Download Genes

Gene Circuits -2
Shu-Ping Lin, Ph.D.
Institute of Biomedical Engineering
E-mail: [email protected]
Website: http://web.nchu.edu.tw/pweb/users/splin/
Date: 10.20.2010
Genes

Flow of information in gene expression from genes in the form
of DNA to mRNA by transcription and from mRNA to proteins in
the form of translation.
TRANSCRIPTION
TRANSLATION
DNA
mRNA
Ribosome
(a) Prokaryotic cell. In a cell lacking a nucleus, mRNA
produced by transcription is immediately translated into proteins
without additional processing.
Polypeptide
Nuclear
envelope
TRANSCRIPTION
DNA
RNA PROCESSING
Pre-mRNA
mRNA
Ribosome
TRANSLATION
Polypeptide
(b) Eukaryotic cell. The nucleus provides a separate
compartment for transcription. The original RNA
transcript, called pre-mRNA, is processed in various
ways before leaving the nucleus as mRNA.
Genes Are Found on Both Strands
In eukaryotes, coding regions (exons) are separated noncoding sequences
(introns).
Exons, introns and regulatory sequences of a gene all lie on the same strand
and the start and stop codons are at the 5’ and 3’-end of the strand respectively.
TRANSCRIPTION
RNA PROCESSING
DNA
Pre-mRNA
5 Exon Intron
5 Cap
30
31
1
Intron
Exon
Exon
3
Poly-A tail
104
105
146
Pre-mRNA
Coding
segment
mRNA
Ribosome
Introns cut out and
exons spliced together
TRANSLATION
Polypeptide
mRNA
5 Cap
1
5 UTR
Poly-A tail
146
3 UTR
Gene Anatomy
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Regulation of gene expression is mostly controlled at
transcription. But some level of regulation also occurs post
transcription and translation.
Prokaryotic Genes: Bacterial DNA contains mostly
uninterrupted protein-coding regions in which genes clustered
on DNA subunits called Operons.
Cluster of Genes lie on the same DNA strand that encode
proteins with related functions all controlled by a single
Regulatory circuit.
Operon consists of promoter (P) and operator (O), regulatory
sequences, coding regions of each gene and small spacers
between genes.
* Lac Operon: (Digestion of the
milk sugar lactose, such as
gene lacZ, lacY, lacA)
Lac Operon
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Digestion of milk sugar lactose: lac Z, lac Y, and lac A
Lac Z: contains 3075 bases and encodes β-galactosidase, which splits the
disaccharide lactose into monosaccharides glucose and galactose
Lac Y: 1254 bases and called lactose permease to encode transmembrane protein by
using energy of electrochemical gradient across membrane to pump lactose into cell
Lac A: 612 b (shortest) and encodes thiogalactoside transacetylase, which degraded
small carbon compounds
RNA polymerase (RNAp): protein complex, low affinity for DNA but in promoter
regions, binds to promoter for initiating transcription
Lac I: lac repressor (repressor protein), binds to operator region and then blocks
interaction of RNAp with gene-regulatory segment and prevents gene transcription
Catabolic gene activating protein (CAP): activator protein (DNA binding
protein), binds to promoter to attract/stimulate RNAp to the lac operon and enhance
transcription rate
Eukaryotic Genes
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Distribution of eukaryotic genes differs from prokaryotic genes
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Transcription factor (TF): conserved, regulate transcription by binding regulatory sites
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Only a fraction of eukaryotic DNA codes for proteins or RNA molecules: 1.1% of
human genome represents protein-coding genes
Coding regions are not continuous ( Split genes), contains exons and introns. Introns
often account for most of the gene size.: BRCA-1 (Chr 17) 100,000 bp. Codes for a
protein of 1863 aa rest is introns (~98%). BRCA-2 (Chr 13) 200,000 bp. Codes for a protein
of 3418 aa.
Eukaryotic genes may have multiple sequences regulating the same gene. Promoter
is the regulatory element closest to the first exon. Regulator sites distant from the first exon
are called enhancers. Some of these sequences may be as far as 50,000 bp upstream.
General TF: many are not specific to a given gene, but function as regulatory proteins for multiple genes
Specific TF: regulate cell-signaling pathways, exposing cell to external stimulus can increase synthesis
or activation of these TF
TFIID binding to TATA box at promoter and concluding with activation of RNA
polymerase II (RNApII) Initiate transcription
Start codon: placed a few base
pairs after transcription
initiation site
Stop codon (poly A sequence): in
last exon
DNA Looping
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Both exons and introns are transcribed into premature mRNA. 
Introns are excised and exons are brought together before mRNA
leaves nucleus and enters cytoplasm for translation.
Activator proteins bound to enhancer transiently bind to RNApII by
looping out intervening DNA.
Folding DNA enables proteins bound to distant regulatory sites to
physically interact with proteins bond to proximal sites.
DNA looping might enable
transcription factors bound to
enhancers to physically interact
with the components of the
transcription initiation complex
assembled at the promoter.
Activators: TFs bind to
enhancers, opening up DNA at
transcription-initiation sites
Human Chromosomes
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Protein-coding genes: linearly arranged on the chromosomes of
multicellular organisms
Approximate locations of thousands of genes on human
chromosomes have been identified.  Light (genes are
distributed, high GC concentrations ) and dark bands
Provide significant
information about the
function of genes and
progression of certain
diseases such as cancer 
Gene mutations cause
cancer occur in celldivision-cycle-regulating
genes
Gene mapping: closely
related organisms
Gene Circuits
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Gene expression can be explained through mathematical
models based on Boolean networks
Regulation of Gene
Expression in Prokaryotes
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E.coli genome decoded 101 - 105 copies of proteins depending on the
need Transcription rate can vary 1000 fold
Regulation of (milk sugar) lactose catabolism:
Glucose
No lactose
b-gal
No allolactose
Glucose
Lactose
b -gal
Allolactose binds LacR
Allolactose
High Affinity Regulation
of Lac Operon
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Multiple logic network
Involves
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Catabolic gene activating Protein (CAP): stimulate transcription only
when form complex with cAMP
cAMP signaling molecule (hunger signal): indicate absence of glucose
and lead to synthesis of enzymes
CAP + cAMP
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RNAp
Lac Operon also involves feedback control
Regulation of Lac Operon
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Prokaryotic cell structure is simple allowing for rapid movement
of ligands affecting gene expression.
Lac repressor (R) bound to the operator unless it forms a
complex with allolactose (A) in a glucose-poor and lactose-rich
environment  Blocks transcription
Lac Z: contains 3075 bases and encodes β-galactosidase, which
splits the disaccharide lactose into monosaccharides glucose
and galactose
RNAp binds to operator
sites and initiates
transcription only when
CAP protein (form a
complex with second
messanger cAMP) binds
to promoter
Regulatory Logic of
Eukaryotic Genes
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Multiple regulatory switches encode gene-specific transcription
factors  Include regulatory sites: (A) proximal; (B) distal enhancer
and (C) distal repressor
Module A, 3 regulatory sites, 2 of which, when occupied with TAs,
amplify and relay signals arriving from distal modules to basal
promoter (BP) of gene
Transcription is
represses when both C
and A1 are occupied
with their specific TFs.
A2 occupied  Acts as a
switch to B  Amply
signal coming from B
and relay to BP
A3 dominates signal for
transcription.
Time Course of
Transcription/Regulation
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Prokaryotic cell structure is simple allowing for rapid movement
of ligands affecting gene expression.  Within seconds
Eukaryotic gene transcription depends on the concentrations of
TFs in the nucleus. H form complex with Rab at promoter Pab
1. Polymerase transcribes gene “a”  Produce protein A
2. Enough A binds to Rc  Produces C
3. Sufficient C binds to Rb  Produces B
Bionetworks
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Proteins and gene regulatory elements that also include
metabolic pathways
Synthesis of aromatic amino acids - Phe, Tyr, Trp By plants and
bacteria (not in humans)
A  B, B  C, C  D, D  F
B  E, E  E’, E’  F, F  G
DNA
molecule
Gene 2
Gene 1
Gene 3
DNA strand 3
A C C A A A C C G A G T
(template)
5
TRANSCRIPTION
mRNA
5
U G G U U U G G C U C A
3
Codon
TRANSLATION
Protein
Trp
Amino acid
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DNA:
Phe
3’
Gly
Ser
,
, ,
,
,
,
,
,
, , ,
|
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accgtgattcatcccgccaacaaccgctattatac
,
,
, , ,
,
,
,
,
, ,
, 5’
1. mRNA 5’ UGG,CAC,UAA,GUA,GGG,CGG,UUG,UUG,GCG,AUA,AUA,UG 3’
Amino-acid sequence
WH*VGRLLAII
2. mRNA 5’ U,GGC,ACU,AAG,UAG,GGC,GGU,UGU,UGG,CGA,UAA,UAU,G 3’
Amino-acid sequence
GTK*GGCWR*Y
3. mRNA 5’ UG,GCA,CUA,AGU,AGG,GCG,GUU,GUU,GGC,GAU,AAU,AUG 3’
Amino-acid sequence
ALSRAVVGDNM