Transcript second of Chapter 10: RNA processing

Chapter 10
Transcription
RNA
processing
Translation
Jones and Bartlett Publishers © 2005
Terms (mRNA)
•
•
•
•
primary transcript
coding sequence (open reading frame, ORF)
prokaryotic mRNA = primary transcript
Eukaryotic transcripts are converted into
mRNA through RNA processing:
– Modification of the 5’ end
– Extension of 3’ end
– Excision of untranslated embedded sequences.
The Central Dogma
Polycistronic messages
• In bacteria and eukaryotic organelles,
sometimes more than one polypeptide is
encoded in a single message.
• These are called polycistronic mRNAs.
Polycistronic mRNAs
Eukaryotic Transcription
• In eukaryotes, three different polymerases
are used.
• No polycistronic mRNAs are produced.
(All eukarytoic mRNAs are monocistronic.)
• The mRNA molecules are chemically
modified in eukaryotes.
Eukaryotic RNA Polymerases
Enzyme
Cellular location
RNA polymerase I
Nucleolus
Product
rRNA
RNA polymerase II
Nucleus
mRNA
snRNA
RNA polymerase III
Nucleus
tRNA,
5S RNA,
snRNA
Promoters for eukaryotic genes
• There is more variability in promoters and
termination sequences.
• TATA box : 5’-TATAAA-3’, -25 bp, with
GC regions near it.
• CAAT box: GCCAATCT, -80 bp
• GC box: GGGCGC, multiple copies.
Promoters in eukaryotes
Modification of primary transcripts
• Three major alterations to mRNA:
– Modified guanosine cap at 5’ end
– 3’ end gets a poly(A) tail.
– pre-mRNA is processed to remove specific
internal sequences by splicing them out.
5’ cap in eukaryotes
Polyadenylation
• The poly(A) tail is added by poly(A)
polymerase.
• About 200 adenosines are added to the 3’
end of the primary transcript.
• Poly(A) tails help geneticists isolate mRNAs
from cells.
RNA Splicing
• In the 1960s, it was observed that RNA
molecules were larger than predicted, based
on protein structure.
• In 1977, internal, non-coding sequences
were discovered.
• These internal, non-coding sequences are
called introns.
Spliceosomes and the GU-AG rule
• Intron-Exon junctions have a conserved
sequence that is recognized by an enzyme
complex (spliceosome).
• The sequence is GU-AG, and is almost
universal.
• A spliceosome has nucleic acids, 8-10
proteins, and snRNAs.
RNA processing
Transcription
and RNA
processing are
coupled
processes.
Molecular details of the splicing of an intron
Introns begin
with a GU and
end with an AG.
There is also an
internal A
nucleotide
where the
branch junction
is formed.
The spliced
intron is known
as a lariat (with
a loop and tail).
Splicing of introns is mediated by small nuclear
ribonucleoprotein particles (snRNPs)
In this
diagram,
only the RNA
component of
the snRNPs
is shown.
Base pairing
between
snRNA and
the intron is
involved in
the splicing
reaction.
DNA-RNA hybridization showing the parts of DNA
that are missing from the mature mRNA and the
part of the mRNA (polyA tail) missing in the DNA
Mutation in a splice site may result in the
retention of the entire (or part) of an intron
Exon-shuffle model
• Introns may play a role in gene evolution.
• In some proteins, each exon has its own
independent folding characteristics.
• Folding domains (=exons) can be grouped
together to give new proteins with new
functions.
• This is called the exon-shuffle model.
• Not all genes have domain boundaries that
correlate to exons.
Transcription
occurs in the
nucleus, as does
RNA
processing.
Export to the
cytoplasm
happens before
translation
occurs.
Translation of mRNA into amino acid sequences
• The genetic code consists of three bases.
• A codon is the ‘code word’ for each amino
acid.
• 4 x 4 x 4 =64, but only 20 amino acids are
used in proteins, so the genetic code is said
to be degenerate.