In this article we will discuss about the Transcriptional and Post-Transcriptional Regulation of Gene Expression in Eukaryotes.
Transcriptional Regulation of Gene Expression in Eukaryotes:
The variation in the rate of transcription often regulates gene expression. Interactions between RNA polymerase II and basal transcription factors leading to the formation of the transcription initiation complex influence the rate of transcription. Other transcription factors change the rate of transcription initiation by binding to promoter sequences. The rate of transcription is also influenced by enhancers and silencers.
This is a site for regulation of transcription. Every structural gene in eukaryotes has the promoter site which consists of several hundred nucleotide sequences that serve as the recognition point for RNA polymerase binding, located at a fixed distance from the site where transcription is initiated.
Eukaryotic promoters require the binding of a number of protein factors to initiate transcription. Promoter regions are recognized by RNA polymerase II, which transcribes primarily mRNA, consists of short DNA sequences usually located within 100 bp upstream (in the 5′ direction) of the gene.
The promoter regions of most eukaryotic gene contain several specific regions such as:
(a) TATA box
(b) CAAT box
(c) GC box (Fig. 17.9).
Variation in the rate of transcription often regulates gene expression. Interactions between RNA polymerase II and basal transcription factors lead to formation of transcription initiation complex (TIC) at the TATA box.
It is located about 25-30 bases upstream from the initial point of transcription, it consists of an 8 bp consensus sequence composed of A = T base pairs (TATAAA) only, but flanked on either side by G=C rich regions. Mutation in TATA box reduces transcription or may alter the initiation point. TATA box is also known as Hogness box.
Many promoters contain other components and also bear the consensus sequence like GGCCAATC which is situated at the region 70-80 bp from the start site, it can function in both 5-3′ or a 3-5′ orientation. Mutational analysis showed that CAAT box plays the strongest role in determining the efficiency of the promoter.
Another element often seen in some promoter regions, called the GC box, has the consensus sequence GGGCGG and is found at about position -110, often occurs in multiple copies, the GC elements bind transcription factors and function more like enhancer.
Binding of RNA Polymerase II to Promoters:
The binding of RNA polymerase II to its promoter site requires a number of transcriptional factors (TPs).
Promoters have multiple binding sites for transcription factors each of which can influence transcription. TF IID is the first transcriptional factor to bind close to the promoter at an initiator site about -20 to -10 base pairs before the transcriptional start site, i.e., at the TATA boxes, so it is also called TATA box binding protein (TBP).
TF IID may also interact with other transcriptional factors like TF IIA, TF MB and TF ME. A complex consisting of all transcriptional factors determine which RNA polymerase binds and which gene can be transcribed, and the complex is called pre-initiation complex.
The transcription factors have a modular structure containing DNA binding, dimerization and transactivation domains.
DNA binding domains contain three motifs: helix-turn-helix, zinc fingers and basic domains which occur in combination with dimerization domains.
Dimerization domains contain two motifs: leucine zippers and helix-loop-helix.
Dimerization allows the formation of homo- and heterodimers creating transcription factors with diverse functions. Transactivation domains have no motifs but are often enriched with acidic amino acid, glutamines or pro-lines. They interact with a variety of proteins at different stages during transcription. Transcription factors can also repress transcription by direct or indirect mechanisms.
The transcriptional factors are produced constitutively, but except these there are some transcriptional activators (TAs) which bind to the enhancer site situated many hundreds base pairs from the promoter site.
These transcriptional activators are induced proteins, i.e., synthesized only in response to specific signals, which on binding with DNA forms the loop back on itself when they interact with the TFs near the promoter. This interaction between enhancer site and initiation site is usually necessary for transcription above a basal level (Fig. 17.10).
Co-activators are activator proteins that often connect TFs and TAs and may be essential for expression of gene at high level.
There are many ways by which negative control of transcription takes place in eukaryotes.
These can be divided into 3 main categories:
(1) Inhibition of DNA binding;
(ii) Blocking of activation;
(iii) Silencing, i.e., transcriptional activation factor (TAP) cannot bind with transcription initiation complex (TIC) due to presence of silencer factor.
Like an enhancer, a silencer also functions irrespective of its position (many thousands base pairs away) and orientation relative to the gene, whose expression it controls. The silencer factor (a protein) either locks the transcription initiation complex or makes it unavailable for activating factors or it disorganizes the transcription initiation complex (Fig. 17.11).
Among the various models, the Britten and Davidson model for regulation of protein synthesis in eukaryotes is most popular. This model is also called genes controlled by one sensor site is termed as battery.
This model assumes the presence of four classes of sequences (Fig. 17.12a):
(i) Producer genes:
It is comparable with the structural gene of a prokaryotic operon.
(ii) Receptor site:
It is comparable to operator gene in bacterial operon and one such receptor site is always assumed to be present adjacent to each producer gene or a set of producer gene.
(iii) Integrator gene:
It is comparable to regulator gene and is responsible for synthesis of an activator RNA that may or may not give rise to proteins before it activates the receptor site.
(iv) Sensor site:
A sensor site regulates the activity of integrator gene, which can be transcribed only when the sensor site is activated by agents like hormones and proteins, changes the pattern of gene expression. In this model the genes (producer gene and integrator gene) are involved in RNA synthesis whereas receptor and sensor sites are those sequences which help only in recognition without taking part in RNA synthesis.
It is proposed in this model that receptor sites and integrator genes may be repeated a number of times to control the activity of a large number of genes in the same cell. Repetition of receptor ensures that same activator recognises all of them and several enzymes of one pathway are simultaneously synthesized.
When the transcription of same gene is needed at different developmental stages, it can be achieved by multiplicity of receptor sites and integrator genes.
Each producer gene may have several receptor sites, each responding to one activator (Fig. 17.12b) so that a single activator thus can recognize several genes at a time. One sensor site may regulate the activity of several integrators and different activators may activate the same gene at different times. An integrator gene may also fall in cluster with same sensor site (Fig. 17.12c).
Regulation of Gene Expression by Hormones:
Hormones influence target cells by activating gene transcription. Steroid hormones on entering cells, bind steroid hormone receptor protein, releasing it from an inhibitory protein. The receptor dimerizes and is trans-located to the nucleus where it binds to target gene promoters activating transcription.
Polypeptide hormones bind receptor proteins on the surface of target cells. Signal transduction triggers gene activation in which a sequential activation of several proteins by phosphorylation takes place.
Post-Transcriptional Regulation of Gene Expression in Eukaryotes:
Post-transcriptional regulation of gene expression may occur in different ways.
Regulation of Processing:
Post-transcriptional modes of regulation also occur in many organisms where the eukaryotic nuclear RNA transcripts are modified prior to translation, non-coding introns are removed, the remaining exons are precisely spliced together and the mRNA is modified by the addition of cap at the 5′ end and a poly-A tail after end.
The message is then complexed with proteins and exported to the cytoplasm. Each of these processing steps offers several possibilities for regulation, for example, several alternative splicing pathways of a single pre-mRNA transcript to give multiple mRNAs and regulation of the stability of mRNA itself. This leads to the synthesis of different proteins or isoforms in the same time and space.
Regulation of Translation:
Regulation at translational level occurs in different ways:
(i) Activation and repression of translation:
In eukaryotes the activator protein binds to mRNA and leads to the formation of hairpin structure which helps in ribosome binding with mRNA by the exposure of 5′ end. The translational repressor protein (IRE-BP) controls ferritin synthesis by down-regulation and transferring receptor synthesis by up-regulation.
(ii) Regulation by phosphorylation machinery:
Translational repressor protein may regulate the translation in eukaryotic system or regulation of translation is brought about by modification of general components of translational machinery.
Reversible phosphorylation machinery is involved in the regulation of gene expression, as the phosphorylated or dephosphorylated forms of the components of translational machinery should identify a specific mRNA from the bulk mRNA population.