Physiology 3140A Lecture Notes - Lecture 30: Small Nucleolar Rna, Non-Coding Rna, Small Interfering Rna
2 May 2018
School
Department
Course
Professor
Cell Physiology Lecture 30
Non coding RNA
Epigenetics
Heritable changes in gene expression that do not involve any change in DNA sequence but modifications in
the chromatin.
Three general types:
1. Modification of histone core proteins
- Could be phosphorylation, ubiquination, methylation, sumoylation and acetylation
- Can be associated with repression or activation
o Includes chromatin remodeling proteins
2. DNA methylation
- Generally at cystidines
- Usually associated with gene repression
3. microRNAs/non coding RNA
- Affect transcription, silence genomic regions or alter RNA processing all leading to changes in RNA
accumulation and expression
o Third level of epigenetic regulation
o Regulating expression of genes but not causing any changes in the actual DNA sequence
Non-coding (ncRNA)
- Known protein coding gene exons compose less than 3% of the human genome
- Based on the ENCODE* database, approximately 76% of the human genome is transcribed
- Most of these transcripts do not code for protein called non-coding RNAs (ncRNAs)
- Separated into two classes: short ncRNAs and long ncRNAs
o MOST FALL IN THE SHORT – LESS THAN 200 BP
- Long ncRNAs are arbitrarily defined as being longer than 200 nucleotides
Types of ncRNA:
- Transfer RNA (tRNA) – most common
- Ribosomal RNA (rRNA) – most common
- snoRNA – small nucleolar RNA; guide modifications of other RNAs
- snRNA – small nuclear RNA; aid in the processing of pre-mRNA
- piRNA – piwi-interacting RNA; gene silencing of retrotransposons
- exRNA – extracellular RNA; possibly cell communication
o People that have specific diseaseas have different expression of extracellular RNA in the blood
o Assess changes in it and how it reflects a disease or developmental problem
- siRNA – small interfering RNA; promotes mRNA degradation
- microRNA (miRNA) – RNA silencing and post-transcriptional regulation
o Similar to how it works with siRNA but differences in how they target genes and how they are
expressed
- scaRNA – Small Cajal body RNA; role in RNA maturation
- lncRNA – long non-coding RNAs
find more resources at oneclass.com
find more resources at oneclass.com
microRNAs:
- Short
- Hairpin derived RNAs ~20-24 nucleotides long
o FUNCTIONAL PART
- Typically bind the 3’ UTR of mRNA leading to repression
o 3’ UTR is the target for microRNA
- Computational analysis suggests that 60% of protein-coding RNAs can be targeted by miRNAs
- Recent studies describe 2,588 miRNAs (http://www.mirbase.org/)
o Some are cell-specific, condition-specific (temporal time point)
- Typically produces subtle (<2-fold) reduction in protein levels
o Effects on RNA are subtle – many of them only cause small change in expression
o Expression might be enough to cause difference in the protein works or functional outcome
- Have been linked to regulation of developmental processes, fine tuning gene expression, disease
progression and chemotherapeutic resistance
- miRNAs can exist, and be transcribed, as a cluster. Therefore, multiple miRNAs produced
- Most protein coding RNA – most of them have sequences within their untranslated region, that is a
target for microRNA
- Many of the microRNA may be able to target multiple genes
- MicroRNA are often expressed as a tandem of RNA
o Transcript made – there will be 3-4 microRNA expressed
- 2 THINGS:
o 1) Get one large transcript making 3+ microRNA
o 2) Can have one microRNA that might target 10-20 genes
- If you want to target a pathway, the most efficient way of doing it is by having one microRNA target all
the parts of the pathway
o Even though it is made into 3 different microRNA, they all might target same pathway OR
they do not regulate the same pathway but use the same promoter region upstream to
generate the expression
- NOT EVERY NEW MICRORNA NEEDS NEW PROMOTER AND REGULATORY ELEMENTS
siRNAs:
- More of a research tool
o Proteins expressed in lower organisms and have a function that can be used for research
o Not expressed in high mammalian cells
- Originates from dsRNA and is usually 21-22 nucleotides long
o Ds-RNA makes them more stable and more susceptible to particular types of RNA
- Primary role in repression is to cleave mRNA
o ONLY INVOLVED IN DNA REPRESSION (whereas microRNA had multiple functions)
- Often the response to viral infection and provide a perfect match to its target (therefore typically
has a single target)
o While microRNA target multiple genes, siRNA target a SINGLE gene
- Often used as a research tool to knock down expression by as much as 85-90%
o Over express the siRNA and target the specific gene and knock it down
o
- Unsure if siRNA is produced in mammalian cells
find more resources at oneclass.com
find more resources at oneclass.com
- Differences in some places, but in others the use the same machinery
MicroRNA
- miRNA – have gene that is expressed
o Similar to mRNA transcript
- Secondary folding comes about – there is complementarity
o Fold the RNA like this because it increases stability & protects it from degradation
o tRNA are the same way!!! – secondary structure is folded up with complementarity
- NOTE: the microRNA are not the perfect match
o They will not perfectly match what it is targeting, but it has enough complimentary to still
bind
- They are outside the nucleus and they interact with DICER complex
- DICER cuts off regions of either the siRNA or microRNA that are not important for targeting
o Now have 20-25 bp sequence of RNA, double stranded, that is now going to be used for
targeting
- How do you take the double stranded RNA and separate it
o Have one copy of the strand that is going to target RNA – done by RISC-Argonaute complex
- Type of argonaute protein that is interacting differs between siRNA and microRNA
Occurrence
Configuration
Length
Length
Complementarity
to target mRNA
Biogenesis
Action
Function
Clinical
uses
miRNA
Occur
naturally in
plants and
animals (All
cell
populations)
GENERATED
AS Single
stranded
(may fold back
up on
themselves to
generate
double
strands)
19–25
nt
Not exact, and
therefore a single
miRNA may target
up to hundreds of
mRNAs
Expressed by
genes whose
purpose is to
make miRNAs,
but regulate
genes other
than the ones
that expressed
them
May expressed
as group of
microRNA
Inhibit
translation
of mRNA –
bind +
prevent
translation
Regulators
(inhibitors)
of genes
(mRNAs)
MODULATE
EXPRESSION
(small
changes)
Possible
therapeutic
uses.
Can be
used as
potential
diagnostic
and
biomarker
tools
siRNA
Naturally in
plants and
lower
animals.
Unclear if in
mammals.
Double
stranded
21–22
nt
100% perfect
match, and
therefore siRNAs
knock down
specific genes,
with minor off-
target exceptions
Regulate the
same genes that
express them
Cleaves
mRNA
Bind mRNA
& cause
cleavage +
degradation
Act as gene-
silencing
guardians in
plants and
animals
SILENCING
Valuable
laboratory
tools to
knock
down
genes. In
clinical
trials as
possible
therapeutic
agents
find more resources at oneclass.com
find more resources at oneclass.com
Document Summary
Heritable changes in gene expression that do not involve any change in dna sequence but modifications in the chromatin. Could be phosphorylation, ubiquination, methylation, sumoylation and acetylation. Known protein coding gene exons compose less than 3% of the human genome. Based on the encode* database, approximately 76% of the human genome is transcribed. Long ncrnas are arbitrarily defined as being longer than 200 nucleotides. Computational analysis suggests that 60% of protein-coding rnas can be targeted by mirnas: functional part. Typically produces subtle (<2-fold) reduction in protein levels: expression might be enough to cause difference in the protein works or functional outcome. Have been linked to regulation of developmental processes, fine tuning gene expression, disease progression and chemotherapeutic resistance. Mirnas can exist, and be transcribed, as a cluster. Most protein coding rna most of them have sequences within their untranslated region, that is a target for microrna. Many of the microrna may be able to target multiple genes.
