Splicing: Function, Tasks, Role & Diseases

Splicing represents a crucial process during transcription in the nucleus of eukaryotes, in which mature mRNA emerges from pre-mRNA. In this process, introns that are still present in the pre-mRNA after transcription are removed, and the remaining exons are combined to form the final mRNA.

What is splicing?

The first step in gene expression is called transcription. In this process, RNA is synthesized, using DNA as its template. The central dogma of molecular biology is that the flow of genetic information is from the information carrier DNA to RNA to protein. The first step in gene expression is transcription. In this process, RNA is synthesized, using DNA as a template. DNA is the carrier of genetic information, which is stored there with the help of a code consisting of the four bases adenine, thymine, guanine and cytosine. During transcription, the RNA polymerase protein complex reads the base sequence of the DNA and produces the corresponding “pre-messenger RNA” (pre-mRNA for short). In this process, uracil is always inserted instead of thymine. Genes are composed of exons and introns. Exons are those parts of the genetic material that actually encode genetic information. Introns, on the other hand, represent non-coding sections within a gene. The genes stored on the DNA are thus interspersed with long segments that do not correspond to amino acids in the later protein and do not contribute to translation. A gene can have up to 60 introns, with lengths between 35 and 100,000 nucleotides. On average, these introns are ten times longer than exons. The pre-mRNA formed in the first step of transcription, also often referred to as immature mRNA, still contains both exons and introns. This is where the process of splicing begins. The introns must be removed from the pre-mRNA and the remaining exons must be linked together. Only then can the mature mRNA leave the nucleus and initiate translation. Splicing is mostly carried out with the help of the spliceosome. This is composed of five snRNPs (small nuclear ribonucleoprotein particles). Each of these snRNPs consists of an snRNA and proteins. Some other proteins that are not part of the snRNPs are also part of the spliceosome. Spliceosomes are divided into major and minor spliceosome. Major spliceosome processes more than 95% of all human introns, and minor spliceosome mainly handles ATAC introns. For explaining splicing, Richard John Roberts and Phillip A. Sharp were awarded the Nobel Prize in Medicine in 1993. For their research on alternative splicing and the catalytic action of RNA, Thomas R. Cech and Sidney Altman received the Nobel Prize in Chemistry in 1989.

Function and task

In the process of splicing, the spliceosome forms anew each time from its individual parts. In mammals, snRNP U1 first attaches to the 5′-splice site and initiates the formation of the rest of the spliceosome. The snRNP U2 binds to the branching site of the intron. Following this, the tri-snRNP also binds. The spliceosome catalyzes the splicing reaction by two successive transesterifications. In the first part of the reaction, an oxygen atom from the 2′-OH group of an adenosine from the “branch point sequence” (BPS) attacks a phosphorus atom of a phosphodiester bond in the 5′-splice site. This releases the 5′-exon and the intron circulates. The oxygen atom of the now free 3′-OH group of the 5′-exon now binds to the 3′-splice site, connecting the two exons and releasing the intron. The intron is thereby brought into a schligen-shaped conformation, called lariat, which is subsequently degraded. In contrast, spliceosomes play no role in autocatalytic splicing (self-splicing). Here, the introns are excluded from translation by the secondary structure of the RNA itself. Enzymatic splicing of tRNA (transfer RNA) occurs in eukaryotes and archeae, but not in bacteria. The process of splicing must occur with extreme precision exactly at the exon-intron boundary, since a deviation by only a single nucleotide would lead to the incorrect coding of amino acids and thus to the formation of completely different proteins. Splicing of a pre-mRNA can vary due to environmental influences or tissue type. This means that different proteins can be formed from the same DNA sequence and thus the same pre-mRNA.This process is called alternative splicing. A human cell contains about 20,000 genes, but is capable of forming several hundred thousand proteins due to alternative splicing. About 30% of all human genes exhibit alternative splicing. Splicing has played a major role in the course of evolution. Exons often encode single domains of proteins, which can be combined in various ways. This means that a large variety of proteins with entirely different functions can be generated from just a few exons. This process is called exon-shuffling.

Diseases and disorders

Some inherited diseases can arise in close association with splicing. Mutations in the noncoding introns do not normally lead to defects in protein formation. However, if a mutation occurs in a part of an intron that is important for the regulation of splicing, this can lead to defective splicing of the pre-mRNA. The resulting mature mRNA then encodes defective or, in the worst case, deleterious proteins. This is the case, for example, in some types of beta-thalassemia, a hereditary anemia. Other representatives of diseases that arise in this way include Ehlers-Danlos syndrome (EDS) type II and spinal muscular atrophy.