See also RNA capping and splicing

Transcription in eukaryotic cells occurs in the nucleus followed by translation in the cytoplasm.

Transcription is the transfer of information from a double stranded DNA molecule to a single RNA molecule. Like DNA, RNA is a linear polymer made up of four types of nucleotides linked together by phosphodiester bonds. It differs from DNA in that (1) its sugar units are riboses (OH attached to the 2’C) rather than deoxyriboses, (2) uracil is substituted for the base thymine (uracil lacks the methyl group which thymine has), (3) it is single stranded except in some viruses and (4) it does not form a double helix although it can fold back onto itself to form double helical regions.

Most of the genes which are transcribed into RNA specify the amino acid sequence of proteins. The RNA which is transcribed from such genes is referred to as messenger RNA (mRNA). However, the final product after transcription of many genes is not protein but rather the RNA itself. Like proteins, these RNAs serve as enzymatic and structural components for a wide variety of processes in the cell. 3 major classes of such RNAs are small nuclear RNA (snRNA) which direct the splicing of pre-mRNA to form mRNA, ribosomal RNA (rRNA) which form the core of ribosomes and transfer RNA (tRNA) molecules which serve in translation of the mRNA. Transfer RNA molecuels have amino acids coavlently attached to one end and an anticodon that can base pair with an mRNA codon at the other end.

Because DNA is double-stranded and RNA is single-stranded, only one of the two DNA strands needs to be copied. The strand that is copied is called the template strand. The RNA transcript’s sequence is complementary to the template strand. The strand of DNA not used as a template is called the coding or sense strand becasue it has the same “sense” as the RNA. It has the same sequence as the RNA transcript, except that U is used instead of T. The template strand is referred to as the anti-sense strand.

Another type of RNA is the SRP RNA. In eukaryotes, where some proteins are synthesized by ribosomes on the roughg ER, this process is mediated by the signal recognition particle or SRP that contains both RNA and proteins.

Yet another type of RNA are small RNAs which includes both micro RNA (miRNA) and small interfering RNA (siRNA) which are involved in the control of gene expression.

Transcription starts with the opening and unwinding of a small portion of the DNA double helix to expose the bases on each DNA strand. Transcription runs in the 5′ to 3′ direction so that the RNA produced by transcription starts from the 3′ end. In contrast to DNA replication, the RNA single strand product does not remain hydrogen bonded to the DNA template but is rather displaced just behind the region where the ribonucleotides are being added. This release means that many RNA copies can be made from the same gene.

The enzymes which transcribe DNA are called RNA polymerases. They differ from DNA polymerases in that 1) they catalyze the linkage of ribonucleotides, not deoxyribonucleotides, 2) they can start an RNA chain without a primer and 3) they lack the high fidelity or accuracy of DNA polymerases (there is an error rate of about 1 of 104 nucleotides copies into RNA in comparison of 1 of 107 by DNA polymerase).

Eucaryotics have 3 types of RNA polymerases which transcribe different types of genes. RNA polymerase I transcribes the 5.8S, 18S and 28S rRNA genes. RNA polymerase II transcribes all the protein coding genes (mRNA) along with some small nucleolar RNAs (snoRNAs) and small nuclear snRNA genes. RNA polymerase III transcribes tRNA genes, 5S rRNA genes and some other snRNA genes.

RNA polymerase II requires the help of a large set of proteins called general transcription factors which must assemble at the promoter with the polymerase before transcription can begin. The proteins are called “general” because they must assemble on all promoters used by RNA polymerase II. The assembly starts with the binding of a short double helical DNA sequence composed mainly of T and A nucleotides about 25 nucleotides upstream from the transcription start site, referred to as the TATA box by a subunit, TBP (for TATA box binding protein), of one of the general transcription factors, TFIID (for transcription factor II). After RNA polymerase II has been guided onto the promoter DNA to form a transcription initiation complex, it gains access to the template strand by another general transcription factor, TFIIH which contains a DNA helicase. Some eukaryotes also use a GC box rather than a TATA box which serve as promoter sequences for the transcription factor SP1. Compare Bacteria

Transcription in the eukaryotic cell is very complex and requires more proteins than it does on purified DNA. Even other regulatory proteins called enhancers or transcriptional activators bind to specific sequences in DNA sometimes several thousand nucleotide pairs away from the transcription start site to help RNA polymerase and the general factors assemble at the promoter. The enhancers which can be located anywhere with respect to the gene (5′ of the promoter, 3′ of the gene or even in an intron of the gene). In addition, enhancers attract ATP-dependent chromatin remodeling complexes and histone acetylases which allow greater accessibility to the DNA present in chromatin and thereby facilitate the assembly of the transcription ininitiation machinery on DNA.

These proteins bind to specific nucleotide sequences within promoters and enhancers and act either to enhance or suppress their activity. Such proteins have been identified by a variety of techniques like DNA footprinting.

Once transcription has started, RNA polymerase also require elongation factors which are proteins that help polymerases move along the DNA template. In order to deal with DNA supercoiling (1 large DNA supercoil will form to compensate for each 10 nucleotide pairs that are unwound, DNA topisomerase enzymes are necessary to remove this tension. Click Here to see what happens to a mRNA after transcription.

Ribosomal RNA genes exist in multiple copies in order to produce necessary quantities of rRNA. This is important because, unlike mRNA, rRNAs are not translated. The final produce from transcription in the case of rRNA is the rRNA itself and thus amplification can not be achieved through multiple rounds of amplification. There are 4 types of eukaryotic rRNA. 3 of the 4 types (18S, 5.8S, and 28S) which are transcribed by RNA polymerase I, are made by chemically modifying and cleaving a single large precursor rRNA and the 4th (5S) is transcribed by a different polymerase RNA polymerase III. The specific positions at which chemical modifications are made as well as the cleavage of the precursor rRNA into the mature rRNA are made by a large class of RNAs called small nucleolar RNAs (snoRNAs) so named because they perform their functions in a subcompartment of the nucleus called the nucleolus which is a large aggregate of macromolecules including the rRNA genes, snoRNPs and other proteins.

Prokaryotic Transcription:

Unlike eukaryotes, bacteria have a single RNA polymerase. Accurate initiation of transcription requires two sites in DNA; one called a promoter that forms a recognition and binding site for the RNA polymerase and then the actual start site. The polymerase also needs a termination site.

The first based transcribed is referred to as +1. Bases upstream of the start site receive negative numbers starting at -1. The promoter is a short sequence found upstream of the start site; it is not transcribed by the polymerase. Two 6 base sequences are common to bacterial promoters; one is located 35 nt upstream and the other is located 10 nt upstream of the start site. The binding of RNA polymerase to the promoter is the first step in transcription. Promoter binding is controlled by the alpha subunit of the RNA polymerase holoenzyme, which recognizes the -35 sequence in the promoter and positions the RNA polymerase at the correct start site, oriented to transcribe in the correct direction.  Once bound to the promoter, the RNA polymerase begins to unwind the DNA helix at the -10 site. The polymerase covers a region of about 75 bp but unwinds only about 12-14 bp.

The region containing the RNA polymerease, the DNA template and the growing RNA trasncript is called the transcription bubble because it contains a locally unwound “bubble” of DNA.

The end of a bacterial transcripton unit is makred by terminator sequences that singal “stop” to the polymerease. The simplest temrinators consist of a series of G-C base pairs followed by a series of A-T base pairs. The RNA transcript of this stop region can form a double stranded structure in the GC region called a hairpin, which is followed by four or more uracil ribonucleotides. Formation of the hairpin casues the RNA polyemrease ot pause, placing it direclty over the fun of four uracils. The pairing of U with the DNA’s A is the weakest of the four hybrid base pairs and it is not strong enoguh to hold the hybrid strands when the polymerase pauses.

In prokaryotes, the mRNA produced by transcripion begins to be translated before transcription is finished. Thus, transciption and translation are ocupled. As soon as a 5′ end of the mRNA becomes availabe, ribosomes are loaded onto this to being translation.  This is not the case with eukaryotes where transcription occurs in the nucleus and is followed by translation in the cytoplasm.