# mRNA Processing in Eukaryotes: Steps and Mechanisms

## Introduction to mRNA Processing

### What is mRNA Processing?

In eukaryotic cells, the primary transcript produced by RNA polymerase II—termed pre-mRNA—is not directly translated into protein. Instead, it undergoes a series of covalent modifications collectively known as mRNA processing. These modifications convert the nascent pre-mRNA into a mature messenger RNA (mRNA) that is competent for nuclear export, translation, and ultimately, protein synthesis. The three canonical processing events are 5' capping, splicing, and 3' polyadenylation. In addition, RNA editing and base modifications can further alter the final mRNA sequence.

This is a fundamental distinction from prokaryotes, where [transcription and translation](/knowledge/molecular-biology/transcription-translation) are coupled in the cytoplasm, and mRNA is typically used immediately without extensive modification. In prokaryotes, polycistronic mRNAs can encode multiple proteins, and there is no nuclear compartment to separate transcription from translation. Eukaryotes, by contrast, transcribe mRNA in the nucleus, process it extensively, and only then export the mature mRNA to the cytoplasm for translation. This spatial and temporal separation provides an additional layer of regulatory control over gene expression.

### Why Eukaryotes Need mRNA Processing

The need for mRNA processing arises from the split nature of eukaryotic genes. Most eukaryotic genes contain coding sequences (exons) interrupted by non-coding sequences (introns). If introns were retained in the final mRNA, they would disrupt the open reading frame and produce non-functional or truncated proteins. Splicing removes these introns and joins exons in the correct order.

The 5' cap and 3' poly(A) tail serve multiple protective and functional roles. They protect the mRNA from 5'→3' and 3'→5' exonucleases, respectively, and they are recognized by translation initiation factors and RNA-binding proteins that control mRNA localization, stability, and translation efficiency. Without these modifications, the mRNA would be rapidly degraded, and translation would be inefficient. The processing events are also tightly coupled to transcription itself, with the C-terminal domain of RNA polymerase II acting as a platform to recruit processing factors. This coupling ensures that processing is efficient and accurate, and it allows the cell to coordinate the rate of transcription with the rate of mRNA maturation.

## The 5' Cap Addition

### Mechanism of Capping

The 5' cap is a modified guanosine nucleotide linked to the first transcribed nucleotide of the pre-mRNA via a 5'→5' triphosphate bridge. This unusual linkage distinguishes the cap from all other internal nucleotides, which are linked 3'→5'. The cap structure is 7-methylguanosine (m⁷G), and its addition occurs co-transcriptionally, almost as soon as the nascent RNA emerges from the RNA polymerase II exit channel—typically when the transcript is only 20–30 nucleotides long.

Capping proceeds through three enzymatic steps:

1. **RNA 5'-triphosphatase** removes one phosphate from the 5' end of the nascent transcript, converting the 5' triphosphate (pppN) to a diphosphate (ppN).
2. **Guanylyltransferase** (also called capping enzyme) transfers a GMP moiety from GTP to the diphosphate end, forming the 5'→5' triphosphate linkage (GpppN).
3. **Guanine-N7-methyltransferase** adds a methyl group to the N7 position of the newly added guanine, producing the mature cap structure (m⁷GpppN).

In metazoans, these three activities reside in a single bifunctional polypeptide, while in yeast, they are separate proteins. The capping enzymes are specifically recruited to RNA polymerase II via its C-terminal domain (CTD), which is phosphorylated on serine 5 during [transcription initiation](/knowledge/molecular-biology/transcription-initiation). This ensures that capping is restricted to RNA polymerase II transcripts and does not occur on rRNA or tRNA, which are transcribed by RNA polymerases I and III.

For many mRNAs, a second layer of methylation occurs on the ribose of the first and second nucleotides (2'-O-methylation), producing cap1 and cap2 structures. These additional methylations are added by cap-specific methyltransferases and are recognized by the innate immune system as "self" RNA, a distinction that is relevant for [mRNA-based vaccines](/knowledge/molecular-biology/difference-between-mrna-and-non-mrna-vaccine), where the synthetic cap structure must mimic the natural cap to avoid triggering an immune response.

### Functions of the 5' Cap

The 5' cap has several critical functions:

- **Protection from 5'→3' exonucleases:** The cap structure is not recognized by the major 5'→3' exonuclease Xrn1, which would otherwise rapidly degrade the mRNA. Decapping is a regulated step in mRNA turnover, and the [mRNA stability](/knowledge/molecular-biology/mrna-stability) of a transcript is in large part determined by how long its cap remains intact.
- **Translation initiation:** The cap is bound by the eukaryotic translation initiation factor 4E (eIF4E), which, together with eIF4G and eIF4A, forms the eIF4F complex. This complex recruits the 40S ribosomal subunit to the 5' end of the mRNA, a step that is rate-limiting for translation. The [translatability of mRNA sequences](/knowledge/molecular-biology/translatability-of-mrna-sequences) is therefore directly influenced by cap recognition.
- **Splicing enhancement:** The cap-binding complex (CBC), which binds the cap in the nucleus, promotes splicing of the first intron and facilitates mRNA export.
- **Nuclear export:** The CBC is recognized by the nuclear export machinery, helping to direct the mature mRNA out of the nucleus.

## Splicing: Removal of Introns

### The [Spliceosome Machinery](/knowledge/molecular-biology/spliceosome-machinery)

Splicing is the process by which introns are removed and exons are joined. This reaction is catalyzed by the spliceosome, a large and dynamic ribonucleoprotein complex composed of five small nuclear RNAs (snRNAs)—U1, U2, U4, U5, and U6—and more than 100 associated proteins. The snRNAs associate with proteins to form small nuclear ribonucleoproteins (snRNPs, pronounced "snurps").

The spliceosome recognizes specific sequence elements at the intron boundaries:

- The **5' splice site** (5'SS) has a consensus sequence of GU (or GURAGU, where R is a purine).
- The **branch point** is located 18–40 nucleotides upstream of the 3' splice site and contains an adenine that is the nucleophile in the first transesterification reaction.
- The **3' splice site** (3'SS) ends with AG, preceded by a polypyrimidine tract.

These consensus sequences are recognized by base-pairing with the snRNAs. The U1 snRNA base-pairs with the 5'SS, U2 snRNA base-pairs with the branch point, and U6 snRNA interacts with U2 and the 5'SS to position the reactive groups. The spliceosome assembles in a stepwise manner, with the U4/U6.U5 tri-snRNP joining last, followed by extensive conformational rearrangements that are driven by ATP hydrolysis by DExD/H-box RNA helicases.

### Splicing Reaction Steps

The chemistry of splicing involves two sequential transesterification reactions, which are described in detail in the [mRNA splicing](/knowledge/molecular-biology/mrna-splicing) article. Here is the ordered sequence:

1. **Assembly:** U1 snRNP binds the 5'SS, and U2 snRNP binds the branch point, forming the A complex. The U4/U6.U5 tri-snRNP joins to form the B complex.
2. **First transesterification:** The 2'-hydroxyl of the branch point adenine attacks the phosphate at the 5'SS. This cleaves the 5' exon from the intron and forms a lariat intermediate, in which the 5' end of the intron is covalently linked to the branch point adenine via a 2'→5' phosphodiester bond.
3. **Second transesterification:** The 3'-hydroxyl of the 5' exon attacks the phosphate at the 3'SS. This joins the two exons and releases the intron as a lariat structure.
4. **Disassembly:** The lariat intron is debranched by a debranching enzyme and degraded. The snRNPs are recycled for another round of splicing.

The entire reaction is catalyzed by RNA, with the U6 snRNA playing a central catalytic role, making the [spliceosome a ribozyme](/knowledge/molecular-biology/spliceosome-a-ribozyme). The reaction is reversible in principle, but in the cell it is driven forward by the energy of ATP hydrolysis used during assembly and conformational rearrangements.

### Alternative Splicing

Alternative splicing is the process by which different combinations of exons are joined to produce multiple distinct mRNAs from a single primary transcript. This is a major source of protein diversity in eukaryotes. It is estimated that over 95% of human multi-exon genes undergo alternative splicing, and the resulting isoforms often have distinct, even opposing, functions.

The main patterns of alternative splicing are:

- **Exon skipping:** An entire exon is excluded from the mature mRNA.
- **Intron retention:** An intron is retained in the mature mRNA, often leading to a truncated protein or nonsense-mediated decay.
- **Alternative 5' splice site selection:** A different 5'SS is used, extending or shortening the upstream exon.
- **Alternative 3' splice site selection:** A different 3'SS is used, extending or shortening the downstream exon.
- **Mutually exclusive exons:** Only one of two or more exons is included.

The choice of splice sites is regulated by **splicing enhancers** and **silencers**, which are cis-acting RNA sequences, and by **trans-acting factors** such as the SR proteins (serine/arginine-rich) and hnRNPs (heterogeneous nuclear ribonucleoproteins). SR proteins generally promote exon inclusion by binding enhancers and recruiting the spliceosome, while hnRNPs generally repress splicing by binding silencers and blocking spliceosome assembly. The relative concentrations of these factors in different cell types or developmental stages determine the splicing pattern, providing a powerful mechanism for tissue-specific gene regulation.

## 3' Polyadenylation

### Cleavage and Polyadenylation Factors

The 3' end of the pre-mRNA is processed by a two-step reaction: endonucleolytic cleavage followed by the addition of a poly(A) tail. This process is directed by two cis-acting elements in the pre-mRNA:

- The **[polyadenylation signal](/knowledge/molecular-biology/polyadenylation-signal)** (AAUAAA), located 10–30 nucleotides upstream of the cleavage site.
- A **G/U-rich downstream element**, located 20–40 nucleotides downstream of the cleavage site.

These elements are recognized by a large multiprotein complex. The key factors include:

- **CPSF** (cleavage and polyadenylation specificity factor), which recognizes the AAUAAA signal.
- **CstF** (cleavage stimulation factor), which binds the downstream G/U-rich element.
- **CFI and CFII** (cleavage factors I and II), which are required for the cleavage reaction.
- **PAP** (poly(A) polymerase), which catalyzes the addition of the poly(A) tail.
- **PABPN1** (poly(A)-binding protein nuclear 1), which binds the growing poly(A) tail and controls its length.

The cleavage reaction occurs first, releasing the downstream RNA, which is rapidly degraded. PAP then adds adenosine monophosphates to the 3' hydroxyl of the cleavage site. The initial poly(A) tail is approximately 200–250 nucleotides long in mammalian cells, though the length is controlled by PABPN1, which binds the tail and stimulates processive polyadenylation while limiting its length. In yeast, the tail is shorter, around 60–70 nucleotides.

### Poly(A) Tail Functions

The poly(A) tail serves several essential functions:

- **mRNA stability:** The poly(A) tail is bound by cytoplasmic poly(A)-binding protein (PABPC), which protects the mRNA from 3'→5' exonucleolytic degradation. Deadenylation—the progressive shortening of the poly(A) tail—is the first step in the major pathway of mRNA decay. The [property of mRNA](/knowledge/molecular-biology/property-of-mrna) stability is therefore closely tied to poly(A) tail length, and [mRNA instability](/knowledge/molecular-biology/mrna-unstable) is often initiated by accelerated deadenylation.
- **Translation initiation:** PABPC interacts with eIF4G, which is bound to the 5' cap via eIF4E. This interaction circularizes the mRNA, bringing the 5' and 3' ends into proximity. This circularization stimulates translation initiation by promoting ribosome recycling and re-initiation.
- **Nuclear export:** The poly(A) tail is recognized by the nuclear export receptor, which helps transport the mature mRNA through the nuclear pore complex.
- **Quality control:** The presence of a poly(A) tail distinguishes properly processed mRNAs from unprocessed or partially processed transcripts, which are retained in the nucleus and degraded.

## RNA Editing and Modifications

### Base Modification and Editing

RNA editing is a post-transcriptional process that changes the nucleotide sequence of the mRNA, thereby altering the protein product. This is distinct from splicing, which removes introns, and from capping and polyadenylation, which modify the ends. RNA editing can involve the substitution, insertion, or deletion of nucleotides.

The two most common types of RNA editing in eukaryotes are:

- **C-to-U editing:** Catalyzed by the APOBEC family of cytidine deaminases. The best-studied example is in the human APOB gene, where a C-to-U change in the mRNA creates a premature stop codon, producing a truncated protein (APOB48) in the intestine instead of the full-length APOB100 in the liver.
- **A-to-I editing:** Catalyzed by the ADAR (adenosine deaminase acting on RNA) family of enzymes. Adenosine is deaminated to inosine, which is read as guanosine by the translation machinery. A-to-I editing is widespread in the human transcriptome, particularly in Alu repeat elements in introns and untranslated regions, and it can alter codons, splice sites, and miRNA binding sites.

RNA editing is a regulated process that can be tissue-specific and developmentally regulated. It expands the coding capacity of the genome without requiring changes to the DNA sequence.

### Epitranscriptomics

Beyond RNA editing, mRNAs carry a variety of covalent modifications to their nucleotides. The most abundant internal modification in mRNA is **N6-methyladenosine (m⁶A)**, which is added co-transcriptionally by the METTL3/METTL14 methyltransferase complex and removed by the demethylases FTO and ALKBH5. m⁶A is recognized by reader proteins, such as YTHDF2, which promote mRNA degradation, and YTHDF1, which promotes translation.

The study of these reversible RNA modifications is called **epitranscriptomics**. m⁶A affects nearly every aspect of mRNA metabolism, including splicing, export, translation, and stability. Other modifications include 5-methylcytosine (m⁵C), N1-methyladenosine (m¹A), and pseudouridine (Ψ). The dynamic and reversible nature of these modifications provides another layer of post-transcriptional gene regulation, and their dysregulation has been linked to cancer, obesity, and neurological disorders.

## Coupling of Processing with Transcription

### The CTD Code

All three major processing events—capping, splicing, and polyadenylation—occur co-transcriptionally, while the pre-mRNA is still being synthesized by RNA polymerase II. This coupling is mediated by the C-terminal domain (CTD) of the largest subunit of RNA polymerase II, RPB1. The CTD consists of a repeated heptapeptide sequence, Tyr-Ser-Pro-Thr-Ser-Pro-Ser, which is repeated 52 times in humans and 26 times in yeast.

The CTD is dynamically phosphorylated during the transcription cycle, and this phosphorylation pattern constitutes a "CTD code" that recruits different processing factors at different stages:

- **Serine 5 phosphorylation (Ser5P)** is added during [transcription initiation](/knowledge/molecular-biology/transcription-initiation) by the kinase CDK7 (part of TFIIH). Ser5P recruits the capping enzyme, ensuring that capping occurs early in transcription.
- **Serine 2 phosphorylation (Ser2P)** is added during transcription elongation by the kinase CDK9 (part of the P-TEFb complex). Ser2P recruits splicing factors and the 3' processing machinery.
- **Serine 7 phosphorylation (Ser7P)** is also present during elongation and is important for the processing of certain snRNAs.
- **Tyrosine 1 phosphorylation (Tyr1P)** is enriched at the 5' ends of genes and may help prevent premature recruitment of 3' processing factors.

The phosphorylation state of the CTD is dynamic: phosphatases remove phosphates as the polymerase transcribes, and the pattern changes as the polymerase moves from the promoter to the terminator. This ensures that processing factors are recruited at the correct time and place.

### Processing Factors and Transcription

The physical coupling of transcription and processing has several important consequences. First, it increases the efficiency of processing: the processing factors are present at high local concentrations near the nascent RNA, so they can act as soon as the relevant sequences emerge from the polymerase. Second, it provides a quality control mechanism: if processing is defective, transcription can be slowed or terminated, preventing the production of aberrant mRNAs. Third, it allows for feedback regulation: processing factors can influence transcription elongation, and transcription rate can influence splicing decisions.

For example, a slow RNA polymerase II can allow more time for weak splice sites to be recognized, promoting exon inclusion, while a fast polymerase can promote exon skipping. This kinetic coupling between transcription and splicing is an important mechanism for the regulation of alternative splicing.

## Methods to Study mRNA Processing

### RNA Sequencing and Bioinformatics

RNA sequencing (RNA-seq) is the most widely used method to study mRNA processing on a genome-wide scale. In a typical RNA-seq experiment, total RNA is isolated, poly(A)-selected to enrich for mRNA, fragmented, reverse-transcribed into cDNA, and sequenced on a high-throughput platform. The resulting reads are aligned to the genome, and the coverage and junction patterns are analyzed.

RNA-seq can reveal:

- **Splicing patterns:** Reads that span exon-exon junctions indicate which splice sites are used. Tools such as rMATS and MISO quantify alternative splicing events and compare them between conditions.
- **Polyadenylation site usage:** Reads that map to the 3' ends of genes can reveal alternative polyadenylation. Specialized protocols such as 3'Seq or PolyA-seq enrich for 3' ends to quantify poly(A) site usage.
- **RNA editing:** Mismatches between the RNA-seq reads and the reference genome can indicate RNA editing events, though this requires careful filtering to exclude sequencing errors and SNPs.

RT-PCR (reverse transcription-[polymerase chain reaction](/knowledge/molecular-biology/polymerase-chain-reaction)) is a more targeted method. In a typical RT-PCR experiment, RNA is reverse-transcribed using an oligo(dT) primer or random hexamers, and the resulting cDNA is amplified by PCR. Primers flanking a region of interest can be used to detect splicing isoforms, which are separated by gel electrophoresis or quantified by real-time PCR. For example, to detect exon skipping, primers are designed in the flanking exons; the PCR product will be shorter if the exon is skipped. The cycling conditions typically involve 30–40 cycles of denaturation at 95°C for 30 seconds, annealing at 55–60°C for 30 seconds, and extension at 72°C for 30 seconds per kilobase.

### In Vitro Processing Assays

In vitro assays are used to dissect the biochemical mechanisms of mRNA processing. These assays use nuclear extracts, typically derived from HeLa cells, which contain all the necessary processing factors.

- **In vitro capping assay:** A radiolabeled RNA substrate is incubated with HeLa nuclear extract in the presence of GTP and S-adenosylmethionine (the methyl donor). The reaction is incubated at 30°C for 30–60 minutes, and the products are analyzed by gel electrophoresis. The cap structure can be detected by its resistance to nuclease digestion or by immunoprecipitation with anti-cap antibodies.
- **In vitro splicing assay:** A radiolabeled pre-mRNA substrate containing two exons and one intron is incubated with HeLa nuclear extract under splicing conditions (typically 20 mM HEPES pH 7.9, 3.2 mM MgCl₂, 0.5 mM ATP, 20 mM creatine phosphate, and 2.6% polyvinyl alcohol) at 30°C for 1–2 hours. The RNA is then extracted and analyzed by denaturing polyacrylamide gel electrophoresis. The lariat intermediate and lariat intron migrate more slowly than linear RNAs of the same length, allowing the reaction products to be identified.
- **In vitro polyadenylation assay:** A radiolabeled RNA substrate containing the AAUAAA signal and downstream element is incubated with HeLa nuclear extract in the presence of ATP. The reaction is incubated at 30°C for 30–60 minutes, and the products are analyzed by gel electrophoresis. The addition of the poly(A) tail is detected as a smear of higher molecular weight products.

These assays are powerful tools for identifying the sequence elements and protein factors required for processing, and they can be used to test the effects of mutations or inhibitors.

## Common Pitfalls and Misconceptions

### Order of Processing Events

A common error is to think of mRNA processing as a strictly sequential pathway in which capping, splicing, and polyadenylation occur in a fixed order. In reality, these events are co-transcriptional and can overlap. Capping occurs first, when the transcript is only 20–30 nucleotides long, but splicing of the first intron can begin while the polymerase is still transcribing downstream exons. Polyadenylation occurs last, after transcription has terminated, but the 3' processing factors are recruited to the CTD early in elongation.

Another misconception is that all introns are removed before the mRNA is exported. In fact, some introns can be retained in the cytoplasm, and a process called "intron retention" can produce mRNAs that are translated into truncated proteins or targeted for nonsense-mediated decay. The order of processing is therefore not fixed, and the cell has quality control mechanisms to detect and degrade improperly processed mRNAs.

### Nuclear vs. Cytoplasmic Processing

All three canonical processing events—capping, splicing, and polyadenylation—occur in the nucleus. The mature mRNA is then exported to the cytoplasm for translation. However, some processing events can also occur in the cytoplasm:

- **RNA editing** can occur in both the nucleus and the cytoplasm, depending on the enzyme and the target.
- **m⁶A modification** can be added or removed in the cytoplasm, and it can affect translation and stability.
- **Poly(A) tail length** is dynamically regulated in the cytoplasm by deadenylases and cytoplasmic poly(A) polymerases. Cytoplasmic polyadenylation can activate translation of maternal mRNAs during oocyte maturation and early development.

Students often confuse the nuclear processing of pre-mRNA with the cytoplasmic events that regulate mRNA stability and translation. It is important to remember that the 5' cap and poly(A) tail are added in the nucleus, but their functions in translation and stability are mediated by cytoplasmic factors.

### Common Confusions

- **Capping vs. polyadenylation:** The 5' cap is added to the 5' end and involves a modified guanosine; the poly(A) tail is added to the 3' end and consists of a homopolymer of adenosines. They are added by completely different enzymes and serve different functions.
- **Splicing vs. RNA editing:** Splicing removes introns and joins exons; RNA editing changes individual nucleotides within the mRNA. Splicing is catalyzed by the spliceosome; RNA editing is catalyzed by deaminases.
- **Introns vs. exons:** Introns are removed during splicing; exons are retained in the mature mRNA. The terms are relative to the final mRNA, not to the genomic sequence.
- **Pre-mRNA vs. mRNA:** Pre-mRNA is the primary transcript that contains introns; mRNA is the mature, processed transcript that is exported to the cytoplasm. The terms are not interchangeable.

## Frequently Asked Questions

### What are the steps of mRNA processing in eukaryotes?

The three canonical steps are: (1) 5' capping, in which a 7-methylguanosine cap is added to the 5' end; (2) splicing, in which introns are removed and exons are joined; and (3) 3' polyadenylation, in which the pre-mRNA is cleaved and a poly(A) tail is added. These events occur co-transcriptionally in the nucleus. Additional modifications, such as RNA editing and base methylation, can also occur.

### What are the types of mRNA processing?

The main types are 5' capping, splicing (including alternative splicing), and 3' polyadenylation. In addition, RNA editing (C-to-U and A-to-I) and epitranscriptomic modifications (such as m⁶A) are considered forms of mRNA processing. These processes are collectively referred to as [RNA processing](/knowledge/molecular-biology/rna-processing).

### Why is mRNA processing important?

mRNA processing is essential for producing a functional, stable mRNA that can be translated into protein. It removes non-coding introns, protects the mRNA from degradation, promotes translation initiation, and enables nuclear export. Processing also provides opportunities for regulation: alternative splicing and RNA editing can produce multiple protein isoforms from a single gene, and the [importance of mRNA](/knowledge/molecular-biology/mrna-important) in gene expression is largely determined by these processing events.

### Where does mRNA processing occur?

The three canonical processing events occur in the nucleus, co-transcriptionally. The mature mRNA is then exported to the cytoplasm for translation. Some modifications, such as RNA editing and m⁶A methylation, can also occur in the cytoplasm, and poly(A) tail length is dynamically regulated in the cytoplasm.

### What is the role of the 5' cap in mRNA processing?

The 5' cap protects the mRNA from 5'→3' exonucleases, promotes translation initiation by binding eIF4E, enhances splicing of the first intron, and facilitates nuclear export. It is added co-transcriptionally when the nascent transcript is only 20–30 nucleotides long.

### How does alternative splicing affect gene expression?

Alternative splicing allows a single gene to produce multiple mRNA isoforms, which can encode proteins with different, even opposing, functions. This greatly expands the coding capacity of the genome and provides a mechanism for tissue-specific and developmental regulation of gene expression. It is estimated that over 95% of human multi-exon genes undergo alternative splicing.

### What is the function of the poly(A) tail?

The poly(A) tail protects the mRNA from 3'→5' degradation, promotes translation initiation by interacting with eIF4G, and facilitates nuclear export. The length of the poly(A) tail is dynamically regulated and is a key determinant of mRNA stability. Deadenylation is the first step in the major pathway of mRNA decay.

## Key Takeaways

- mRNA processing in eukaryotes consists of three canonical co-transcriptional events: 5' capping, splicing, and 3' polyadenylation, all occurring in the nucleus.
- The 5' cap (m⁷G) protects the mRNA from degradation and is essential for translation initiation via eIF4E binding.
- Splicing removes introns via two transesterification reactions catalyzed by the spliceosome, a large RNA-protein complex; alternative splicing generates protein diversity.
- The poly(A) tail is added by PAP after cleavage of the pre-mRNA and is bound by PABP, which protects the mRNA and promotes translation.
- The C-terminal domain of RNA polymerase II coordinates processing by recruiting processing factors through its phosphorylation pattern (the CTD code).
- RNA editing and epitranscriptomic modifications such as m⁶A add further layers of regulation and expand the coding capacity of the genome.
- mRNA processing is a major point of gene regulation, and defects in processing are linked to numerous human diseases, including cancer and neurological disorders.

## Further Reading

- Boreikaitė V, Passmore LA. *3'-End Processing of Eukaryotic mRNA: Machinery, Regulation, and Impact on Gene Expression*. Annual review of biochemistry. 2023. [PubMed 37001138](https://doi.org/10.1146/annurev-biochem-052521-012445)
- Martinez NM et al. *Pseudouridine synthases modify human pre-mRNA co-transcriptionally and affect pre-mRNA processing*. Molecular cell. 2022. [PubMed 35051350](https://doi.org/10.1016/j.molcel.2021.12.023)
- Fliedner A et al. *Variants in SCAF4 Cause a Neurodevelopmental Disorder and Are Associated with Impaired mRNA Processing*. American journal of human genetics. 2020. [PubMed 32730804](https://doi.org/10.1016/j.ajhg.2020.06.019)
- Uggenti C et al. *cGAS-mediated induction of type I interferon due to inborn errors of histone pre-mRNA processing*. Nature genetics. 2020. [PubMed 33230297](https://doi.org/10.1038/s41588-020-00737-3)
- Biswas J et al. *Aberrant pre-mRNA processing in cancer*. The Journal of experimental medicine. 2024. [PubMed 39316554](https://doi.org/10.1084/jem.20230891)
- Biswas B, Vagner S. *Genotoxic stress impacts pre-mRNA 3'-end processing*. BioEssays : news and reviews in molecular, cellular and [developmental biology](/blog/careers/developmental-biology). 2024. [PubMed 39030821](https://doi.org/10.1002/bies.202400037)

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* [MAPK Pathway: Mechanism, Function, and Clinical Relevance](/knowledge/molecular-biology/mapk-pathway)
* [Mammalian Cell Culture Bioreactors: A Practical Guide](/knowledge/molecular-biology/mammalian-cell-culture-bioreactor)
* [Nucleotide Formation: Biosynthesis and Assembly of DNA/RNA Building Blocks](/knowledge/molecular-biology/nucleotide-formation)