microRNA

MicroRNA (Greek micros, small ' ), abbreviated miRNA or miR are short, highly conserved, non-coding RNAs that play an important role in the complex network of gene regulation, particularly in gene silencing. MicroRNAs regulate gene expression highly specific to the post-transcriptional level. miRNAs generally have a size of 21 to 23 nucleotides (nt) on.

• 5.1 Important microRNAs in human cell

Mechanism of gene regulation

Gene regulation occurs in animals by binding of microRNAs to the 3 ' untranslated region (3'- UTR) of the mRNA of target genes can be inhibited depending on the complementarity of the binding sequence and the proteins involved either the translation or degraded by cutting. Partial complementarity leads to translational inhibition, whereas perfect base pairing leads to the degradation of the target mRNA. During a long time it was considered that both of these mechanisms dominating the translational inhibition, recent studies revealed that the degradation of the target mRNA relative terms is responsible for a large proportion of the inhibition of protein production.

History

MiRNAs were first described in 1993, the name of microRNA was coined only in 2001, however. In the nematode Caenorhabditis elegans, the gene lin - 4 coded surprisingly not a protein but for two small RNA molecules having a length of about 60 nt and about 20 nt. The longer the molecule is based on current knowledge, the pre - microRNA dar. In the molecule of about 20 nucleotides is the mature microRNA. The smaller RNA lin -4 regulates the gene lin -14 by complementarily binds through base pairing to the 3'- UTR of the lin -14 mRNA and reduces the translation of the mRNA. With the discovery of another miRNA ( let-7 ) showed that it is probably not a rare phenomenon in C. elegans. Let-7 regulates the gene lin -41. Since it was evolutionarily conserved genes in let-7 and lin -41, it became clear that the mechanism of microRNA regulation could be applied to other multi-cellular organisms. In recent years the knowledge about microRNAs have grown steadily. The miRBase database shows an increase of over 4000 sequences in the last two years. The PubMed database is experiencing a significant increase in publications on microRNAs. The biological functions of most miRNAs are still unknown. After computer-based predictions about 20-30 % of the genes might be regulated by microRNAs in the human genome. It can be assumed that several hundred to a few thousand different microRNAs are encoded.

Biogenesis

Transcription and activity is described in terms of mammals. With variations, such as the participating protein components of the active mechanism in the different species is comparable. The genes for the miRNAs are located in the genome, they are transcribed by an RNA polymerase II or III. The resulting primary transcript having a length of 500 to 3000 nucleotides and has the conventional poly -A tail at the 3 ' end as well as a 7- methylguanosine cap at the 5' end. The primary transcript is called primary microRNA (pri - miRNA) and layered together to create a loop. The RNase III ( Drosha ) and the dsRNA - binding protein DGCR8 ( Pasha corresponds in Drosophila melanogaster) form a microprocessor complex in the nucleus of a cell through the pri-miRNA into an approximately 70-80 nucleotides major precursor microRNA ( pre-miRNA ) is processed. The pre-miRNA, thus building a characteristic hairpin structure ( hairpin ). The pre-miRNA is actively exported by Exportin -5 in the presence of Ran-GTP as a cofactor of the nuclear pores of the nuclear membrane into the cytoplasm. In the cytoplasm, the pre - miRNAs are cleaved by the RNase III enzyme Dicer into 17-24 nt ds- miRNAs. Dicer interacts with the dsRNA - binding protein TRBP (RDE -4 in C. elegans and Drosophila melanogaster Loquacious in ), making the miRNA duplex is unwound and single-stranded. Here, starting at the end with the lower thermodynamic stability. Depending forms of miRNA strand with the 5 'terminus at its end the mature miRNA, which is also called guide RNA. The opposite strand is marked in annotation with an asterisk. This miRNA * may possibly have a regulatory effect in a few cases. The mature miRNA is incorporated into a ribonucleoprotein ( miRNP ), which has a great resemblance to the RISC complex of the RNAi pathway. This miRISC complex the activity of target genes by two methods can be down-regulated. This depends on the degree of complementarity between the miRNA and the mRNA of the target gene, as well as RNA-binding proteins of the Argonaute family. In partial agreement of the binding sequence, the translation is inhibited by binding. At high complementarity, the target mRNA is cut. The mechanisms of action of microRNAs and the so-called small interfering RNAs (siRNAs ) have clear parallels. siRNAs are processed by the RNase III Dicer from long double-stranded RNAs ( dsRNAs ) and nt long single-stranded RNAs included as 21-28 in the siRISC complex. This complex mediates mRNA degradation.

RNA interference

A breakthrough in science is the discovery that artificially induced double-stranded RNA in C. elegans leads to an efficient and specific gene knockdown. This mechanism is known as RNA interference (RNAi). For the discovery of the mechanism of RNA interference, the two U.S. scientists Andrew Z. Fire and Craig C. Mello received in 2006 the Nobel Prize in Physiology or Medicine.

Expression

MicroRNAs have very different expression patterns and are differentially regulated in the development and physiological processes. The expression is usually organized tissue-specific manner along with nearby genes. MicroRNAs may be located within exons or introns protein-coding genes or in non-coding regions. Some microRNAs have their own promoters. A few microRNAs are in a cluster and are transcribed together.

Current Research

The study of microRNA is a new and very topical issue in molecular and cell biology. Working with miRNA proves more difficult than with mRNAs because they are degraded rapidly due to their small size and ubiquitous occurrence of degradation enzymes. The quantification of miRNA therefore requires independent work under cooling and specially prepared, RNase - free equipment. Conventional manner, is carried out the investigation of miRNA expression levels by rewriting the RNA synthetic DNA (cDNA) and subsequent quantification by means of quantitative PCR. Recently, it is also possible to measure miRNA expression in specific microarray platforms, a method that is growing in popularity. Although this allows the simultaneous inventory of hundreds of miRNAs, but the post-processing of the data, as is conventional microarrays, complicated and often connected to great variation. However, this can often be compensated for by the subsequent analysis of the data sets, in the best case additionally comprise information about the regulated mRNAs.

In recent years, it was found that the miRNAs as important regulators of Genübersetzung ( Translation) after Genüberschreibung ( transcription) act. This is done by specific attachment to mRNA molecules, thus hampering their translation into proteins, is completely prevented or relieved.

In mammalian cells so far have over 800 different miRNAs are detected; in humans are currently over 1,800 different miRNA species are known in collections, so-called libraries are present (11/ 2013 ) The comparison of invertebrate and vertebrate cells shows that the structure of some of these molecules is highly conserved, suggesting a suggests important common evolutionary function in very different species.

Experimental and informatics studies strongly suggest that each miRNA can regulate some mRNA molecules, and that 20-30 % of all human genes are also controlled by miRNAs.

The type and number produced in the nucleus miRNA molecules often show a close correlation with the level of development of the cell ( cell division, differentiation into specific cell types, apoptosis (programmed cell death in errors) ). The miRNAs thereby cooperate in a regulatory network of transcription factors ( TFn ). Thus, current studies show the critical role of miRNAs in early developmental processes of animals, such as neurogenesis, myogenesis (muscle formation), cardiogenesis ( heart formation, ) and hematopoiesis ( blood formation). Although probably arisen independently, miRNAs play an important role in plants.

Furthermore, it was suggested that miRNAs are very important for the suppression of cellular transformation such as tumor formation, since an incorrect formation of miRNA molecules into cells enhances these undesirable processes. Deregulation of miRNAs has been observed in various types of tumors and tumor-specific characteristics seem to have the influence of the well-known as a tumor suppressor p53 protein on the maturation of different miRNAs that inhibit cell growth, this conclusion suggests likewise.

Recent research shows that certain miRNAs for the maintenance of pluripotency and self-renewal of embryonic stem cells are important. microRNAs could therefore represent useful molecular biological tools for the manipulation of stem cells in the future.

For research purposes, cellular microRNA molecules can be inhibited by means of complementary Antagomirs.

Recent results suggest that miRNA is incorporated by nutrition and influences processes in the body.

The base sequences of the different miRNAs are identical or almost identical in mammals. These matches are (eg miRBase ) verifiable on the Internet via database queries. At the same time there is an organ specificity in all species, so that identical miRNA types take over their duties as part of the modulation of protein synthesis in the same organs of different mammals.

2013 has provided 190,000 euros for a current research project of the German Cancer Research Center Heidelberg, the German Cancer Aid. The scientists want to find out the effect of microRNAs in cancer cells and prevent the spread of malignant cells in the body. The researchers assume that " specific microRNAs, cancer cells can reprogram so that they spread in the body ."

Important microRNAs in human cell

The microRNA -335 ( 5p ) whose Transkriptvorlage is located in the gene sequence for MEST, has been recognized as an important regulator in the development of malignant tumors, but also recognized as carcinogenic Oncomir.

The microRNA -145 decreased the expression of Oct-4, a stem cell- type gene. Therefore, your failure promotes the development of cancer stem cells.

The microRNAs miR -302 family are typical of human embryonic stem cells and are regulated by the pluripotency genes Oct-4 and Sox -2.