Épissage

At the higher organizations (Eucaryote S), the Gène S which code Protéine S consist of a continuation of Exon S and alternate Intron S. Ex: Exon1-Intron1-Exon2-Intron2-Exon3.

At the time of the transcription pre-ARNm is synthesized, this one will be spliced to give place to mature ARN m known as . The épissage is a process which occurs in the core of the cell and which consists of the excision of will introns and in the binding of exons. Mature ARNm , made up of only let us exons, is then exported towards the Cytoplasme to be translated into protein.

The épissage is ensured by a whole of complexes ribonucleoproteic called Spliceosome collectively (épissage = splicing in English). Each complex, called snRNP for small nuclear RiboNucleoProtein , contains a ARN and several proteins. There also exists will only introns car-épissables, i.e. capable of exciser, in the Mitochondrie S, the Plaste S and some Bactérie S.

By analogy with the operation of the Ribosome, one thinks that in the spliceosome, in fact the ARN is catalytic and thus that the spliceosome is a Ribozyme. The exact catalytic mechanism of the spliceosome is still unknown

Structure of will introns

The junctions intron/exon contain characteristic nucleotidic sequences which are preserved. These sequences are recognized by the spliceosome. The intron contains in more one sequence internal, known as limps of connection . This limps of connection comprises a Adénosine which plays a central role in the process of épissage.

The épissage itself is carried out in two times, one first of all has an attack Nucléophile of the 2 ' - OH of the Ribose of the adenosine of limps of connection on phosphate of the junction exon-intron into 5 '. After this cut, 3 ' - OH released on the level of the exon upstream attacks phosphate of the junction intron-exon downstream. The products of this reaction are on the one hand both exons bound correctly and on the other hand, the intron cyclized on the level of the adenosine of limps of connection. Because of its particular form, this form of the intron is called lasso (in: lariat ). The lasso is finally opened by an enzyme of disconnection in order to be able to be recycled

Mechanism of épissage

snRNP

The snRNP or small nuclear ribonucleoproteins are components of the Spliceosome present in the core of the cells Eucaryotes. The canonical process of épissage calls upon five snRNP, called U1, U2, U4, U5 and U6. Each one of these snRNP is composed of a ARN, is called SnARN, and several proteins. Among associated proteins, some are common to all the snRNP and others are specific of each one of them. The common proteins are called proteins Sm , they are seven and join to form a ring heptameric surrounding a segment of the snARN.

The specific proteins are of variable number, there are for example three for U1 of them: U1A, U1C and U1-70K.

Process of épissage

The process of épissage is a dynamic mechanism which follows a precise succession of events during which different the snRNP are assembled and are disassembled spliceosome. The sequence is the following one:

U1 joins junction 5 ' intron.
U2 joins limps of connection.
U4 associated with U6 brings closer U1 and U2, thus carrying out a bridge between junction 5 ' of the intron and limps it of connection.
U5 joins in its turn and brings closer edges 3 ' and 5 ' of let us exons to suture.
U4 and U1 leaves the complex.
2 ' - OH of has limps of connection cuts junction 5 ' of the intron.
3 ' - OH of nucleotide in 3 ' of the exon upstream cuts the other junction.
spliced ARNm and the intron in " lasso" are released.

Association of the components of the spliceosome

The majority of the interactions between ARNm and the snRNP pass by basic pairings between portions of nucleotidic sequences complementary. Thus for example the snARN U1 contains a sequence complementary to the sequence consensus found to the junction exon-intron into 5 ', which enables him to bind to this area of the pre-ARN messengers. There exist also pairings ARN-ARN between different the snARN.

Introns autocatalytic

In certain protozoa (unicellular organizations eucaryotes), certain intronic ARN do not need spliceosomes, they are enough to catalyze the reactions of épissage (" coil-splicing") without external protein contribution. This relatively rare phenomenon concerns will introns them formants a Ribozyme. One touve two kinds of will introns autocatalytic: Groups 1 and 2.

Alternate Épissage

At Eucaryotes only, it is a stage necessary to return a ARN viable messenger to the translation. This stage consists with the délétion of will introns or of let us exons which will not be necessary to the coding of protein, with the addition of a methylated cap into 5 ' and of a polyadénylé passing (also called tail poly A) into 3 '. These last elements will be necessary to the displacement of the ARN messenger of the core towards the cytoplasm, where it will be translated by the ribozymes, like with its stability, since it will not be contestable any more by ARNs nucléosidases.

Indeed the spliceosome recognizes signals of épissage, as for a radio signal, these signals of épissage are more or less strong, which implies that the spliceosome recognizes them more or less well.

These signals are simply specific sequences of nucleotides.

The weak signals are called “signals of épissage alternate”, they will make it possible pre-ARNm to be spliced in several ARNm mature . By opposition the strong signals are called “constitutive signals”.

Thus a gene can code several Protéine S. The alternate épissage plays a very important part in the development of the cells, the organization of fabrics and even in the development of an individual. (ex: the gene Slx for the differentiation of the sex at the Drosophila). Today, it is allowed that nearly 60% of genes at the human being undergo the alternate épissage, it is the phenomenon which explains the surprise of the researchers who during the analysis of the human Génome calculated that the latter would contain between 25 and 30.000 genes. And the dogma “1 gene for 1 protein” is not any more.

In certain extreme cases, the alternate épissage makes it possible only one gene to code more proteins than all the others joined together. It is the case for Dscam at the Drosophile which can code to 96.000 different proteins (in theory).

The alternate épissage thus makes it possible a gene to code several ARNm different. It can be done in several ways.

the lasso always does not include/understand a intron only. For a gene containing a great number of let us exons, the formed lasso can include/understand 2 will introns and 1 exon instead of only one intron. One will have then exons not-represented in mature ARNm, and thus in protein.
One can find several promoters within the same gene. So ARNm mature do not contain that exons them downstream from the promoter, which involves the translation of several different proteins according to the promoter used at the time of the épissage.
One can find sequences polyA (sequence of residues adénine which protects ARNm from the exonucléases) inside ARNm in addition to the sequence polyA which one finds at end 3 ' of ARNm. According to the sequence polyA which will be cut before translation, there will be thus different translated proteins.

The alternate épissage is an important process of regulation of the form of genes. Two consequences of the alternate épissage contribute to it: First is known under initial NMD (Not-Judicious RNA Mediated Decay): at the time of the alternate épissage, the addition or withdrawal of exon, or even modification of the length of such or such exon (compared to ARNm " canonique") can involve a shift of the framework of reading (the nucleotides are read three by three by ribosome during the translation), and of this fact of causing the premature appearance of a codon STOP, what is called a change nonsense. ARNm thus formed are recognized by a whole of proteins and ranges before the translation can take place. This process is not only one mechanism of " control qualité" but also allows, by increasing the rate of the épissages alternate producers of code STOP premature, to decrease the number of translated proteins. The second consequence is quite simply the fact that a change in the sequence of ARNm is reflected on that of protein and thus potentially on its physicochemical capacities, and makes it possible to control its function. For example: protein CD45 plays a part in the activation of the lymphocytes T at the time of an immunizing response to an infection. When the lymphocytes T are at rest and thus potentially activables, the longest shape of protein is expressed (it comprises the totality of the nine exons). When the lymphocytes T have just been activated, ARNm of CD45 are spliced alternatively and let us exons them 4,5,6 are excisés. The form of CD45 thus produced is thus shorter and cannot play its part of activator. This prevents the lymphocytes T from receiving a signal of activation too a long time and thus to start a disproportionate immunizing response.

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