thumb|Pre-mRNA is the first form of RNA created through transcription in protein synthesis. The pre-mRNA lacks structures that the messenger RNA (mRNA) requires. First all introns have to be removed from the transcribed RNA through a process known as splicing. Before the RNA is ready for export, a Poly(A)tail is added to the 3' end of the RNA and a 5' cap is added to the 5' end.

thumb|Micrograph of gene transcription of ribosomal RNA illustrating the growing primary transcripts

A primary transcript is the single-stranded ribonucleic acid (RNA) product synthesized by transcription of DNA, and processed to yield various mature RNA products such as mRNAs, tRNAs, and rRNAs. The primary transcripts designated to be mRNAs are modified in preparation for translation. For example, a precursor mRNA (pre-mRNA) is a type of primary transcript that becomes a messenger RNA (mRNA) after processing.

Pre-mRNA is synthesized from a DNA template in the cell nucleus by transcription. Pre-mRNA comprises the bulk of heterogeneous nuclear RNA (hnRNA). Once pre-mRNA has been completely processed, it is termed "mature messenger RNA", or simply "messenger RNA". The term hnRNA is often used as a synonym for pre-mRNA, although, in the strict sense, hnRNA may include nuclear RNA transcripts that do not end up as cytoplasmic mRNA.

There are several steps contributing to the production of primary transcripts. All these steps involve a series of interactions to initiate and complete the transcription of DNA in the nucleus of eukaryotes. Certain factors play key roles in the activation and inhibition of transcription, where they regulate primary transcript production. Transcription produces primary transcripts that are further modified by several processes. These processes include the 5' cap, 3'-polyadenylation, and alternative splicing. In particular, alternative splicing directly contributes to the diversity of mRNA found in cells. The modifications of primary transcripts have been further studied in research seeking greater knowledge of the role and significance of these transcripts. Experimental studies based on molecular changes to primary transcripts and the processes before and after transcription have led to greater understanding of diseases involving primary transcripts.

Production

The steps contributing to the production of primary transcripts involve a series of molecular interactions that initiate transcription of DNA within a cell's nucleus. Based on the needs of a given cell, certain DNA sequences are transcribed to produce a variety of RNA products to be translated into functional proteins for cellular use. To initiate the transcription process in a cell's nucleus, DNA double helices are unwound and hydrogen bonds connecting compatible nucleic acids of DNA are broken to produce two unconnected single DNA strands. One strand of the DNA template is used for transcription of the single-stranded primary transcript mRNA. This DNA strand is bound by an RNA polymerase at the promoter region of the DNA.

thumb|Transcription of DNA by RNA polymerase to produce primary transcript

In eukaryotes, three kinds of RNA—rRNA, tRNA, and mRNA—are produced based on the activity of three distinct RNA polymerases, whereas, in prokaryotes, only one RNA polymerase exists to create all kinds of RNA molecules. RNA polymerase II of eukaryotes transcribes the primary transcript, a transcript destined to be processed into mRNA, from the antisense DNA template in the 5' to 3' direction, and this newly synthesized primary transcript is complementary to the antisense strand of DNA. Activation of transcription depends on whether or not the transcription elongation complex, itself consisting of a variety of transcription factors, can induce RNA polymerase to dissociate from the Mediator complex that connects an enhancer region to the promoter.

Histone modification by transcription factors is another key regulatory factor for transcription by RNA polymerase. In general, factors that lead to histone acetylation activate transcription while factors that lead to histone deacetylation inhibit transcription. Acetylation of histones induces repulsion between negative components within nucleosomes, allowing for RNA polymerase access. Deacetylation of histones stabilizes tightly coiled nucleosomes, inhibiting RNA polymerase access. In addition to acetylation patterns of histones, methylation patterns at promoter regions of DNA can regulate RNA polymerase access to a given template. RNA polymerase is often incapable of synthesizing a primary transcript if the targeted gene's promoter region contains specific methylated cytosines— residues that hinder binding of transcription-activating factors and recruit other enzymes to stabilize a tightly bound nucleosome structure, excluding access to RNA polymerase and preventing the production of primary transcripts.

Transcription stress

DNA damages arise in each cell, every day, with the number of damages in each cell reaching tens to hundreds of thousands, and such DNA damages can impede primary transcription. The process of gene expression itself is a source of endogenous DNA damages resulting from the susceptibility of single-stranded DNA to damage. Otherwise stated, the newly synthesized primary transcripts are modified in several ways to be converted to their mature, functional forms to produce different proteins and RNAs such as mRNA, tRNA, and rRNA.

Processing

The basic primary transcript modification process is similar for tRNA and rRNA in both eukaryotic and prokaryotic cells. On the other hand, primary transcript processing varies in mRNAs of prokaryotic and eukaryotic cells.

In complex eukaryotic cells, one primary transcript is able to prepare large amounts of mature mRNAs due to alternative splicing. Alternative splicing is regulated so that each mature mRNA may encode a multiplicity of proteins. thumb|Alternative splicing of the primary transcript The effect of alternative splicing in gene expression can be seen in complex eukaryotes which have a fixed number of genes in their genome yet produce much larger numbers of different gene products.

In Drosophila and Aedes, hnRNA (pre-mRNA) size was larger in Aedes due to its larger genome, despite both species producing mature mRNA of similar size and sequence complexity. This indicates that hnRNA size increases with genome size.

In HeLa cells, spliceosome groups on pre-mRNA were found to form within nuclear speckles, with this formation being temperature-dependent and influenced by specific RNA sequences. Pre-mRNA targeting and splicing factor loading in speckles were critical for spliceosome group formation, resulting in a speckled pattern.

Recruiting pre-mRNA to nuclear speckles significantly increased splicing efficiency and protein levels, indicating that proximity to speckles enhances splicing efficiency.

Research has also led to greater knowledge about certain diseases related to changes within primary transcripts. One study involved estrogen receptors and differential splicing. The article entitled, "Alternative splicing of the human estrogen receptor alpha primary transcript: mechanisms of exon skipping" by Paola Ferro, Alessandra Forlani, Marco Muselli and Ulrich Pfeffer from the laboratory of Molecular Oncology at National Cancer Research Institute in Genoa, Italy, explains that 1785 nucleotides of the region in the DNA that codes for the estrogen receptor alpha (ER-alpha) are spread over a region that holds more than 300,000 nucleotides in the primary transcript. Splicing of this pre-mRNA frequently leads to variants or different kinds of the mRNA lacking one or more exons or regions necessary for coding proteins. These variants have been associated with breast cancer progression. In the life cycle of retroviruses, proviral DNA is incorporated in transcription of the DNA of the cell being infected. Since retroviruses need to change their pre-mRNA into DNA so that this DNA can be integrated within the DNA of the host it is affecting, the formation of that DNA template is a vital step for retrovirus replication. Cell type, the differentiation or changed state of the cell, and the physiological state of the cell, result in a significant change in the availability and activity of certain factors necessary for transcription. These variables create a wide range of viral gene expression. For example, tissue culture cells actively producing infectious virions of avian or murine leukemia viruses (ASLV or MLV) contain such high levels of viral RNA that 5–10% of the mRNA in a cell can be of viral origin. This shows that the primary transcripts produced by these retroviruses do not always follow the normal path to protein production and convert back into DNA in order to multiply and expand.

See also

  • cis-splicing
  • Outron
  • trans-splicing
  • Transcription (biology)
  • Transcriptome

References

  • Scienceden.com RNA Article