1. RNA post-transcriptional processing

2. RNA Processing
Very few RNA molecules are transcribed directly into the final mature RNA.
Most newly transcribed RNA molecules (primary transcripts) undergo various alterations to yield the mature product
RNA processing is the collective term used to describe the molecular events allowing the primary transcripts to become the mature RNA.

3. primary transcript mature RNA. Cytoplasm Nucleus or Nucleolus
RNA processing
Removal of nucleotides
addition of nucleotides to the 5’- or 3’- ends
modification of certain nucleotides
mature RNA.

4. Processing Events in Prokaryotes vs Eukaryotes

5. endonucleases to cut at specific sites within a precursor RNA
(1) Removal of nucleotides by both endonucleases and exonucleases
endonucleases to cut at specific sites within a precursor RNA
exonucleases to trim the ends of a precursor RNA
This general process is seen in prokaryotes and eukaryotes for all types of RNA

6. (2) Addition of nucleotides to 5’-or 3’-ends of the primary transcripts or their cleavage products.
Add a cap and a poly(A) tail to pre-mRNA
AAAAAA

7. tRNA 3-D structure

8. tRNA processing in prokaryotes
Mature tRNAs are generated by processing longer pre-tRNA transcripts, which involves
specific exo- and endonucleolytic cleavage by RNases D, E, F and P (general) followed by
base modifications which are unique to each particular tRNA type.

9. Primary transcripts RNase D,E,F and P tRNA with mature ends
Base modifications
mature tRNAs

10. tRNA processing in eukaryotes
The pre-tRNA is synthesized with a
16 nt 5’-leader,
a 14 nt intron and
two extra 3’-nucleotides.

12. Primary transcripts forms secondary structures recognized by endonucleases
5’ leader and 3’ extra nucleotide removal
tRNA nucleptidyl transferase adds 5’-CCA-3’ to the 3’-end to generate the mature 3’-end
Intron removal

13. Processing of mRNA: prokaryotes
There is essentially no processing of prokaryotic mRNA, it can start to be translated before it has finished being transcribed.
Prokaryotic mRNA is degraded rapidly from the 5’ end

14. Processing of mRNA in eukaryotes
Pre-mRNA molecules are processed to mature mRNAs by
5’-capping
3’Cleavage and polyadenylation
splicing

15. Eukaryotic mRNA processing: overview

16. 5’ Capping
Very soon after RNA Pol II starts making a transcript, and before the RNA chain is more then nt long, the 5’-end is chemically modified.
7-methylguanosine is covalently to the 5´ end of pre-mRNA.
Linked 5´  5´
Occurs shortly after initiation

17. 7-methylguanosine (m7G)

18. Function of 5´cap Protection from degradation
Increased translational efficiency
Transport to cytoplasm
Splicing of first exon

19. 3’ Cleavage and polyadenylation
In most pre-mRNAs, the mature 3’-end of the molecule is generated by cleavage followed by the addition of a run, or tail, of A residues which is called the poly(A) tail.

20. RNA polymerase II does not usually terminate at distinct site
Pre-mRNA is cleaved ~20 nucleotides downstream of polyadenylation signal (AAUAAA)
~250 AMPs are then added to the 3´ end
Almost all mRNAs have poly(A) tail

21. Function of poly(A) tail
Increased mRNA stability
Increased translational efficiency
Splicing of last intron
AAAAAA

22. Splicing
The process of cutting the pre-mRNA to remove the introns and joining together of the exons is called splicing.
It takes place in the nucleus before the mature mRNA can be exported to the cytoplasm.

23. Introns: non-coding sequences
Exons: coding sequences
RNA splicing: removal of introns and joining of exons
Splicing mechanism must be precise to maintain open reading frame
Catalyzed by spliceosome (RNA + protein)