Download Other RNA Processing Events

Other RNA Processing Events
Chapter 16
Ribosomal RNA Processing
• rRNA genes of both eukaryotes and bacteria
are transcribed as larger precursors must be
processed to yield rRNAs of mature size
- Several different rRNA molecules are
embedded in a long precursor and each must
be cut out
Eukaryotic rRNA processing
• The rRNA are separated by regions called
nontranscribed spacers (NTSs).
• Transcribed spacers - regions of the gene that
are transcribed as part of rRNA precursor
- and then removed in processing of precursor
to mature rRNA species.
Eukaryotic rRNA Processing
• Ribosomal RNAs are made in eukaryotic
nucleoli as precursors that must be processed
to release mature rRNAs
• Order of RNAs in the precursor is (fig: 16.2)
–
–
–
–
18S
5.8S
28S in all eukaryotes
Exact sizes of the mature rRNAs vary from one
species to another
Eukaryotic rRNA Processing
• rRNA processing –
nucleolus – small nucleolar
RNAs (snoRNAs) +
proteins = Small nucleolar
ribonucleoproteins
(snoRNPs)
• E1, E2 and E3 – interact
with distinct regions in prerRNA
Bacterial rRNA Processing
• Bacterial rRNA precursors contain tRNA
and all 3 rRNA
• rRNA are released from their precursors by
RNase III and RNase E
– RNase III is the enzyme that performs at least the
initial cleavages that separate the individual
large rRNAs
– RNase E is another ribonuclease that is
responsible for removing the 5S rRNA from the
precursor
Processing Bacterial rRNA
Transfer RNA Processing
• Transfer RNAs are made in all cells as overly
long precursors
– These must be processed by removing RNA at
both ends
• Nuclei of eukaryotes contain precursors of a
single tRNA
• In bacteria - precursor may contain one or
more tRNA
- Sometimes a mixture of rRNAs and tRNAs
Cutting apart Polycistronic
Precursors
• In processing bacterial RNA that contain more
than one tRNA
– First step is to cut precursor up into fragments with
just one tRNA each
• Cutting between tRNAs in precursors having 2 or more
tRNA
• Cutting between tRNAs and rRNAs in precursors
– Enzyme that performs both chores is the RNase III
Forming Mature 5’-Ends of tRNA
• Extra nucleotides are
removed from the 5’ends of pre-tRNA in
one step by an
endonucleolytic
cleavage catalyzed by
RNase P
• RNase P from bacteria
and eukaryotic nuclei
have a catalytic RNA
subunit called M1 RNA
Processing 3’-Ends of tRNA
RNase D, RNase BN, RNase T, RNase PH, RNase II and polynucleotide
phosphorylase (PNPase)
Forming mature 3’-Ends of tRNA
• RNase II and polynucleotide phosphorylase
cooperate
– To remove most of extra nucleotides at the end of
a tRNA precursor
• RNases PH and T are most active in
removing the last 2 nucleotides from RNA
– RNase T is the major participant in removing very
last nucleotide
Trans-Splicing
• Splicing that occurs in all eukaryotic species
is called cis-splicing because it involves 2 or
more exons that exist together in the same
gene
• trans-splicing has exons that are not part of
the same gene at all - may not even be on
the same chromosome
The Mechanism of Trans-Splicing
• Trans-splicing occurs in several organisms
– Parasitic and free-living worms
– First discovered in trypanosomes
• Trypanosome mRNA are formed by transsplicing between a short leader exon and any
one of many independent coding exons
Trans-Splicing
Trans-Splicing Scheme
• Branchpoint adenosine
within the half-intron
attached to the coding
exon attacks the junction
between the leader exon
and its half-intron
• Creates a Y-shaped
intron-exon intermediate
analogous to the lariat
intermediate
Mechanism of Editing
• Unedited transcripts can be found along with
edited versions of the same mRNAs
• Editing occurs in the poly(A) tails of mRNAs
that are added posttranscriptionally
• Partially edited transcripts have been isolated always edited at their 3’-ends but not at their
5’-ends
Role of gRNA in Editing
• Guide RNAs (gRNA)
could direct the
insertion and deletion of
UMPs over a stretch of
nucleotides in the
mRNA
• When editing is done,
gRNA could hybridize
near the 5’-end of newly
edited region
Guide RNA Editing
• 5’-end of the first gRNA hybridizes to an
unedited region at the 3’-border of editing I the
pre-mRNA
• The 5’-ends of the rest of the gRNAs hybridize
to edited regions progressively closer to the 5’end of the region to be edited in the premRNA
• All of these gRNAs provide A’s and G’s as
templates for the incorporation of U’s missing
from the mRNA
Mechanism of Removing U’s
• Sometimes the gRNA is missing an A or G to pair
with a U in the mRNA
– In this case the U is removed
• Mechanism of removing U’s involves
– Cutting pre-mRNA just beyond U to be removed
– Removal of U by exonuclease
– Ligating the two pieces of pre-mRNA together
• Mechanism of adding U’s uses same first and last
step
• Middle step involves addition of one or more U’s
from UTP by TUTase
RNA Interference
• RNA interference occurs when a cell encounters
dsRNA from:
– Virus
– Transposon
– Transgene
• Trigger dsRNA is degraded into 21-23 nt fragments
(siRNAs) by an RNase III-like enzyme called Dicer
• Double-stranded siRNA, with Dicer and Dicerassociated protein R2D2 form a complex called
complex B
Complex B
• Complex B delivers the siRNA to the RISC loading
complex (RLC)
– Separates 2 strands of siRNA
– Transfers guide strand to RNA-induced silencing complex
(RISC) that introduces a protein- Ago2
Complex B
• The guide strand of siRNA base-pairs with target
mRNA in the active site of PIWI domain of Ago2
– Ago2 is an RNase H-like enzyme known as a slicer
– Slicer cleaves the target mRNA in middle of the region of
its base-pairing with the siRNA
– ATP-dependent step has cleaved RNA ejected from RISC
which then accepts a new molecule of mRNA for
degradation
•
•
•
This project is funded by a grant awarded under the President’s Community Based Job Training Grant as
implemented by the U.S. Department of Labor’s Employment and Training Administration (CB-15-162-06-60).
NCC is an equal opportunity employer and does not discriminate on the following basis:
against any individual in the United States, on the basis of race, color, religion, sex, national origin, age disability,
political affiliation or belief; and
against any beneficiary of programs financially assisted under Title I of the Workforce Investment Act of 1998
(WIA), on the basis of the beneficiary’s citizenship/status as a lawfully admitted immigrant authorized to work in
the United States, or his or her participation in any WIA Title I-financially assisted program or activity.
Disclaimer
• This workforce solution was funded by a grant awarded under the
President’s Community-Based Job Training Grants as implemented
by the U.S. Department of Labor’s Employment and Training
Administration. The solution was created by the grantee and does
not necessarily reflect the official position of the U.S. Department of
Labor. The Department of Labor makes no guarantees, warranties,
or assurances of any kind, express or implied, with respect to such
information, including any information on linked sites and including,
but not limited to, accuracy of the information or its completeness,
timeliness, usefulness, adequacy, continued availability, or
ownership. This solution is copyrighted by the institution that
created it. Internal use by an organization and/or personal use by
an individual for non-commercial purposes is permissible. All other
uses require the prior authorization of the copyright owner.