancer, but what drives such metabolic reprogramming remains incompletely clear. We report here a novel transcriptional repressor-mediated negative regulation of glycolysis. ZBTB7A, a member of the POK (POZ/BTB and Krüppel) transcription repressor family, directly binds to the promoter and represses the transcription of critical glycolytic genes, including GLUT3, PFKP, and PKM. Analysis of The Cancer Genome Atlas (TCGA) data sets reveals that the ZBTB7A locus is frequently deleted in many human tumors. Significantly, reduced ZBTB7A expression correlates with up-regulation of the glycolytic genes and poor survival in colon cancer patients. Remarkably, while ZBTB7A-deficient tumors progress exceedingly fast, they exhibit an unusually heightened sensitivity to glycolysis inhibition. Our study uncovers a novel tumor suppressor role of ZBTB7A in directly suppressing glycolysis.