Structural Organization of the Template
RNA synthesis takes place at strictly defined regions of the genome. The unit of transcription is the transcription unit (or transcription complex), which is a DNA fragment bounded by two key regulatory elements:
- Promoter — a specific nucleotide sequence (site) where RNA polymerase binds to the template and initiates assembly.
- Termination site — the region where the synthesis of the polynucleotide chain is completed.
A key feature of eukaryotes is that a single transcription unit generally contains only a single gene. This isolated arrangement allows the cell to individually transcribe different genes, tightly regulating their activity according to its needs.
Eukaryotic RNA Polymerases
In the nucleus of a eukaryotic cell, biosynthesis is catalyzed by massive multi-subunit enzymes known as RNA polymerases. They not only form phosphodiester bonds but also possess binding sites for regulatory factors. Based on the type of product synthesized, three types of enzymes are distinguished:
- RNA polymerase I — specializes in the synthesis of ribosomal RNA precursors (pre-rRNA).
- RNA polymerase II — is responsible for creating messenger RNA precursors (pre-mRNA).
- RNA polymerase III — catalyzes the assembly of transfer RNA precursors (pre-tRNA).
Building of the new chain proceeds according to the principle of complementarity. Adenine (A) in DNA pairs with uracil (U) in RNA, guanine (G) pairs with cytosine (C), and thymine (T) pairs with adenine (A).
Initiation Stage
The process is launched in the promoter region, where a critical role is played by the TATA-binding protein (TATA-box factor). It recognizes a specific nucleotide sequence ("-TATA-") and binds to it.
Binding of the TATA-box factor "activates" the promoter, facilitating its interaction with RNA polymerase. Docking of the enzymatic complex onto the DNA dramatically increases its affinity for initiation factors (A, B). Together, they achieve local unwinding of approximately one turn of the DNA double helix to gain direct access to the template strand for the start of polymerization.
Elongation and Termination
Following successful initiation, elongation begins — the active growth of the pre-RNA strand. Synthesis of the molecule always proceeds in an antiparallel fashion, in the 5′ to 3′ direction, while the enzyme itself moves along the template from the 3′ to 5′ end. At this stage, elongation factors (E, H, F) are recruited, which enhance enzyme activity and maintain local strand separation.
The dynamics of the DNA helix are such that ahead of the sliding polymerase, the turns unwind, and behind it, they rewind. The overall reaction equation is as follows:
aGTP + bATP + cUTP + mCTP → (DNA template, Mg2+, RNA polymerase) → RNA + (a+b+c+m)H4P2O7
The process concludes at the termination stage. When the complex reaches the termination site, local strand unwinding makes this region accessible to termination factors. They halt the polymerase, accelerating the release of the primary transcript and the dissociation of the enzyme from the DNA complex.