mRNA
The mRNA is then exported into the cytoplasm through nuclear pores . There protein biosynthesis takes place on ribosomes . Here, the nucleotide sequence of the mRNA is translated into the amino acid sequence of a polypeptide ( translation ) using tRNAs . An mRNA can be used several times; eventually it will be dismantled.
A mRNA ( English messenger RNA ), including messenger RNA called, is a single-stranded ribonucleic acid (RNA) as a transcript of a gene associated with partial section of the deoxyribonucleic acid (DNA) is the genetic information for the structure of a protein in a cell contains.
The sequence of nucleobases ( base sequence ) of certain sections of the DNA molecules of cells carries the information for the construction of proteins . It is expressed in that it is transcribed into RNA molecules and read and translated as a message from an mRNA to ribosomes . For this purpose, an mRNA with a specific nucleotide sequence is created ( transcription ) using the DNA template under the enzymatic effect of an RNA polymerase . During translation, it serves as a template for ribosomal protein biosynthesis and, as the coding sequence, then specifies the sequence of amino acids ( amino acid sequence ) of the polypeptide chain to be formed, i.e. the primary structure of a specific protein.
In the case of drugs based on mRNA, cells then use the information brought in to produce the protein that is effective in each case. This can act as an antigen in RNA vaccines , for example .
transcription
During transcription , a section of the DNA is rewritten into a single RNA strand. This happens under the action of the enzyme RNA polymerase . The codogenic DNA strand serves as a template for the construction of an RNA strand, which then codes as mRNA for protein . In prokaryotes (living beings without a nucleus) this process of transcription takes place in the cytoplasm , in eukaryotes (living beings with a nucleus) the nuclear genome is transcribed in the karyoplasm of the nucleus of a cell.
The actual transcription process by an RNA polymerase is the same in prokaryotes and eukaryotes. In prokaryotic cells (procytes), however, ribosomes can already attach to the mRNA that has not yet been fully synthesized and begin translation . In this way, the synthesis of proteins can start at the same time during transcription , which enables special forms of gene regulation (see attenuation ). In eukaryotes, on the other hand, the primary RNA transcript (pre-mRNA) is first subjected to various processes in the cell nucleus (processed, see below ) and only then exported as mRNA from the nucleus into the cytoplasm, where the ribosomes are located.
Prokaryotes have only one type of enzyme, RNA polymerase, which is used to build an RNA polynucleotide . Eukaryotes, on the other hand, have different types of RNA polymerases; their RNA polymerase II catalyzes the synthesis of pre-mRNA.
| enzyme | functional RNA |
|---|---|
| RNA polymerase I. | rRNA |
| RNA polymerase II | pre-mRNA (among others) |
| RNA polymerase III | tRNA (among others) |
| RNA polymerase IV, V | siRNA |
An essential difference between prokaryotic and eukaryotic mRNA is that prokaryotic mRNA is mostly polycistronic , while eukaryotic mRNA is usually monocistronic . This makes it possible for prokaryotes to have the information of several genes - one behind the other on the DNA - on a single mRNA transcript and to begin with the synthesis of the encoded proteins (translation) during the mRNA synthesis (transcription). Such a jointly transcribed area of functionally related genes on the DNA is called an operon .
Eukaryotic pre-mRNA processing
In eukaryotic cells, a mature mRNA is created by processing its precursor, the hnRNA (English heterogeneous nuclear RNA ) or pre-mRNA (English precursor messenger RNA , pre-mRNA). These process steps are still running from the nucleus - only then the mRNA through nuclear pores get into the cytoplasm where then to ribosomes , the protein synthesis takes place.
- At the 5 'end , it is synthesized in the transcription First, the RNA gets a 5'-cap structure (English cap "cap"). This cap consists of a modified form of guanosine , 7-methylguanosine (m 7 G). The cap structure protects the RNA from degradation by nucleases and allows the cap-binding complex , which u. a. is important for core export. Even after it has been transported into the cytosol , the 5′-cap plays a role in the recognition of the mRNA by the small ribosomal subunit and thus in the beginning of translation.
- At the 3 'end , synthesized last in the transcription, the RNA undergoes polyadenylation . In this process, a poly-A tail consisting of 30 to 200 adenine nucleotides is attached. This also protects the mRNA from enzymatic degradation. In addition, it facilitates both the core export and the translatability of the mRNA - also through interactions with the cap at the other end.
- By splicing (English splicing ), from the original transcript (hnRNA) certain RNA segments as introns removed, so no longer contribute to the coding information; the remaining sections are connected to one another and then represent the expressed genetic information as exons . This process takes place in the spliceosome , a complex of the hnRNA and the so-called snRNPs (English small nuclear ribonucleoproteins ) - consisting of the snRNAs U1, U2, U4, U5 and U6 and around 50 proteins - as well as other splicing factors. Through alternative splicing can order from the same hnRNA different mRNAs arise which translated can lead to different proteins.
Various regulatory processes in the cell also intervene at this point. About antisense RNA and RNA interference mRNA can be degraded and so the translation can be prevented. Furthermore, in some cases the so-called RNA edition takes place , whereby nucleotides in an mRNA are changed. An example of this is the apolipoprotein B mRNA, which is edited in some tissues and not in others. The edition creates a second stop codon upstream , which leads to a shorter protein (with a different function) during translation.
Translation
During translation , the coding sequence of nucleobases of the mRNA nucleotides is translated (translated) into the amino acid sequence of the polypeptide chain of a protein . The nucleotide sequence of an open reading frame is read in steps of three and each base triplet is assigned a specific amino acid using tRNA molecules and this is linked to the previous one via a peptide bond . This process takes place on the ribosomes in the cytoplasm and represents the actual protein biosynthesis . In eukaryotic cells, the ribosomes can be free or attach to the membrane of the endoplasmic reticulum .
Each ribosome that recognizes and binds to an mRNA then translates the coding nucleotide sequence of the mRNA into the corresponding amino acid sequence of a protein according to the genetic code . The amino acid molecules required for this are carried by tRNA molecules and taken from the cytoplasm of the cell. While a prokaryotic mRNA often contains several coding sections ( polygenic mRNA ), eukaryotic mRNAs are usually monocistronic and thus contain only one section with a coding sequence.
A ribosome only translates one mRNA at a time and builds the encoded polypeptide once according to its specifications. The ribosome then detaches from the mRNA. However, several ribosomes can attach to an mRNA at the same time and each synthesize a polypeptide chain. An mRNA can also be read ribosomally several times in succession. The number of protein molecules formed therefore depends on the number of translation processes carried out. The longer an mRNA molecule exists, the more often it can be used for this purpose.
Degradation
The mRNA is later enzymatically broken down by a ribonuclease (RNase) and broken down into its nucleotides, which can then be used again to build up new RNA molecules. With this decomposition , also called degradation, the life of an mRNA molecule ends. However, the duration of availability until degradation by nucleases in the cell can vary and is important for regulating protein biosynthesis. The degradation process often takes place in eukaryotic cells in specific structures, the so-called P-bodies in the cytoplasm. Since mRNA can also be temporarily stored here for a new translation instead of being broken down, additional modes of regulation are possible.
Cleaning and proof
RNA can be isolated by RNA purification , e.g. B. for RNA with a poly-A tail by using an oligo-dT column. Detection is carried out by Northern blot or, after RT-PCR, by qPCR or DNA sequencing .
mRNA vaccines
Since mRNA is rapidly broken down by enzymes in a cell, research into drugs based on RNA, such as vaccines , was initially hesitant . A vaccination in which the mRNA coding for a specific protein is administered can trigger an immune response when the mRNA is taken up in cells, the protein is ribosomally synthesized and this is then presented to the immune system as an antigen extracellularly. This method can be used for various purposes, for example in cancer therapy, as well as for influenza vaccines and for rabies vaccines .
Several projects are currently involved in pre-clinical or clinical studies with the development of mRNA vaccines to protect against the disease COVID-19 caused by the SARS-CoV-2 coronavirus .
literature
- James E. Darnell , Harvey Lodish, David Baltimore : Molecular Cell Biology . de Gruyter, Berlin et al. 1993, ISBN 3-11-011934-X (4th edition. Harvey Lodish: Molecular Cell Biology. Spectrum Akademischer Verlag, Heidelberg et al. 2001, ISBN 3-8274-1077-0 ).
- Benjamin Lewin: Molecular Biology of Genes . Spektrum Akademischer Verlag, Heidelberg et al. 1998, ISBN 3-8274-0234-4 .
- William S. Klug, Michael R. Cummings, Charlotte A. Spencer: Genetics . 8th, updated edition 2007, ISBN 978-3-8273-7247-5 .
- G. Haimovich, DA Medina et al. a .: Gene expression is circular: factors for mRNA degradation also foster mRNA synthesis. In: Cell. Volume 153, number 5, May 2013, pp. 1000-1011, doi : 10.1016 / j.cell.2013.05.012 . PMID 23706738 .
Individual evidence
- ↑ B. Weide, JP Carralot, A. Reese, B. Scheel, TK Eigenler, I. Hoerr, HG Rädenee, C. Garbe, S. Pascolo: Results of the first phase I / II clinical vaccination trial with direct injection of mRNA . In: Journal of immunotherapy. Volume 31, Number 2, 2008 Feb-Mar, pp. 180-188, doi : 10.1097 / CJI.0b013e31815ce501 , PMID 18481387 .
- ↑ B. Weide, S. Pascolo, B. Scheel, E. Derhovanessian, A. Pflugfelder, TK Eigenler, G. Pawelec, I. Hoerr, HG R Bäumenee, C. Garbe: Direct injection of protamine-protected mRNA: results of a phase 1/2 vaccination trial in metastatic melanoma patients. In: Journal of immunotherapy. Volume 32, Number 5, June 2009, pp. 498-507, doi : 10.1097 / CJI.0b013e3181a00068 , PMID 19609242 .
- ↑ M. Fotin-Mleczek, KM Duchardt, C. Lorenz, R. Pfeiffer, S. Ojkić-Zrna, J. Probst, KJ Kallen: Messenger RNA-based vaccines with dual activity induce balanced TLR-7 dependent adaptive immune responses and provide antitumor activity. In: Journal of immunotherapy. Volume 34, Number 1, January 2011, pp. 1-15, doi : 10.1097 / CJI.0b013e3181f7dbe8 , PMID 21150709 .
- ^ FB Scorza, N. Pardi: New Kids on the Block: RNA-Based Influenza Virus Vaccines. In: Vaccines. Volume 6, number 2, April 2018, p., Doi : 10.3390 / vaccines6020020 , PMID 29614788 , PMC 6027361 (free full text).
- ↑ N. Armbruster, E. Jasny, B. Petsch: Advances in RNA Vaccines for Preventive Indications: A Case Study of A Vaccine Against Rabies. In: Vaccines. Volume 7, number 4, September 2019, p., Doi : 10.3390 / vaccines7040132 , PMID 31569785 , PMC 6963972 (free full text).