RNA processing
all chemical modifications required to generate the final RNA product
Exons
coding sequences
Introns
noncoding regions
Coding sequences in eukaryotic genes
- In many eukaryotic genes the coding sequences are not continuous.
- The coding sequences (exons) are interrupted by non-coding sequences (introns); both exons and introns are transcribed into RNA
- RNA processing steps are required to remove intron to generate mRNA for translation; the mRNA has only the sequences of the exons.
- Number of introns and exons varies in different genes.
- The human dystrophin gene is 2 million bases long and contains 78 introns (99% DNA of the gene)
Processing of mRNA
- RNA is modified in the nucleus by
- Capping at the 5` end
- Addition of a poly A tail at the 3` end
- Removal of introns
- RNA editing
- Mature mRNA is transported through nuclear pores to the cytoplasm, where it is translated.
mRNA capping
a methylated guanine base is added, in reverse orientation, at the 5` end of the primary transcript
Role of cap
- enhances splicing of the mRNA.
- required for transport of the mRNA to the cytoplasm.
- increases the translatability of the mRNA.
- protects the mRNA against nuclease degradation.
Polyadenylation of mRNA
The enzyme Poly A polymerase adds the poly A tail
Polydenylation of mRNA functions
Protects from nuclease attack.
Helps in its transport into the cytoplasm
Helps ribosomes in recognizing mRNA as a molecule to be translated
Removal of introns
The ends of nuclear introns are defined by the GU-AG rule
Intron removal by splicesomes
Self splicing introns (ribozymes)
Intron removal by splicesomes
- Intron removal for most eukaryotic genes is catalysed by spliceosome that are made of small nuclear ribonucleoproteins (snRNPs, pronounced snurps) and additional proteins.
- Each snRNP contain one or more of snRNA (small nuclear RNA).
Self splicing introns (ribozymes)
Do not require snRNPs for splicing
Commonly found in plant and fungal organellar genomes
Alternative splicing
One gene can yield different proteins by alternative splicing
Changing the splice sites may:
- introduce termination codons (shown by asterisks)
- change reading frames.
Why do eukaryotes have introns
- Some of the miRNAs come from introns.
- Alternative splicing can generate different types of mRNA and hence different polypeptides from the same gene.
- In humans there are roughly 35,000 genes that code for more then 100,000 proteins.
- Sex determination in drosophila is through alternative splicing
- Role in evolution
- Exons often code for a domain in a multidomain polypeptide chain.
- Presence of introns make exons shuffling easier allowing different exons from different genes coming together to generate novel genes.
RNA editing
- A process in which information changes at the level of mRNA
- Nucleotide sequence of mRNA must be the same as sequence of the gene (without introns)
- There are some exceptions
- In trypanosome mitochondria:
The cytochrome oxidase coxII mRNA contains extra 4Us
The coxIII mRNA contains extra 407 Us and lacks 19 Us
RNA editing by addition or deletion of bases
- Pre-edited RNA base pairs with a guide RNA on both sides of the region to be edited.
- The guide RNA provides a template for the insertion of uridines.
- The mRNA produced by the insertions is complementary to the guide RNA
- Addition or deletion of U residues occurs by cleavage of the RNA, removal or addition of the U, and ligation of the ends
- The reactions are catalyzed by a complex of enzymes under the direction of guide RNA.
RNA editing by base conversion
- Sometime an existing base in mRNA is converted into another base.
- The sequence of the apo-B gene is same in intestine and liver, but the sequence of the mRNA is modified in the intestine by a base change that creates a termination codon
Processing of transfer RNAs (tRNA)
- The precursor tRNA is cleaved to produce individual tRNA
- An intron is removed by splicing
- Bases are added at the 3` end
- A number of bases are modified; the unusual bases in tRNAs are the result of this modification
Processing of ribosomal RNA (rRNA)
- All rRNAs get transcribed as a single RNA
- A number of bases and 2`-C of some ribose sugars are modified by methylation
- The long chain is cleaved to produce intermediate rRNA
- The intermediates are trimmed to produce mature rRNAs.
