Genes direct the synthesis of a functional product, such as protein or tRNA, rRNA, etc. It is known as gene expression. The information present in the DNA is used to synthesise proteins or other molecules.
Gene expression occurs in two main steps, i.e. transcription and translation. First, the information present in DNA is used to synthesise the complementary RNA molecule and then a polypeptide chain is synthesised using the RNA template. This way a gene is expressed in the form of a functional product.
Gene expression is tightly regulated at various steps and it controls the type as well as the amount of proteins synthesised. Gene expression can be regulated at the transcription or translation level.
Mechanism of Gene Expression
Genes are the short stretch of DNA that code for a functional product. Genes are the basic functional unit of inheritance. They are passed on from parents to offspring. The information contained in the DNA is referred to as ‘genotype’ and the expression of genes in terms of functional products or observable traits is referred to as ‘phenotype’. Gene expression leads to the formation of proteins and also to the synthesis of non-coding RNA, such as tRNA, rRNA, etc. All three main RNAs are required for the synthesis of proteins.
Genes govern all cellular functions. There are thousands of genes present in an organism that decide the fate and function of a cell. A gene is expressed in a cell to perform one or many functions. It helps a cell respond to various internal and external changes. Genes control the synthesis of proteins and proteins control the structure, metabolic functions and development of an organism.
The two steps involved in the synthesis of a protein are transcription and translation.
Transcription
It is the process by which RNAs are synthesised using DNA templates. The enzyme RNA polymerase catalyses the synthesis of RNA. It is a DNA-dependent enzyme. There are multiple RNA polymerases in eukaryotes that synthesise different RNA molecules. The RNA polymerase II catalyses the synthesis of pre-mRNA or hnRNA that acts as a template for protein synthesis.
The synthesis of RNA takes place in the 5’ to 3’ direction and the DNA strand with polarity 3’ to 5’ acts as a template. The RNA thus produced, contains the same sequence as the 5’ to 3’ strand (coding strand) of the DNA, except for thymine that is replaced by uracil in RNA.
The process of transcription is a three-step process:
- Initiation – The RNA polymerase binds to the promoter that is present towards the 5’-end of the structural gene (coding strand) to initiate transcription.
- Elongation – Nucleoside triphosphates are used as a substrate and each base is added as per the complementary base sequence in the template strand of the DNA.
- Termination – When the RNA polymerase reaches the terminator region present downstream of the structural gene, the nascent RNA and RNA polymerase are released and the transcription ends.
In eukaryotic cells, the RNA polymerase I transcribes rRNAs, RNA polymerase II transcribes pre-mRNA transcript and RNA polymerase III transcribes tRNA, 5srRNA and snRNA.
All three RNAs, i.e. mRNA, rRNA and tRNA are required for protein synthesis, but mRNA acts as a template for translation. The hnRNA or pre-mRNA is further processed to form a mature and fully functional mRNA that is ready for protein synthesis. The mRNA undergoes splicing, capping and tailing in the nucleus. The fully processed mature mRNA is transported out of the nucleus for protein synthesis.
Translation
Translation is the process of protein synthesis using mRNA as a template. The three base sequences or a codon in the mRNA codes for one amino acid and this way the information contained in mRNA is translated to a polypeptide chain of amino acids. The sequence of amino acids in the polypeptide chain is defined by the sequence of bases in the mRNA transcript.
The gene or a DNA segment that codes for a protein is known as a cistron. In eukaryotes, genes are generally monocistronic, i.e. structural gene codes for one mRNA and one protein. In prokaryotes, structural genes are generally polycistronic, i.e. multiple mRNAs and proteins are synthesised using the same promoter and operator.
The process of translation starts with the addition of an amino acid to the specific tRNA (charging of tRNA) that acts as an adapter molecule. tRNA brings amino acids to ribosomes for protein synthesis. Translation is also carried out in three steps, i.e. initiation, elongation and termination.
- Initiation – The process of translation initiates with the binding of ribosomal subunits and the recognition of the start codon (AUG) by initiator tRNA (Met-tRNA).
- Elongation – The polypeptide chain continues to grow as specific amino acid-tRNAs sequentially bind to mRNA codons and amino acids are added to the growing polypeptide chain by peptide bonds.
- Termination – Translation terminates on reaching one of the stop codons (UAA, UAG or UGA) and the polypeptide chain is released.
Regulation of Gene Expression
Regulation of gene expression is required for controlling the amount, location, type and timing of functional products synthesis. It is necessary for the proper functioning and maintenance of cellular machinery. It enables a cell to produce the specific gene product, in the required quantity and at the right time. Therefore, it enables cells to respond to changing environments. Genes are regulated according to physiological, metabolic and environmental conditions.
Control of gene expression forms the basis of cellular differentiation, i.e. how different cells mature to perform different functions even after having the same genome. Some genes are switched on/off in one cell and some in another cell. It is a key to development, homeostasis, morphogenesis and also for evolution. The development of an embryo to an adult is a result of coordinated regulation of the expression of various gene sets.
Regulation of gene expression can occur at various levels:
- At the time of formation of RNA, i.e. transcription.
- At the time of protein synthesis, i.e. translation.
- In eukaryotes, gene expression is also regulated at the time of processing and exporting of mRNA.
Transcription initiation is an efficient and primary control point. There are regulatory sequences that are present upstream or downstream to the promoter region of a transcription unit. The accessibility of the promoter site to which the RNA polymerase binds and initiates the transcription is regulated by accessory proteins. The regulatory proteins bind to the operator region that is present adjacent to the promoter region. The regulatory protein can be both an activator as well a repressor. If a repressor protein binds to the promoter, it inhibits the binding of RNA polymerase to the promoter site and thereby hinders transcription.
Example of Gene Expression and Regulation
The lac operon provides a classic example of a transcriptionally regulated system in prokaryotes. It is a polycistronic gene, wherein a number of mRNAs are transcribed from structural genes having a common promoter and operator regions. Mostly, genes in an operon work in tandem or have related functions.
The lac operon consists of three structural genes z, y and a, that are regulated by a regulatory gene i.
- z gene – codes for β-galactosidase.
- y gene – codes for permease.
- a gene – codes for transacetylase.
- i gene – codes for inhibitor or repressor protein of the lac operon.
Beta-galactosidase catalyses the hydrolysis of lactose into glucose and galactose. Permease increases the cell permeability to β-galactosides and transacetylase transfers an acetyl group from acetyl-CoA to β-galactosides. The repressor protein binds to the operator region and inhibits the transcription of the lac operon.
In E.coli, β-galactosidase is required for the hydrolysis of lactose. Hence, lactose acts as an inducer of the lac operon. When lactose is present as the source of energy, the operon is switched on, but when lactose is not present, bacteria do not need the enzyme β-galactosidase. Hence, the lac operon is switched off.
Negative Regulation
Normally, the lac operon is switched off due to the continuous formation of repressor protein from the i gene. The repressor binds to the operator region and makes the promoter region inaccessible to RNA polymerase, thereby inhibiting the transcription of z, y and a gene. When an inducer, such as lactose or allolactose are present, then they act as inducers of the operon. The inducer binds to the repressor and inactivates it, thereby enabling the RNA polymerase to access the promoter site for transcription of the genes. Here, the substrate of the enzyme regulates the synthesis of the enzyme. This regulation of a gene by the repressor is known as negative regulation.
Positive Regulation
The lac operon is also under positive regulation by the absence of glucose catabolites. When glucose level is low, the levels of cAMP are high and they readily bind to catabolite activator protein (CAP). The cAMP-CAP complex binds to the region that is upstream to the operator of the lac operon. It facilitates the binding of RNA polymerase and enhances the gene expression of the lac operon.
When glucose level is high, the cAMP level is decreased due to the presence of glucose catabolites. This decreases the efficiency of binding of RNA polymerase and thereby decreases the gene expression of lac operon even when lactose is present.
This is how lac operon is regulated and operated to enable bacteria to use glucose or lactose as a source of energy.
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