Summary Summery lectures Translational Genomics

Translational genomics
Genome architecture
DNA is present in two places in the cell±
1. DNA in the nucleus
2. DNA in the mitochondria

Functional DNA
- Protein coding
- Non coding
- Regulatory element
genes can be spliced different
- This can lead to different isoforms in different tissue




Small ncRNA: mechanism of action
Biogenesis of miRNAs
- transcribed by RNA polymerase II
as pri-miRNA
- processing by endoribonuclease
Drosha and double-stranded RNA-
binding protein DGCR8 (called
pasha in flies and nematodes).
o Drosha has two RNase III
domains that each cut one
strand of the stem of the
pri-miRNA with a 2bp
offset
o ~ 60nt stemloop called pre-miRNA
- Exporting out of the nucleus into the cytoplasm by exportin5
- Processing pre-miRNA by endoribonuclease Dicer
o Dicer has also two RNase III domains.
o Dicer cuts both strands near the loop
o Generating of a miRNA duplex (of ~ 22 nt)
 this duplex contains a ~2 nt 3’ overhang on each end.
 miRNA paired to its passenger strand (miRNA*)
o Loading of miRNA duplex in argonaute
 Passenger strand degradation
 5’ U or A is preferred
o Forming of the silencing complex (RISC) -> pairing of the mature miRNA to mRNA
sides
 Translational inhibition (translation initiation or elongation) and inducing degradation of
target RNA
Feingold syndrome 2: miR-17~92 deletion

,Long ncRNA

NAT: natural antisense transcript
- Can work cis and trans
o Cis: from the same
template
o Trans: other gene




Mechanism
a. Inhibition of transcription
b. Blocking of splicing by binding a specific exon
c. Recruitment of RNAi  inhibition of gene
expression

Regulatory elements
- Core promotor elements: surrounding the
transcriptional start site
(-40 - +40bp)
- Promotor proximal elements: upstream of the core promotor
(-200 - -40bp)
- Enhancer/silencer: further than 200bp upstream/downstream or even far away
 Different regulatory DNA elements in eukaryotes are differentiated by distance from start
site

Core promotor elements: minimal sequence required for transcription initiation
- TATA box
- Initiator element
- Downstream promotor Element (DPE)
Promotor-proximal elements: modulation of transcription
- Often gene- or cell type-specific
Enhancers/silencers: modulation of transcription
- Often gene- or cell type-specific
- Can work from a distance
- Orientation-independent

Regulatory regions are diverse in eukaryotes
TATA-box: classical (core promotor)
o Fulfils similar function as enhancers and promoter-proximal elements
The core promotor
Core promotor: consists of the combination of distinct sequence motifs
- TATA:
Conserved sequence called the TATA box found 26-31bp upstream the transcription start
site. Positions RNA pol II for transcription initiation
- Inr

, Initiator sequence; instead of TATA box some eukaryotic genes contain an alternative
promotor element called the initiator.
- BRE
- DPE

‘junk’ DNA
 Transposable elements (TEs): ‘jumping’ genes
o 45% of human genome
o <0.05% active
o The most abundant: Alu elements (10% genome)
Two classes
1. Class I: retrotransposons
a. LINEs
b. SINEs
c. LTR
2. Class II: DNA transposons

Example: family with dilated cardiomyopathy
They found that there was an exon spliced in
that normally isn’t there  Alu repeat

They found differential expression of DMD
isoforms
 They didn’t have the normal exon in
the cardiac muscle: possible
explaining the cardiac problems

Also, differential protein (dystrophin)
expression
 Really clearly in heart, no expression


Genomic imprinting
What is the most extreme example of the effect of epigenetic modification on gene expression?
- DNA methylation
DNA methylation  imprinting  inactivation

Paternal and maternal genomes contain the same set of genes  embryo from two paternal or two
maternal embryos cannot develop.
- Paternal and maternal genomes must be epigenetically modified
 Imprinting turns on or off certain genes in either sperm or egg  implying a memory of the
genes being in sperm or egg
o At least 80 imprinted genes identified in mammals
- Maternal and paternal genomes make different contributions to embryonic development
o Two maternal genomes: relatively well developed embryos; extra-embryonic tissue
poorly developed
o Two paternal genomes: well-developed extra-embryonic tissue; embryo itself is
abnormal

, Epigenetics:
- DNA methylation  gene repression
o Always metalation of the C in a CG  methylation of Cytosine by methyltransferase
to 5-Mehylcytosine
- Histone modifications  gene repression and/or activation
- Non-coding regulatory RNAs
o Attracting of polycomb group proteins  transcriptional repressors via chemical
modifications of histone proteins

Genomic imprinting = essential for normal development
 Deregulation results in complex genetic diseases

Example: Chr15q11-q13: same deletion causes to very different
syndromes
1. Angelman syndrome: maternal deletion  loss of maternal
gene expression = paternal uniparental disomy
a. ID, laugh a lot unexpectedly, ataxia, no speech,
epilepsy, typical face, friendly
2. Prader-Willi syndrome: Paternal deletion  loss of paternal
gene expression = maternal uniparental disomy
a. Neonatally: hypotonia, feeding probles
b. First decade of life: obesitas, small, mild ID, hypogonadism, behavioural problems