Name: rna splicing and the spliceosome
Uploaded: 2020-11-23T11:47:36.000Z
Duration: 1 min 32 s
Channel: JoVE

When pre-mRNA is transcribed from DNA, it contains exons, protein-coding sequences, and introns, non-coding regions. RNA splicing removes the introns and attaches the exons together.
This is catalyzed at the spliceosome, a large assembly of small nuclear ribonucleoproteins, or snRNPs &#8212; complexes made up of small nuclear RNAs and proteins.
The spliceosome is composed of 5 snRNPs, U1, U2, U4, U5 and U6, and several other proteins.
First, U1 snRNP binds the 5&#39; splice site while the branch point sequence containing A is recognized by other proteins and later replaced by U2 snRNP.&#160;
Next, a complex of U4/U6 and U5 snRNPs binds to the 5&#39; splice junction and assists in bringing the 5&#8217; end to the branch point A. A transesterification reaction between the branch point A and the 5&#39; splice site creates a loop called a lariat.
Following this, another transesterification reaction occurs between the 3&#39; end of the upstream exon and the 5&#39; end of the next exon. This cleaves off the lariat containing the intron, leaving the exons attached to each other.&#160;

rna splicing

RNA Splicing

Splicing is the process by which eukaryotic RNA is edited before its translation into protein. The RNA strand transcribed from eukaryotic DNA is called the primary transcript. The primary transcripts that become mRNAs are called precursor messenger RNAs (pre-mRNAs). Eukaryotic pre-mRNA contains alternating sequences of exons and introns. Exons are nucleotide sequences that code for proteins, whereas introns are the non-coding regions. In RNA splicing, introns are removed and exons are bonded together.
Splicing is Mediated by the Spliceosome
Splicing occurs in the nucleosome and is mediated by a complex of proteins and small RNAs called small nuclear ribonucleoproteins (snRNPs). snRNPs, together with other proteins, form the spliceosome, which recognizes specific nucleotide sequences at the ends of the exon and intron. First, it binds to a GU-containing sequence at the 5&#39; end of the intron and to a branch point sequence containing an A towards the 3&#39; end of the intron. In a number of carefully-orchestrated steps, other snRNPs then bring the branch point close to the 5&#39; splice site. Subsequently, a chemical reaction cleaves the 5&#39; end of the intron from its upstream exon and attaches it to the branch point, forming a loop called a lariat. To release the lariat, the AG-containing sequence of the intron near the 5&#39; end of the downstream exon reacts with the 3&#39; end of the upstream exon. This reaction patches the two exons together, concluding the splicing process.
Splicing Allows the Expression of Several Proteins from a Single Gene
Typically, exons are joined together in the order in which they appear in a gene. However, during alternative splicing, different combinations of exons in pre-mRNA are combined to form mature mRNA. This produces several different proteins from a single pre-mRNA transcript.
Different patterns of alternative splicing include exon skipping, alternative 5&#39; or 3&#39; splice sites, and intron retention. These patterns are guided by the length of exons or introns and the strength of the splicing signal at the splice sites. Because of this, exons that are shorter than other exons may be overlooked by the spliceosome and omitted from the mature mRNA. In contrast, introns that are significantly shorter than other introns may evade removal by the spliceosome and are retained in the mature mRNA. As a result, alternative splicing generates variants of mature mRNA that were copied from the same stretch of DNA. The RNA sequence variants produce different proteins with additional or fewer amino acids, shifts in the reading frame, or a premature stop codon. These protein isoforms have different biological properties, including function, cellular localization, and interaction with other proteins, thereby playing a vital role in tissue- and environment-specific gene expression.
Abnormal Splicing Can Cause Diseases
Errors in splicing can produce aberrant protein isoforms, which may contribute to diseases, including cancer. For instance, alternative splicing of the BCL2L1 gene generates a long and short protein isoform&#8212;BCL-XL and BCL-XS, respectively&#8212; through the use of alternative 5&#39; splice sites. The longer BCL-XL isoform promotes cell survival and is highly expressed in several types of cancers (e.g., blood, breast, and liver cancers). Expression of the short BCL-XS isoform that promotes cell death is suppressed in cancer.

Splicing is the process by which eukaryotic RNA is edited before its translation into protein. The RNA strand transcribed from eukaryotic DNA is called ...

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JoVE Core

Molecular Biology

Unit 1

Transcription: DNA to RNA

KEY TERMS AND CONCEPTS

rna splicing and the spliceosome

RNA splicing is a step in gene expression that helps turn pre-mRNA into mature mRNA. During this process, the cell removes introns and joins the remaining exons together. The finished mRNA can then carry the correct coding sequence forward.

The spliceosome carries out this work. It is a large complex made of proteins and RNA that recognizes the splice sites on pre-mRNA. After binding, it cuts out the intron and links the exons in order.

A key intermediate in splicing is the lariat. The intron folds into a looped shape as it is removed. This structure helps the spliceosome complete the cut-and-join process before the mature mRNA is released.

RNA Splicing and the Spliceosome

what is gene expression?

Gene Expression: From DNA to Proteins

Gene expression is the process that uses DNA instructions to make proteins. Proteins help cells do many of their jobs, so gene expression is a key part of how cells work. The process starts with transcription, when a gene’s DNA sequence is copied into RNA. In eukaryotic cells, this RNA is then processed by splicing. Splicing removes noncoding regions and joins the coding parts of the message. After transcription and splicing, the RNA is ready for translation. During translation, the cell...

rna structure

RNA Folding and Base Pairing

RNA structure depends on how its nucleotides pair and fold. RNA is made of ribonucleotides, and their sequence helps determine the molecule’s shape. That shape is important because it affects how RNA works in the cell. Unlike DNA, RNA is usually single-stranded. Even so, it can fold back on itself and form paired regions. These folds create the structure students see when comparing RNA to DNA. The transcript also highlights the role of ribonucleotides in building RNA. Their order along the...

rna stability

How Cells Control RNA Lifetime

RNA stability describes how long an RNA molecule stays intact in a cell. Cells do not let every RNA last the same amount of time. Instead, they control RNA lifetime to help regulate gene expression. Some RNAs are made to be short-lived, while others remain stable longer. This difference matters because RNA helps carry genetic information from DNA and supports protein production. When RNA breaks down faster or slower, the cell can change how much of a protein is made. The transcript also...

bacterial rna polymerase

How Sigma Factors Guide Bacterial Transcription

Bacterial transcription starts with a single RNA polymerase, the enzyme that makes RNA from DNA. Unlike eukaryotes, bacteria use one RNA polymerase to transcribe all genes. Its subunits have different jobs. The multisubunit structure helps the enzyme keep catalytic activity, assemble correctly, interact with DNA and RNA, and regulate its own activity. In most genes, the transcription start site is a single base upstream of the coding sequence. RNA polymerase works efficiently, but it does not...

types of rna

RNA in Protein Synthesis

RNA helps cells use genetic information to make proteins. This topic focuses on the main RNA types involved in that process: messenger RNA, ribosomal RNA, and transfer RNA. Messenger RNA, or mRNA, carries the genetic message from DNA to the ribosomes. Ribosomal RNA, or rRNA, is part of the ribosome, the cell structure where proteins are made. Transfer RNA, or tRNA, brings the correct amino acids to the ribosome so the protein chain can grow. These RNA types work together during protein...

transcription

Transcription and RNA Formation

Transcription is the process that makes RNA from DNA. It begins when RNA polymerase, the enzyme that copies genetic information, binds to a gene on the DNA strand. The DNA opens so the cell can read the template strand. As RNA polymerase moves along the template strand, it builds a complementary RNA copy. The RNA strand carries the message from DNA in a form the cell can use later. This step is a key part of gene expression. The process ends when the RNA molecule is released and the DNA...

transcription factors

How Transcription Factors Control Gene Activity

Transcription factors control gene activity by helping turn genes on or off. They are proteins that bind to specific DNA sequences and influence whether a gene is copied into RNA. Some transcription factors act as activators. Others act as repressors. Activators help increase transcription, while repressors help reduce it. These proteins work as part of gene regulation, which is the control of when and how strongly a gene is expressed. Transcription factors help cells respond to signals and...

rna polymerase ii accessory proteins

Transcription Regulators and Chromatin Access

Transcription regulators can help control gene expression by acting in different ways. Some proteins contact RNA polymerase directly. Others work through adaptors, mediators, histone-modifying proteins, and nucleosome remodelers. Direct activation is seen in bacteria and in some eukaryotic genes. In these cases, upstream activation sequences sit near the promoter, and activator proteins interact directly with the transcription machinery. In prokaryotes, the catabolite activator protein, or...

eukaryotic rna polymerases

RNA Polymerase II and mRNA Termination

RNA Polymerase II makes mRNA and then helps end transcription in eukaryotic cells. It is the eukaryotic RNA polymerase that is most similar to bacterial RNA polymerase in structure and folding, even though its action works differently. Eukaryotic RNA polymerases need specific transcription factors to begin and guide RNA synthesis on the template DNA strand. The TATA-binding protein is one factor shared by all three eukaryotic RNA polymerases. These proteins stay attached to the polymerase and...

transcription elongation factors

RNA Polymerase Pausing in Elongation

RNA polymerase pausing helps control transcription elongation as DNA is copied into RNA. The elongation complex can change as it moves along a gene because the DNA sequence is not uniform. During this stage, the polymerase may stop and start several times. In bacteria, these pauses matter because transcription is linked to translation, the process that makes protein from mRNA. In eukaryotes, transcription is linked to mRNA processing instead. Pausing near exon-intron junctions can improve the...

pre-mrna processing

Pre-mRNA Splicing and RNA Protection

Pre-mRNA processing prepares a new RNA transcript for use in the cell. Before the RNA can serve as mature messenger RNA, it must be modified after transcription. These changes help protect the molecule and prepare it for later steps in gene expression. One important change is the addition of a 5' cap at the front end of the RNA. Another is the addition of a poly-A tail at the back end. The RNA is also spliced, which means noncoding regions are removed and the remaining coding sections are...

RNA splicing is a step in gene expression that helps turn pre-mRNA into mature mRNA. During this process, the cell removes introns and joins the remaining exons together. The finished mRNA can then carry the correct coding sequence forward. The spliceosome carries out this work. It is a large complex made of proteins and RNA that recognizes the splice sites on pre-mRNA. After binding, it cuts out the intron and links the exons in order. A key intermediate in splicing is the lariat. The intron...

chromatin structure regulates pre-mrna processing

chromatin-structure-regulates-pre-mrna-processing

How Chromatin Shapes mRNA Splicing

Chromatin structure shapes pre-mRNA splicing in eukaryotic cells. As a gene is being transcribed, the new RNA is processed at the same time before it can leave the nucleus and make a functional protein. For a long time, transcription and pre-mRNA processing were thought to happen one after the other. The transcript shows that they are actually simultaneous and tightly controlled inside the cell. Nucleosome placement and histone changes on a gene can affect how fast RNA polymerase works and how...

nuclear export of mrna

mRNA Quality Control Before Export

mRNA quality control begins before an mRNA leaves the nucleus. Eukaryotic cells use mRNA surveillance to check each transcript for structural and functional problems. If an abnormal mRNA is not removed, it can be translated into a protein that does not work properly. One major problem is a premature stop codon. This kind of mutation places a Stop Codon too early in the reading frame. About 30% of inherited genetic disorders in humans result from these mutations. The faulty mRNAs are broken...

ribosomal rna synthesis

How Ribosomes Are Assembled in Cells

Ribosomes are assembled through a complex process that takes place in several parts of a eukaryotic cell. More than 200 assembly factors help build the final structure. The work happens in the nucleolus, the nucleoplasm, and the cytoplasm. Ribosome biogenesis starts when two different RNA polymerases make 5S and 45S pre-rRNAs. These are primary RNA transcripts that still need to be processed. Ribosomal proteins and assembly factors, which are imported from the cytoplasm, bind to these...

transfer rna synthesis

tRNA Modified Bases and Function

Transfer RNA (tRNA) contains many modified bases that help it work during protein synthesis. In some tRNAs, these chemical changes make up nearly 20% of the total bases. They also help protect tRNA from breakdown by RNases, which are enzymes that cut RNA. These modifications are added after transcription, meaning after the tRNA is made from DNA. Each change is carried out by a specific enzyme with its own base and site preference. Methylation is the most common modification, and at least nine...

the nucleolus

Nucleolus Structure and Ribosome Assembly

The nucleolus is the most prominent substructure inside the nucleus. It was once thought to be an isolated organelle that forms fibrils and granules. Later, in 1931, Heitz described its link with chromosomes and showed that its size and appearance change during the cell cycle. Heitz also found constricted regions on different chromosomes that cluster at specific stages of the cell cycle. These regions are now called nucleolar organizer regions, or NORs. NORs contain the genes that encode...

additional subnuclear structures

Nuclear Bodies in Gene Regulation

The eukaryotic nucleus contains membrane-less nuclear bodies that help organize genetic activity. These subnuclear organelles include nucleoli, Cajal bodies, speckles, and paraspeckles. By separating work into small regions, they create distinct sites for specific reactions inside the nucleus. Their lack of a membrane also makes it easier for components to move between the nuclear bodies and the surrounding nucleoplasm. Cajal bodies are spherical structures inside the nucleus. They contain...

alternative rna splicing

How One Gene Makes Many mRNAs

Alternative RNA splicing lets one pre-mRNA produce different mature mRNAs. It does this by regulating how exons and introns are removed or kept. In constitutive splicing, one gene usually makes one mRNA. In alternative splicing, a single gene can help produce multiple proteins. That is why this process is important for protein diversity. There are five main types of alternative RNA splicing. In exon skipping, some exons, along with all introns, are removed. This is the most common type seen in...

bacterial transcription

Bacterial RNA Polymerase Steps

Bacterial RNA polymerase carries out RNA synthesis from DNA. In bacteria, there is no membrane-bound nucleus. As a result, transcription and translation can happen at the same time on the same DNA template. Bacterial transcription has three main stages: initiation, elongation, and termination. These stages are guided by specific DNA sequences. The promoter regions lie 10 and 35 base pairs upstream of the gene and help RNAP begin transcription. Transcription starts at the DNA base pair labeled...

transcription initiation

RNA Polymerase Recruitment in Transcription

Transcription in eukaryotes begins when RNA polymerase is recruited to the promoter on DNA. In prokaryotes, RNA polymerase can bind the template DNA and start transcription on its own. In eukaryotes, extra proteins called transcription factors must bind first. These transcription factors help bring the correct RNA polymerase to the DNA so transcription can start. Promoters and enhancers, along with their accessory proteins, help control this process tightly. The TATA box is the best-studied...

key terms and concepts

Learn how RNA splicing removes introns, joins exons, and uses the spliceosome and lariat to produce mature mRNA in gene expression.

RNA Splicing and the Spliceosome | JoVE K12

molecular biology

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RNA Splicing and the Spliceosome

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