Transcription Factors

Driving Developmental Progression: Gene Regulatory Network

The necessary expression of various genes during embryonic development or cell differentiation is strictly governed by gene regulatory network that is characterized by extensive interaction between DNA and its associated proteins. The network incorporates integrated action of cis-regulatory element (e.g. core promoter promoter-proximal elements) and cis-regulatory modules (e.g. silencers, insulators, etc.) both in temporal and spatial manner. For the most part, enhancers and transcription factors (TF) play an intermediate role in gene regulatory network. Here, we will introduce 1) the TF mechanisms in mediating the gene regulatory network, 2) the dynamic change of these elements during developmental progression, and 3) the regulatory landscape the accurately regulate gene expression.

TFs in gene regulatory network:

Combinatorial binding of TF results in a precise pattern of transcriptional activity.

TF can occupy a variety of enhancer sets depending on the stage of development and situation. It is suggested that the temporal change in its TF occupancy is not influenced by the timing of the expression of TF but rather by the timing of DNA occupancy controlling the transient nature of the gene regulatory network that promotes the developmental progression.

Different binding modes yield different types of transcriptional output.

TFs are known to interact with different types of cofactors altering the transcriptome. While it is commonly known that concentrations of individual TFs, like NF-kB, is proportional to the enhancer activity, cooperative binding can lead to nonlinear relationship with the degree of occupancy, thus resulting in different transcriptional response such as switch-like effect.
Furthermore, same set of TFs can function differently at different concentration range.

TF engage in cooperative activities with binding proteins for indirect regulation.

Cooperative binding to DNA is due to the direct protein-protein interaction between TFs that bind to adjacent sites on the strand. TF, for instance, interacts with coactivator p300-CREB binding protein (CBP) family, with corepressor, Groucho or with other elements including, mediator, SAGA, TAFIID complexes, to bring about a synergistic effect on transcription.

Chromatin Accessibility and TF binding

TF occupancy is also influenced by the position of nucleosomes in enhancers where histones and TF compete for access to DNA. Also, the interaction between TF and the nucleosome is also affected by the posttranslational modification of the histone tail within the nucleosome. Specific histone modifications are found in the body of active or inactive promoters and transcribed genes and have dynamic and cell-specific patterns in cis-regulatory elements.

In addition, the enhancer architecture has shown to play a crucial role for TF complex binding to the correct sequence motif. This mechanism deals with the motif positioning, which is defined by the relative order, orientation and spacing of motifs within an enhancer. For example, if the sites that allow the binding of TF complex is not spaced correctly, this could lead to helical phasing in which the TF complex fails to bind due to the altered orientation of the DNA.

Enhancers and its impact on developmental progression

The composition of TF bound to the enhancer not only reflects the active state of the enhancer at some point. In fact, it is influenced by gradual dynamic change during development. Continuous events from priming to activation have been suggested to play an important functional role in maintaining the precision of the gene expression program even for events that do not have discernible effects on gene expression .

Pioneer factors are responsible for gene accessibility

Enhancers’ function involved in developmental progression may be regulated not only by TF activity but also by chromatin remodeling. In embryonic development and cell differentiation, most TFs aggregate transcription complexes with specific enhancers to induce nucleosomal rearrangement. This is specifically done by increasing accessibility to other factors. In most cases, however, pioneer TF is necessary but not sufficient to induce the progression of development; other cofactors must be recruited at enhancer site to perform further function. Molecules such as MYOD 1, PAX 5, PU. 1, FOXA 1; HNF 3 α, CCAAT / enhancer binding protein – β (C / EBP β) are known as pioneer TF and they act at the top of the gene regulatory network.

In addition to the repositioning of nucleosomes, protection from DNA methylation within enhancers is also involved in TF enhancer occupancy at developmental stage. Many enhancers have been reported to be cell-specific and have low levels of DNA methylation that are affected by TF binding.

Enhancer Priming and Development

Developmental progression and cellular determinations are regulated by GRN. The continuous expression of TF under GRN function (SOX factor in neurogenesis, or MYF factor in myogenesis) binds to similar target sequences forming structures during development. But this does not necessarily mean that the enhancer-bound TF will induce transcriptional activity.

Priming specific enhancers promote the transitional stage by reducing the rate limiting step of chromatin remodeling. Many enhancers within progenitor cells undergo specific histone tail modification (eg H3K4me1 or H3K4me2), even though these elements are not associated with strong regulatory activity at that stage. On the other hand, improper TF may be restrained at the later stage of development by enhancer priming. Therefore, indiscriminate binding of TF in the enhancer made accessible in the early stages of development needs to prevent inappropriate activation of the primed enhancer.

Changes at enhancer during developmental progression

At early stage, considerable number of enhancer elements are covered by high numbers of nucleosomes to prevent TF binding

Upon expression of pioneer factors, these proteins function to alter chromatin remodeling, rearranging nucleosomes to allow enhancer accessibility for further activation.

At later stage, other TFs along with cofactors begin binding to enhancer regions of open chromatin site

Regulatory landscape

Spatiotemporal expression of genes requires specific genomic structures, including hierarchical interactions between regulatory elements, thereby maintaining precise and robust expression.

Integration of enhancers for gene expression

The developmental genes are regulated by multiple enhancer elements, each controlling a particular spatiotemporal aspect of gene expression. Recent studies have suggested that this multiple enhancers also contains a “secondary” enhancer that overlaps or shows the same spatial activity as the primary enhancer, closer to the gene promoter. In other words, “primary” and “secondary” are distinguished by the physical distance to the gene promoter or the order in which the enhancer was found. The activities of the two elements are nearly equivalent and the secondary enhancer is said to be a shadow enhancer that provides the phenotypic robustness essential for the decision process such as embryogenesis.

Cis-regulatory mechanism of genome

In many genes regulated by remote enhancers, it is thought that a large chromatin loop places enhancer and target genes physically close together, resulting in interaction. Many of the enhancers involved in development also act on not only biologically relevant target genes but also irrelevant adjacent genes. Relative placement of regulatory elements is essential for the output of signals when the remote enhancer is involved in the three dimensional organization of the genome. It is highlighted by the various developmental phenotypes associated with structural changes in the genome affecting the genetic locus of development. Enhancer positions on genes and cis-regulatory elements are constrained by the process of evolution. Thus, as with the enhancer’s output, which is influenced by the relative position of the TF binding site, the transcript is formed by the overall system of loci that includes the relative positions of genes, enhancers and interspersed structural elements.

Summary

Protein-protein interaction by combination of multiple TFs characterize the function of a gene regulatory network. ·

Gene regulation during development is influenced not only by the activation by enhancer’s simple on / off switch but also by the gradual change in developmental stage.

Various regulatory systems are in place to maintain the accuracy and robustness of gene expression.

References

Nature Reviews Genetics 13, 613–626 (2012)  doi:10.1038/nrg3207

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