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Initially, the gene (double-stranded DNA) is copied or transcribed into an virtually identical single-stranded RNA molecule (mRNA). In eukaryotes this process occurs in the nucleus and is catalyzed by the enzyme RNA polymerase. This enzyme attaches itself to a portion of the DNA strand and replicates it, then sending out the mRNA to machinery in the cytoplasm that use the information contained in the gene to make a protein.
Transcription is very similar to DNA replication although different proteins are involved. The most important enzyme is RNA polymerase, an enzyme that influences the synthesis of RNA from a DNA template. For transcription to be initiated, RNA polymerase must be able to recognize the beginning sequence of a gene so that it knows where to start synthesizing an mRNA. It is directed to this initiation site by the ability of one of its subunits to recognize a specific DNA sequence found at the beginning of a gene, called the promoter sequence. The promoter sequence is a unidirectional sequence found on one strand of the DNA that instructs the RNA polymerase in both where to start synthesis and in which direction synthesis should continue. The RNA polymerase then unwinds the double helix at that point and begins synthesis of a RNA strand complementary to one of the strands of DNA. This strand is called the antisense or template strand, whereas the other strand is referred to as the sense or coding strand. Synthesis can then proceed in a unidirectional manner.
Although much is known about transcript processing, the signals and events that instruct RNA polymerase to stop transcribing and drop off the DNA template remain unclear. Experiments over the years have indicated that processed eukaryotic messages contain a poly(A) addition signal (AAUAAA) at their 3' end, followed by a string of adenines. This poly(A) addition, also called the poly(A) site, contributes not only to the addition of the poly(A) tail but also to transcription termination and the release of RNA polymerase from the DNA template. Yet, transcription does not stop here. Rather, it continues for another 200 to 2000 bases beyond this site before it is aborted. It is either before or during this termination process that the nascent transcript is cleaved, or cut, at the poly(A) site, leading to the creation of two RNA molecules. The upstream portion of the newly formed, or nascent, RNA then undergoes further modifications, called post-transcriptional modification, and becomes mRNA. The downstream RNA becomes unstable and is rapidly degraded.
Although the importance of the poly(A) addition signal has been established, the contribution of sequences further downstream remains uncertain. A recent study suggests that a defined region, called the termination region, is required for proper transcription termination. This study also illustrated that transcription termination takes place in two distinct steps. In the first step, the nascent RNA is cleaved at specific subsections of the termination region, possibly leading to its release from RNA polymerase. In a subsequent step, RNA polymerase disengages from the DNA. Hence, RNA polymerase continues to transcribe the DNA, at least for a short distance.
During transcription there needs to be a promoter. A promoter is a certain sequence of DNA which RNA polymerase can bind to. It creates a very tight bond with the promoter. There are at least one promoter for every gene in the genome.
Promoters do three specific jobs. They tell RNA:
- Where to start transcription
- Which strand of DNA to read
- The direction to take from the start.
After the specific promoter has been chosen by the RNA polymerase a process called elongation begins. This process is where the polymerase adds nucleotides (A, U, C, G) to a section of DNA of about 20 amino acids and replicates it. However it is created antiparallel to DNA. DNA is 5' to 3' and RNA transcribes as 3' to 5'. The elongation process will continue until it reaches a certain termination site in the DNA. There is also a specific initiation site the tells where the transcription is to start taking place.
RNA can be spliced to increase the number of proteins it can create. A pre-mRNA (in-mature mRNA) can go through a process of splicing. In the cases of the protein tropomyosin a pre-mRNA can be spliced to create five different end result mature mRNA's. This is the reason why the human body only has 21,000 genes rather than the 100,000 to 150,000 genes estimated by scientists. This type of editing greatly increases the number of mature mRNAs and therefore the number of proteins that can be created from a single gene.
Another way to further alter RNA is called RNA editing. This happens through two separate ways. The first way is called Intersection of Nucleotides. When this happens stretches of U's will be placed into the mRNA to make it longer. An example of this is:
(DNA) ACCTCC - becomes transcribed (mRNA) UGGAGG - becomes edited (Edited mRNA) UGGUUUAGG
The edit above will effectively alter the amino acid sequence from trp-arg, to trp-phe-arg.
The second way for editing to happen is called Alteration of Nucleotides. In this process an enzyme will catalyze and change the way the protein is formed. An example of this process is:
(DNA) GTA - becomes transcribed (mRNA) CAU - becomes edited (Edited mRNA) UAU