Transcription

DNA serves as the template for the synthesis of RNA much as it does for its own replication. The Steps Some 50 different protein transcription factors bind to promoter sites, usually on the 5' side of the gene to be transcribed. An enzyme, an RNA polymerase, binds to the complex of transcription factors. Working together, they open the DNA double helix. The RNA polymerase proceeds to "read" one strand moving in its 3' ? 5' direction. In eukaryotes, this requires — at least for protein-encoding genes — that the nucleosomes in front of the advancing RNA polymerase (RNAP II) be removed. A complex of proteins is responsible for this. The same complex replaces the nucleosomes after the DNA has been transcribed and RNAP II has moved on. As the RNA polymerase travels along the DNA strand, it assembles ribonucleotides (supplied as triphosphates, e.g., ATP) into a strand of RNA. Each ribonucleotide is inserted into the growing RNA strand following the rules of base pairing. Thus for each C encountered on the DNA strand, a G is inserted in the RNA; for each G, a C; and for each T, an A. However, each A on the DNA guides the insertion of the pyrimidine uracil (U, from uridine triphosphate, UTP). There is no T in RNA. Quality control. Occasionally RNA polymerase will select and insert an incorrect, mismatched, ribonucleotide. When this occurs in bacteria (and perhaps in all organisms), the enzyme backs up, removes the incorrect nucleotide (and the one preceding it) and tries again. (Described by Zenkin et al., in the 28 July 2006 issue of Science.)

Synthesis of the RNA proceeds in the 5' ? 3' direction. As each nucleoside triphosphate is brought in to add to the 3' end of the growing strand, the two terminal phosphates are removed. When transcription is complete, the transcript is released from the polymerase and, shortly thereafter, the polymerase is released from the DNA. Note that at any place in a DNA molecule, either strand may be serving as the template; that is, some genes "run" one way, some the other (and in a few remarkable cases, the same segment of double helix contains genetic information on both strands!). In all cases, however, RNA polymerase transcribes the DNA strand in its 3' ? 5' direction.

Prokaryotic vs. eukaryotic transcription

Prokaryotic transcription occurs in the cytoplasm alongside translation. Eukaryotic transcription is primarily localized to the nucleus, where it is separated from the cytoplasm by the nuclear membrane. The transcript is then transported into the cytoplasm where translation occurs. Another important difference is that eukaryotic DNA is wound around histones to form nucleosomes and packaged as chromatin. Chromatin has a strong influence on the accessibility of the DNA to transcription factors and the transcriptional machinery including RNA polymerase. In prokaryotes, mRNA is not modified. Eukaryotic mRNA is modified through RNA splicing, 5' end capping, and the addition of a polyA tail.

Reverse transcription

Some viruses (such as HIV, the cause of AIDS), have the ability to transcribe RNA into DNA. HIV has an RNA genome that is duplicated into DNA. The resulting DNA can be merged with the DNA genome of the host cell. The main enzyme responsible for synthesis of DNA from an RNA template is called reverse transcriptase. In the case of HIV, reverse transcriptase is responsible for synthesizing a complementary DNA strand (cDNA) to the viral RNA genome. An associated enzyme, ribonuclease H, digests the RNA strand, and reverse transcriptase synthesises a complementary strand of DNA to form a double helix DNA structure. This cDNA is integrated into the host cell's genome via another enzyme (integrase) causing the host cell to generate viral proteins which reassemble into new viral particles. Subsequently, the host cell undergoes programmed cell death (apoptosis).

Some eukaryotic cells contain an enzyme with reverse transcription activity called telomerase. Telomerase is a reverse transcriptase that lengthens the ends of linear chromosomes. Telomerase carries an RNA template from which it synthesizes DNA repeating sequence, or "junk" DNA. This repeated sequence of "junk" DNA is important because every time a linear chromosome is duplicated, it is shortened in length. With "junk" DNA at the ends of chromosomes, the shortening eliminates some repeated, or junk sequence, rather than the protein-encoding DNA sequence that is further away from the chromosome ends. Telomerase is often activated in cancer cells to enable cancer cells to duplicate their genomes without losing important protein-coding DNA sequence. Activation of telomerase can be part of the process that allows cancer cells to become immortal.