W13L3 Flashcards
Extension of Epigenetics
Understanding developmental disorders
- Intellectual Disabilities
Combining epigenetics with therapy for cancer
Research tools – iPSCs (induced pluripotent stem cells)
- Reprogramming somatic cells to become embryonic stem cells
- Factors required for reprogramming include Oct4, cMyc, Klf4 and Sox2
- These factors are called pioneer factors since they can promote loosening of the heterochromatin and allow access for other factors
- at level of epigenome, Genome is becoming more active or open for transcription
Non-coding (ncRNA)
Affect transcription, silence genomic regions or alter RNA processing all leading to changes in RNA accumulation and expression
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
long ncRNAs are arbitrarily defined as being longer than 200 nucleotides
- Encyclopedia of DNA Elements
Types of ncRNA
Transfer RNA (tRNA)
- Highly abundant with a well defined function
Ribosomal RNA (rRNA)
- Highly abundant with a well defined function
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
siRNA – small interfering RNA; promotes mRNA degradation
microRNA (miRNA) – RNA silencing and post-transcriptional regulation
lncRNA – long non-coding RNAs
circRNA – circular RNA, long non-coding RNAs
snoRNA – small nucleolar RNA
responsible for posttranscriptional modification and maturation of rRNAs, snRNAs, and other cellular RNAs
most snoRNAs are encoded in the introns of protein-coding or non-coding genes
Typically 60-170 nt long
can regulate mRNA splicing (alternative splicing)
primarily accumulate in the nucleoli
snRNA – small nuclear RNA
average size of 150 nt
exist with proteins in small ribonucleoprotein particles (snRNPs)
catalyze the splicing of pre-mRNA
allow precise alignment and correct excision of introns
found in Cajal bodies within the nucleus
piRNA – piwi-interacting RNA
approximately 24–31 nucleotides
bind to members of the piwi protein family
largest class of small non-coding RNA molecules expressed in animal cells
influence transposon silencing, spermiogenesis, genome rearrangement, epigenetic regulation, protein regulation, and germ stem-cell maintenance
exRNA – extracellular RNA
possibly cell communication
These can include miRNAs that lead to gene silencing in target cells
microRNAs
Hairpin derived RNAs ~20-24 nucleotides long
Typically bind the 3’ UTR of mRNA leading to repression
Computational analysis suggests that 60% of protein-coding RNAs can be targeted by miRNAs
Recent studies describe >5,600 miRNAs (http://www.mirbase.org/)
Typically produces subtle (<2-fold) reduction in protein levels
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
siRNAs
Originates from dsRNA and is usually 21-22 nucleotides long
Primary role in repression is to cleave mRNA
Often the response to viral infection and provide a perfect match to its target (therefore typically has a single target)
Often used as a research tool to knock down expression by as much as 85-90%
Unsure if siRNA is produced in mammalian cells
Dicer, argonaut, and RISC
DICER cleaves both miRNA and siRNA
- becomes ds RNA, which is recognized by RISC complex (RISC = RNA-induced silencing complex)
RISC binds ds RNA
- there is AGO=Argonaute specific for miRNA and siRNA
the RISC - AGO complex gets rid of 1/2 of the ds RNA to make it single stranded so it is complementary to whatever the target is
- for siRNA, passenger RNA is cleaved
- for miRNA, passenger RNA is discarded
OVERALL
Dicer:
- Cleaves double-stranded RNA molecules into short double stranded fragments of around 20 nucleotides
RISC:
- Integration of one strand into a RISC complex
- RISC then targets the complementary sequences in mRNA
- Argonaute is then activated and, depending on the isoform, can cause mRNA cleavage or translation inhibition
Long non-coding RNAs (lncRNAs)
Over 200 nucleotides in length; can reach up to several 100 thousand nucleotides in length
originally thought of as transcriptional noise of RNA polymerases with low fidelity
Almost 15,000 described transcripts in the human genome in 2012
Some lncRNAs have begun to be classified for functions
- Regulating expression of neighboring protein-coding genes
- chromatin modification, transcription and post-transcriptional processing
Barr body
First described by Dr. Murray Barr (Western Professor in Dept. of Anatomy and Cell Biology)
Represents the inactive X chromosome
The entire X chromosome is not inactivated
