Explain the structure of the eukaryotic mRNA that exits the nucleus

The entire journey of DNA to protein starts with the synthesis of messenger RNA, which is one of the critical intermediators in carrying the genetic instructions from nucleus to cytoplasm. This is initiated when RNA polymerase binds to DNA at a specific promoter region because of the assistance of general transcription factors.

Heterogeneous Nuclear RNA Formation

Whereas mammalian cells are treated briefly with either radioactive [3H]uridine or [32P]phosphate under the condition of steady state, the majority of the radioactivity is inserted into huge heterogeneous RNA molecules that are confined in the nucleus. These are termed heterogeneous nuclear RNAs, hnRNAs, which have a high molecular weight up to 80S, or 50,000 nucleotides-and are heterogeneous. Since these cells are then placed into an unlabeled medium, the radioactivity eventually shifts from the large nuclear RNAs into smaller mRNAs appearing in the cytoplasm. This provides definitive proof that hnRNAs are precursors to mRNAs .

The Machinery of mRNA Transcription

RNA polymerase II is the enzyme responsible for synthesizing all mRNA precursors in eukaryotic cells. The enzyme itself has about a dozen different subunits, conserved from yeast to mammals. RNA polymerase II forms a preinitiation complex PIC at the promoter along with GTFs. Components of interest in this pathway include:

TATA-Binding Protein: It is a protein that binds to the TATA box of the promoter region; it causes extreme DNA distortion and initiates PIC assembly.

TATA-Binding Protein complex: The complex includes TBP, which is a part of TFIID that binds to other promoter regions.

Preinitiation Complex Assembly: Following TBP binding, other GTFs (TFIIA, TFIIB) and RNA polymerase II are recruited to the promoter. TFIIH is an enzymatically active GTF responsible for DNA unwinding and phosphorylating RNA polymerase II, thus activating it to transcribe.

The preinitiation complex positions RNA polymerase II to initiate transcription. During elongation, selected GTFs remain bound at the promoter for subsequent rounds of transcription.

CTD Phosphorylation

One of the hallmarks in the structure of this enzyme is the carboxyl-terminal domain of RNA polymerase II, which consists of multiple copies of a heptapeptide sequence. Phosphorylation of specific residues, especially serines within the CTD mediated by various protein kinases, including TFIIH, is one of the most important signals that can commit RNA polymerase II to an active transcriptional form.

This elaborate process therefore denotes the sophisticated machinery of mRNA synthesis required for the regulation of gene expression in eukaryotic cells.

The Role of CTD Phosphorylation in mRNA Processing

The phosphorylation of the CTD of RNAPII is a dynamic process during transcription, which is integral to the mRNA processing pathway. The early phosphorylation event at Ser5 positions within the CTD results in the dissociation of the enzyme from the general transcription factors and promoter DNA, thus enabling RNAPII to proceed along the DNA template. Such modifications also attract proteins that are associated with the initial steps of mRNA processing, including 5' cap formation.

As transcription proceeds, a distinct kinase, P-TEFb phosphorylates the CTD on serines at position #2. This phosphorylation switch enables the recruitment of other processing factors responsible for RNA splicing and polyA addition that allow maturation of the mRNA. The CTD provides a platform from which these processing factors are recruited and released in a highly coordinated and complex series of steps to mature the mRNA. An elongating RNA polymerase II can carry more than 50 components and has a mass greater than 3 million daltons.

Termination of Transcription

In contrast to bacterial systems, where transcription termination is intimately coupled to the formation of mRNA 3' termini, eukaryotic transcription does not utilize specific termination sequences on the DNA. RNA polymerase II can read through the eventual 3' end of the mature mRNA, requiring post-transcriptional processing steps to complete the mRNA 3' terminus.

mRNA Structure

Eukaryotic mRNAs have the following features:

Continuous Coding Sequence: They encode for a specific polypeptide sequence.

mRNA is present in the cytoplasm, which is where translation occurs. The mRNAs are associated with ribosomes during their translation.

Noncoding Regions: Most mRNAs have noncoding regions at the 5' and 3' ends that are not translated into proteins but are essential for regulation purposes. An example of this is globin mRNAs, which contain about 25% of these types of noncoding regions.

Understanding these aspects of mRNA structure and processing are essential to understanding how eukaryotic cells carry out gene regulation.

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