Introduction to Molecular Biology: From Cells to Atoms - Anthony Rees, Michael Sternberg 2002

Nucleic Acids and Genes
Regulation of Gene Expression

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Fig. 28.1.

Gene Expression IN both PROKARYOTES AND EUKARYOTES is regulated by a wide range of mechanisms. Some of these mechanisms operating in bacterial systems are quite well understood, and two of them will be discussed below, whereas much less is known about how regulatory mechanisms function in Eukaryotic Cells.

Prokaryotes are the simplest unicellular organisms whose survival requires only a favorable chemical environment. If a Cell needs a specific metabolite to sustain its metabolic processes, it must be capable of synthesizing the Enzymes required to incorporate it where needed. However, synthesizing such enzymes in the absence of the corresponding metabolite would be wasteful for The Cell.

The hypothesis of Enzyme synthesis induction was proposed by Jacob and Monod in 1961 to explain how bacterial cells respond to environmental changes. Upon introducing lactose (milk sugar) into the medium as a substrate, the level of (ß-galactosidase, an enzyme involved in lactose breakdown, increases 100-fold within the bacterial cell. This type of transcriptional activation is known as induction. Concurrently with lactase, two other Proteins are induced: galactoside permease (a protein that transports lactose across the membrane) and thiogalactoside transacetylase. The three structural genes encoding these proteins are designated as Z, Y, and A, respectively, and together with the operator region, they form the so-called lac Operon. It has been demonstrated that the transcriptional activity of the genes within the operon is regulated by a fourth, regulatory gene. The regulator gene (gene I) in the lac system is located adjacent to the structural genes Z, Y, and A. The existence of gene I was proven by Genetic Methods. When mutant Bacteria lacking gene I (I^- bacteria) were isolated, it was found that in such bacteria the expression of genes Z, Y, and A remains at a high level even in the absence of lactose, i.e., it occurs constitutively. After introducing a DNA fragment containing gene I into I^- bacteria, the expression of genes Z, Y, and A once again became sensitive to the presence of lactose. From this, it was concluded that gene I encodes a diffusible regulatory substance termed a repressor.

A repressor is a protein that blocks gene METABOLISM/31.html">Transcription. In the lac system, the repressor is a tetrameric protein known as the lac repressor. It binds to a specific DNA sequence called the operator.

The operator (O) is a small segment of DNA bordering the first structural gene. The repressor protein can bind to this region, thereby blocking the initiation of transcription. The operator sequence to which the repressor binds contains a palindromic DNA segment. The 2nd-order axis of Symmetry sequence shown in Fig. 28.2 is part of the repressor-binding site in the lac operon.

The promoter (P) is a small DNA segment located just upstream of the operator. It serves as the binding site for RNA polymerase. The repressor-binding site (O) and the promoter region (P) slightly overlap, so that when the repressor is bound to the DNA, RNA polymerase cannot bind to the promoter, and transcription is halted.

An inducer is a low-molecular-weight substance that binds to the repressor, converting it into an inactive form that is no longer capable of binding to the operator. Thus, in the lac system, the inducer is lactose; upon association with lactose, the repressor dissociates from the lac operator. Induction is a form of negative regulation, so named because transcription can proceed only after the repressor is removed. Another variation of negative regulation is known as catabolite repression.

Fig. 28.2.

Repression occurs when the repressor binds to the operator only in a complex with a low-molecular-weight cofactor (corepressor). Such a corepressor is often the end product of Protein Synthesis encoded by the operon. Consequently, if the concentration of this product becomes too high, it binds to the repressor, and its further synthesis is halted. An example of such a system is the Tryptophan operon.

The tryptophan operon consists of an operator and five structural genes (A–E). The latter encode enzymes involved in The Biosynthesis of tryptophan, one of the Essential Amino Acids. As the concentration of tryptophan increases, a point is reached where its further synthesis becomes undesirable, and transcription stops. The "switching off" of transcription occurs as follows. Tryptophan binds to a dimeric repressor (the trp repressor), which is encoded by a separate regulatory gene not part of the operon. This binding induces a conformational change, exposing a region capable of binding to the operator sequence in the DNA. The entire complex then binds to the DNA and blocks the RNA polymerase binding site (the promoter). This is an example of regulation via repression (pathway 1 in Fig. 28.1).

Transcript length variation is another mechanism through which The regulation of operon expression can be achieved in bacteria. For instance, in the trp operon, another regulatory system is utilized alongside repression. It involves the presence of a DNA region located immediately upstream of the first structural gene (trpE), known as the attenuator (pathway 2 in Fig. 28.1). The attenuator is a nucleotide sequence containing a signal that causes premature termination of transcription. In the case of the trp operon, when tryptophan concentrations are high, 90% of all transcripts are terminated after the transcription of just 140 NUCLEOTIDES, well before the Transcription of the structural trp genes begins.

Positive regulation is another mode of gene expression control (not shown in the figure). It differs from negative regulation in that transcription is "switched on" rather than "switched off" following the attachment of a regulatory protein to the operon. A corepressor, or rather a coactivator, binds to the activator protein; the entire complex then binds to the corresponding DNA site, and only then can transcription proceed. An example of this type of regulation is the process mediated by the catabolite activator protein, whose coactivator is cAMP (see Further Reading). In the absence of cAMP (or at low concentrations thereof), which occurs when bacteria proliferate in the presence of glucose, the transcription of certain operons—most notably the lac operon—is blocked. This phenomenon is termed catabolite repression.

EUKARYOTIC ORGANISMS are largely represented by multicellular forms with a high degree of cellular specialization. Although all human cells contain the exact same DNA, vastly different sets of genes are expressed in different Tissues. Therefore, mechanisms must exist whereby certain genes are active (i.e., expressed) while others are not. A variety of mechanisms serve this purpose: transcriptional and translational regulation, post-transcriptional and post-translational regulation, and hormonal regulation.

Transcriptional Regulation is exerted during mRNA synthesis. The average concentrations of individual mRNAs transcribed from different genes vary widely. This is because, for example, the mRNA copies of some genes degrade faster than others, or their synthesis occurs at a slower rate. Regulation can be mediated by DNA-binding proteins or even short RNA fragments that base-pair with DNA, presumably blocking RNA polymerase attachment sites. As a result, The rate of Transcription can be either decreased or increased.

Post-transcriptional regulation operates at the level of mRNA Processing. Even if the transcription rates of two different genes are identical, subsequent mRNA processing—including 5' and 3' end modification and exon splicing—may proceed differently for different mRNAs.

Translational regulation occurs by preventing mRNA from being used as a template for protein synthesis, even though it is present in the Cytoplasm. In the sea urchin oocyte, for instance, abundant mRNA is present, yet no significant protein synthesis occurs until the oocyte is fertilized. Only then do the mRNA molecules undergo modification—acquiring a 5' hairpin Structure (known as a cap) and a 3' poly(A) tail (Chapter 22)—allowing them to enter the normal translational pathway that culminates in the assembly of a protein molecule.

Post-translational regulation relies on the fact that many proteins are synthesized in an inactive form and must undergo subsequent modification. For example, pancreatic β-cells synthesize not Insulin itself, but its precursor, whose polypeptide chain is longer than that of insulin and contains an additional sequence of amino acid residues. Only after this sequence is cleaved by a proteolytic enzyme is the functional hormone produced. Thus, The production of the active hormone can be regulated post-translationally by modulating The activity of the proteolytic enzyme.

Hormonal regulation is a special case of transcriptional regulation; through this pathway, the Organism prompts a cell to "turn on" specific genes in response to an external stimulus. For instance, Steroid Hormones synthesized in specific cells ultimately reach the cytoplasm of their target cells, where a specialized transport protein carries them into The Nucleus. There, they can activate specific genes by directly interacting with Chromatin at designated sites. Each hormone activates its own unique set of genes. In this manner, the cell's response is matched to The Nature of the signal perceived via the corresponding hormone (Chapter 41).