LIF 101 16th October 2016

RECAP

Symbols used in pedigree analysis:

Male
Female
Mating
Parents &
Children:
1 boy; 1
girl
(in order
of birth)
Dizygotic
(non
identical
Twins)

Sex
unspecified
Affected
individuals
Heterozygotes
for autosomal
recessive
Carrier of sex
linked recessive
Death
Abortion or
still birth
Propositus

Monozygotic
(identical
Twins)

Consanguineous
marriage

RECAP

Pedigree Analysis

Normal female

Marriage

Normal male
One these
parent was
therefore a
carrier

1st born

II

Affected

Siblings

Is the mutant
allele dominant
or recessive?

RECAP

Albinism :
Autosomal recessive inheritance

Why this inheritance

is called autosomal?

RECAP

Achondroplasia:
Autosomal dominant inheritence

Why this called

autosomal dominant
dominant ?

RECAP

Sex Determination in humans

Parents

X-linked recessive disease

Female carrier* mates with normal male

RECAP

FxM

XNXA

Sperm

XNY

XN
Eggs
XA

XN

XNXN
normal

F
XAXN
carrier

XNY
normal

M
XAY
affected

Half* her daughters will be carriers

Half* her sons will be affected
*on average

Pedigree Illustrating X-Linked Recessive

Inheritance Pattern

Great grand mother

Grand mother
Nani
Grand Uncle

Uncle (mama)

Nephew

Grand
daughter

Hemophilia : X-linked recessive

Haemophilia A is a genetic deficiency
in clotting factor VIII which causes
increased bleeding and usually affects
males. About 70% of the time it is
inherited as an X-linked recessive trait,
but around 30% of cases arise from
spontaneous mutations.

Empress of India 1876

Queen Victoria (1819-1901)

passed haemophilia A
on to many of her descendants

Haemophilia leads to a severely

increased risk of prolonged
bleeding from common injuries, or
in severe cases bleeds may be
spontaneous and without obvious
cause.

Edward
(Duke of Kent)

Victoria
(Princess of Saxe-Coburg)
Victoria
(Queen of England)

Albert

Victoria

Alice

Frederick

Wihelm II Sophie

George V

George VI

Queen
Elizabeth

Louis

Alfred

Arthur
Helena Louise

Alix Nikolas II of Russia

Waldemar Henry

Prince Philip

Normal Male
Normal Female

Hemophilic Male

Carrier Female

Male died in infancy

Possible hemophilic

Alexic

Leopold

Helen

Henry

Beatrice

Alfonso XII Spain Eugenic

Alfonso

Gonzal
o

nheritance of eye coloration in fruit fly, Drosophila:

An example of sex-linked inheritance

Normal Red

Mutant white

Then test and infer

w+w+

w+w

w+w

ww

w+w

W+

12

Problem
TEST: A yellow body-colored female fruit fly was
mated to a male with normal body color (brown).
RESULT: All the female progeny were brown
(normal) while all the male progeny were yellow.

EXPLAIN: What does this inheritance pattern

signify?

Display this cross

Significance of chromosome numbers in in

heritance
What if individual chromosomes do not separate
during meiosis? Error in chromosome number

The Culprit: Non-disjunction

Older women are more likely than younger women to

evidence chromosome damage and meiotic
irregularities

Pre-natal Diagnosis: Amniocentesis

Will it be possible to
detect a defective
gene by
amniocentesis?

Fetal cells present in

amniotic fluid are then
cultured and analyzed for
chromosomes
(karyotyping) and other
biochemical tests

This baby has

one X
chromosome.
Yet it is a female
baby. Why?

Monosomy: Turners Syndrome: 44+X

Individual chromosome segregation going wrong during meiosis

How can errors

in chromosomal
segregation
result in Downs
syndrome?

Why downs
syndrome does
not suggest
abnormal sex
linked inheritance?

Trisomy of chromosome . 21: Downs

Syndrome: 45+XY/45+XX
Other disorders: Trisomy-13 (47:+13), Trisomy-18 (47:+18),
Klinefelter Syndrome (44+XXY), Triple X syndrome (44+XXX), XYY
trisomy (44+XYY)

People suffering from Down

syndrome can lead a
reasonably normal life

Genes and, chromosomes and some interesting

facts about our inheritance

These cats are known as Calico cats marked

by their patchy coat (fur) coloration . The
coloration gene (or fur pigmentation gene) is
present on the X chromosome (X-linked)

Coat coloration
in XB/Xb cats

B= orange allele
b= Black allele

Possible Female genotypes

Possible male genotypes

However, one of the X chromosomes is

randomly inactivated in female cat body
cells
Female cell where
X chromosome
carrying black
allele allele is
inactivated

Female cell where

X carrying orange
allele is
inactivated

Note the fur/coat coloration in male and female cats

Explain why
male cats
have
uniform coat
color

Explain why these female cats

have uniform coat color

Explain why these female

cats have patchy coat color

How will you name the

gene that regulate eye
coloration?

P1

How will you decide

which one is recessive
and which one is
dominant ?
What will be the consequence
if the while gene is X-linked
and Drosophila female and
male have XX and XY sex
chromosome constitution?

What types and proportions of F2

progeny are expected the white
gene were located on an
autosome?

Predict the phenotypic ratios in the progeny if a

Drosophila female, heterozygous for white mutation
is mated with a white eye-colored male ?
Do you think inheritance of white gene in
Drosophila follows Mendelian laws?
Do you think inheritance of white gene display a
chromosomal basis of inheritance? Which
chromosome?

Significance of chromosome numbers and sets

What if individual chromosomes do not
separate during meiosis? Error in
chromosome number
What if entire sets of chromosome do not
separate? Error in chromosome sets

Polyploidy in plants

Colchicine inhibits
chromosome
segregation

Polyploidy in plants
In nature 3n, 4n, 6n and 8n plants are very
common
Many varieties of wheat, cotton, banana,
tobacco , strawberries etc are polyploids
Polyploidy results in bigger grain or fruit sizes
and high yield which is beneficial
Thus, polyploidy is also artificially induced in
plants by breeders using colchicine.

Potato flowers. Potato is an

autotetraploid.

The cultivated strawberry is an octaploid

hybrid derived from hybridization of two
natural octaploids.

This supermarket banana is most likely the

common variety Cavendish, an autotriploid of
Musa acuminata
Seeds of bread wheat. Wheat is a
allohexaploid. Its genome consists of three
ancestral parental genomes

DNA as a genetic material

First historical demonstration of an chemical entity for genetic

inheritance

Griffiths experiment 1927:discovery of transforming

factor (TF))
Virulent
strain kills
the host

What does this heat killing signify with

respective genetic inheritance

Non-virulent
strain does
not kill
Virulent

Non-virulent

Dies

Heating kills
virulent
strain

Lives

Virulent, heat killed

Lives

Non-virulent + heat
killed
Pneumococcal infection

Dies

Heat-killed
virulent strain
can transfer its
virulence
factor to a
non-virulent
strain

What is genetic
transformation?

Which one was genetically

transformed? Mouse or the bacteria?

Avery, MacLeod, McCarty

Purified the transforming factor (TF) to
investigate its properties.
Salient points:
Treatment with protease did not destroy TF
Treatment with RNase did not destroy TF
Purified TF did not have sugars

Thus TF should be DNA

What about Mendelian
alleles? Are these too
transforming factors,
Mendelian factors?

Do these terms
transforming factors,
Mendelian factors and
DNA carry similar meaning
?

What is there in transforming or Mendelian

factor? And now in DNA?
DNA carries digitized information
Four digits are used to write genetic
codes

Watson and Crick proposed structure of DNA

1953

Journal Nature in its 171st volume on pages 737738 ( 25 April 1953)

Thus, to understand the nature of

genetic information we need to
understand the basic DNA structure and
how information flows:
Within an individual.
From one generation to the other
Within a cell

DNA molecule

RECAP

DNA is a very large molecule made up of a long chain of

sub-units
The sub-units are called nucleotides
Each nucleotide is made up of

a sugar called deoxyribose

a phosphate group -PO4 and
an organic base

A few general features you need to

remember about DNA structure

Chargaffs Rule RECAP

Adenine must pair with Thymine
Guanine must pair with Cytosine
Their amounts in a given DNA molecule will be
about the same.
same

DNA Double Helix

RECAP

H-bonds
3

P
5

O
O

4
2

H-bonds

DNA strands run anti-parellel

RECAP
Mark the features of
DNA that makes it
ideal as a genetic
materials

Sequence of
nucleotides in
DNA strands
could store
genetic
information

Sequence in one
strand serves as the
template for the
sequence in the
other

Central dogma of Molecular

biology

Nature of
information
flow
Flow of information:
nucleic acid to nucleic
acid

Flow of
information
Nucleic acid to
protein

Replication
Transcription

RNA

DNA

Why there is no
loss of
information
during
replication?

Translation

Reverse
transcription

How much of
the information
in DNA goes to
RNA??

protein

In subsequent lectures we
will explore this flow of
information the decoding
process

Flow of genetic information

Replication: synthesis of daughter DNA from parental DNA

Transcription: synthesis of RNA using DNA as the template
Translation: protein synthesis using mRNA molecules as the
template
Reverse transcription: synthesis of DNA using RNA as the template

Test your understanding

Question

Which aspect of
information flow does
these sister chromatids
symbolize ?

How many DNA molecules are present in a

metaphase chromosome?
How many DNA molecules are present in a
chromatid?
Are the DNA sequences of two sister
chromatids are identical?
How many DNA molecules are there in a human
sperm?
Which aspect of
information flow a
sperm represent ?

How many chapters are there in our book of life?? And how
many words/sentences are there in each chapter??
Here book of life is a
metaphor

RECAP

Does a cell read

these chapters one
by one - in a
sequence?

What is implied when

one says that genetic
information is being
readout?

Why did the author say

that human genome is
written in 23 chapters?
Why not 46?
What is a gene/
Mendelian factor:
a letter,
a word or
a sentence?
AUTOBIOGRAPHY OF A
SPECIES IN 23 CHAPTERS

Approximate gene numbers

Chromosome 1 : 4220
Chromosome 2 : 1,491
Chromosome 3 : 1,550

Next lecture:
We would examine the nature of genetic
information and how it flows within a cell

Slide series 6

Flow of genetic information:

Three aspects Transcription, Translation
&
Genetic code

Lets first appreciate the challenge of

transfer of information from DNA
(genes) to protein

Consider the case of Sickle Cell Anemia?

A serious condition in which red blood cells

can become sickle-shaped
Normal red blood cells are smooth and
round. They move easily through blood
vessels to carry oxygen to all parts of the
body.
Sickle-shaped cells dont move easily
through blood. Theyre stiff and sticky and
tend to form clumps and get stuck in blood
vessels.
The clumps of sickle cell block blood flow in
the blood vessels that lead to the limbs and
organs. Blocked blood vessel can cause
pain, serious infection, and organ damage.
While the defect is seen in the protein,
sickle cell anemia is produced by changes in
gene sequence

Normal and Sickled Red Blood Cells in Blood Vessel

Figure B shows abnormal, sickled red blood cells clumping
and blocking the blood flow in a blood vessel. The inset
image shows a cross-section of a sickled red blood cell with
abnormal strands of hemoglobin.

Figure A shows normal red blood cells flowing freely in a

blood vessel. The inset image shows a cross-section of a
normal red blood cell with normal hemoglobin.
Source from http://www.nhlbi.nih.gov/health/dci/Diseases/Sca/SCA_WhatIs.html

Questions?
What could be the reason(s) for the wrong
message from the DNA that produced a
wrong protein (hemoglobin)?
How information encoded in the DNA of
the nucleus is transferred to the cytoplasm
How does this information translate into
protein??

Central dogma of Molecular biology

RECAP

Nature of information flow

Flow of information:
nucleic acid to nucleic
acid

Flow of
information
Nucleic acid to
protein

Replication
Transcription

RNA

DNA

Why there is no
loss of
information
during
replication?

Translation

Reverse
transcription

How much of
the information
in DNA goes to
RNA??

protein

We will now explore this

flow of information: that is,
process of decoding the
genetic message

Flow of genetic information

A gene is transcribed into RNA
which is then translated into the polypeptide

DNA
Transcription

RNA

Translation
Protein

FLOW OF GENETIC INFORMATION FROM DNA

TO RNA TO PROTEIN
Gene (DNA) is a linear sequence of many
nucleotides

DNA is transcribed into a linear sequence

of RNA
RNA is translated into a linear sequence of
amino acids polypeptides (protein)
DNA
Entire business of flow of
genetic information thus
depends on constructing
(transferring information)
one kind of polymer from
another

RNA

Protein
(polypeptide chain

Transcription Step I

T G C A T A G C G C A T
Template DNA Strands unzip locally

Transcription Step II

A C G T A T C G C G T A

DNA

U G C A U A G C G C A U

mRNA

One of the DNA strands serve as a

template and copied as mRNA
(messenger RNA)

A C G T A T C G C G T A
DNA

T G C A T A G C G C A T

RNA U G C A U A G C G C A U
mRNA is then releases

Notice the difference between ribose and

deoxyribose in the figure above. In ribose, carbon
atom #2 carries one hydroxyl group (colored red).
In deoxyribose, carbon atom #2 carries a hyrogen
atom instead of a hydroxyl group.

Chemical structure of RNA

In place of
thymine
in DNA

Cytosin
e
DNA/RNA

Thymin
e
DNA

Pyrimidine bases

Urac
il
RNA

Transcription

RNA nucleotides

RNA polymerase

T C C A A T

T A G G T T A

Direction of
transcription

Newly made
RNA

C A U C C A

T
G

Template
Strand of DNA

 In the nucleus, the DNA helix unzips

And RNA nucleotides line up along
one strand of the DNA, following the
base pairing rules
As the single-stranded messenger
RNA (mRNA) peels away from the
gene
The DNA strands rejoin

Steps in transcription of a gene

RNA polymerase

DNA of gene
Promoter
DNA

Terminator
DNA

1 Initiation

Area shown
In Figure 10.9A

2 Elongation

Growing
RNA

3 Termination

Completed RNA

RNA
polymerase

How does DNA/RNA code for protein?

Strand to be transcribed

DNA

Codon a sequence of
nucleotides that
codes for an
amino acid

Transcription

RNA
Start
condon

Polypeptide Met

Translation

Lys

Stop
condon

Phe

STEP III TRANSLATION

How can four

nucleotides code for 20
amino acids?
If one nucleotide codes for one amino acid then a
maximum of four amino acid could be coded
If two nucleotide code for one amino acid then a
maximum of 16 amino acid could be coded
If three nucleotides code for one amino acid then
a maximum of 64 codons can be generated, while
the number of amino acids are only 20

The dictionary of genetic code (64 potential

codons)

5-3
A
dictionary
offers
translation
from one
language
to the
other. two
Which
languages
are being
translated
in a
genetic
dictionary?

Why do
we need
this
translation
?
Where
does it
take
place?
What is
the name
of this
process?

Hallmarks of genetic code

Triplet : three consecutive nucleotides code of one amino

acid
Redundant : meaning not unique (there are
more than one codon for most amino acids)
Universal: All organisms bacteria to human use
the same genetic codes/dictionary
Comma-less: meaning consecutive codons are
readout into an amino acid sequence without
gaps

How does an universal genetic code impact Biotechnology?

What would have

been the case if
bacterial genetic
code were to be
different from
human?

What do we
understand when we
mean when we say
that genetic code is
universal?

Key players in decoding the genetic message

DNA

Messenger-RNA

Transfer RNA
Amino
acid

Transfer RNA

Leucine

Anti-codon

Codon

How genetic code is readout in the cell

A C G T A T C G C G T A

DNA

T G C A T A G C G C A T
3

U G C A U A G C G C A U

mRNA

Amino
acid

Anti-Codon

5
tRNA

C A G
3

G U C
Codon

Many amino
acids are
specified by
more than one
codondegeneracy
Codons
specifying the
same amino acid
areConnect
calledthe three
points about genetic
synonyms
codes:
redundancy,
degeneracy and
synonymous

Codon-anticodon pairing of two tRNA Leu

molecules

5- 3

5- 3

CUG

Codon

GAC

Anti- codon

3- 5

CUC
GAG
3- 5

Three codons direct chain termination

Three codons, UAA, UAG, and UGA signify

chain termination.
They are not read by tRNAs but by
proteins called release factors (RF1 and
RF2 in bacteria and eRF1 in eukaryotes).

Three Rules

Codons are read in a 5 to 3 direction.

Codons are non-overlapping and the
message contains no gaps.
The message is translated in a fixed
reading frame which is set by the
initiation codon.

Meaning and consequences of non-overlapping & comma-less reading

of the triplet codons
Lets consider a
metaphor of a normal
DNA sequence

FATCATATETHERAT
What is
the
problem
here?
What is
the
consequen
ce?

FATATATETHERAT

Mutation: a change in genetic code

1. Missense mutation: An alternation that
changes a codon specific for one amino acid
to a codon specific for another amino acid.
2. Nonsense or stop mutation: An alternation
causing a change to a chain-termination
codon.
3. Frame-shift mutation: Insertions or deletions
of one or a small number of base pairs that
alter the reading frame.

How the genetic code was cracked?

The use of artificial mRNAs and the availability

of cell-free systems for carrying out protein
synthesis began to make it possible to crack the
code

Codon Assignments from Repeating Copolymers

Organic chemical and

enzymatic techniques were
used to prepare synthetic polyribonucleotides with known
repeating sequences.

Experimental Results:

UUU codes for phenylalanine.

CCC codes for proline.
AAA codes for lysine.
The guanine residues in poly-G firmly
hydrogen bond to each other and form
multistranded triple helices that do not bind
to ribosomes.

polymer
(CU)
3

Codons
Recognized

Amino Acids
Codon
Incorporated Assignment

CUC|UCU|CUC

Leucine

5-CUC-

Serine
UCU
(UG)

UGU|GUG|UGU

Cystine

UGU

Valine
GUG
(AC)

ACA|CAC|ACA

Threonine

ACA

Histidine
CAC
(AG)

AGA|GAG|AGA

Arginine

AGA

What Are Mutations?

Changes in the nucleotide sequence of
DNA
May occur in somatic cells (arent passed
to offspring), but could cause diseases,
such as cancer
May occur in gametes (eggs & sperm)
and be passed to offspring

Nature of point mutations

1. Missense mutation: An alternation that
changes a codon specific for one amino acid
to a codon specific for another amino acid.
2. Nonsense or stop mutation: An alternation
causing a change to a chain-termination
codon.
3. Frameshift mutation: Insertions or deletions
of one or a small number of base pairs that
alter the reading frame.

Point Mutation
Change of a single nucleotide
Includes the deletion,
insertion, or substitution of
ONE nucleotide in a gene

Types of Gene/Point
Mutations
Substitutions
Insertions
Deletions
Frameshift

Frameshift Mutation
Inserting or deleting one or
more nucleotides
Changes the reading
frame like changing a
sentence
Proteins built incorrectly

Gene Mutation
Animation

Problems

Delete C

Insert A

THE

FAT

CAT

THE

FAT

ATA

THE

FAT

ATA

ATE
TET
ATE

THE

BIG

RAT

HEB

IGR

AT

THE

Frame shift mutation

BIG RAT

RBC

Structure
of Hb

Primary sequence of Hb first 26 amino acid

RECALL

Example Point
Mutation
Sickle Cell disease is
the result of one
nucleotide substitution
Occurs in the
hemoglobin gene

Problem
Before the true nature of the genetic
coding process was fully understood, it
was proposed that the message might be
read in overlapping triplets. For example,
the sequence of GCAUC might be read as
GCA CAU AUC

G C A U C
Think of test that would negate this possibility

Consider a gene that specified the structure of hemoglobin.

Arrange the following events in the most likely sequence in
which they would take place
a. Anemia is observed
b. The shape of the oxygen binding site is altered
c. An incorrect codon is transcribed into hemoglobin
mRNA
d. The ovum (female gamete) receive a high radiation dose
e. An incorrect codon is generated in the DNA of the hemoglobin gene
f. A mother (an X-ray technician) accidentally steps in front of an
operating X-ray generator
g. A child dies
h. The oxygen-transport capacity of the body is severely impaired
i. A wrong tRNA is
attached
j. Nucleotide pair substitution occurs in the DNA of the gene for
hemoglobin

Question

If an mRNA of human betaglobin gene is added to

bacterial cell free extract then
human beta globin polypeptide
is formed (translated)
What does this observation signify
with respect to genetic code?

Home assignment
The amino acid sequence shown in the following table was
obtained from the central region of a particular polypeptide
chain in the wild type and several mutant bacterial strains
Codon
1 2 3 4 5 6 7
8
9
a. Wild type phe leu pro thr val thr thr arg trp
b. Mutant 1 phe leu his his gly asp asp thr val
c. Mutant 2
phe leu pro thr met thr thr arg trp
d. Mutant 3

phe leu pro thr val thr thr arg

e. Mutant 4
f. Wild type

phe pro pro arg

phe leu pro ser val thr thr arg trp

For each mutant, say what change has occurred at the

DNA level, whether the change is a base pair
substitution or frame shift mutation, and in which
codon the mutation occurred

Home assignment
In mutant strain X of E. coli, a leucine tRNA that
recognizes the codon 5-CUG-3 in normal cell has been
so altered that is now recognizes the codon 5-GUG-3.
A missense mutation (lets call this mutation Y) that
affects amino acid 10 of a particular protein is
suppressed in mutant X cells (that is when the cells are
mutant for both X and Y, which now appears wild type)

a. What mutational even has occurred in mutant X

cells?
b. What amino acid would normally be present at
position 10 of the protein (without the missense
mutation)?
c. What amino acid is put in at position 10 if the
missense mutations is not suppressed ?
d. What amino acid is put in at position 10 if the
missense mutations is suppressed ?

Slide series 7

Gene regulation
- Prokaryote
- Eukaryotes

Gene regulation

Regulation of the synthesis of a

genes transcript and its protein
product is termed as gene regulation.

Why control gene expression?

Each of our cells carry entire genetic
instructions for our growth, development
and metabolic functions.
Some of these genes are needed to be
expressed all the time: respiration, for
instance. These are also called house
keeping genes
Other genes are not expressed all the time.
These are switched on and off on
demand

Promoter:
RNA Pol binding site in
DNA sequence
upstream of transcription
start point
RNA
polymerase

Promoter

Basic scheme

Gene
Direction of transcription

Transcriptional Regulation is mediated by

controlling the access of the RNA polymerase to
the promoter.

The promoters of genes transcribed by RNA polymerase II consist of

a core promoter and a regulatory promoter that contain
consensus sequences.
Not all the consensus sequences shown are found in all promoters.

We will not go in to these

details either

A point to remember:
Like transcription
initiation there are also
transcription termination
mechanisms

Negative Regulation

Positive Regulation
RNA Polymerase

RNA Polymerase

Binding of
repressor
blocks the
binding of the
RNA Pol to the
promoter

Repressor

X
Activator

An activator
help RNA Pol
bind to the
promoter

X
Two broad strategies for regulation of prokaryotic transcription

In prokaryotes gene regulation takes

place in response to the environment
Glucose

E. coli can use either glucose, which

is a monosaccharide, or lactose,
which is a disaccharide

However, lactose needs to be

hydrolysed (digested) first

So the bacterium prefers to use

glucose when it is available.

On the other hand, when only lactose

is available it turns ON the genes
and therefore the enzymes required
for lactose breakdown

Lactose

Glucose

Galactose

Regulation of the lac Z gene an example of negative regulation

( lacZ gene codes for -galactosidase enzyme that breaks down
lactose )

What can
be the
inducer of
the lac Z
gene?

Answer:
lactose

REPRESSO
R
INDUCE
R
INACTIVE
REPRESSO
R

OPERATOR
:

Binding
site for
When lactose
is absent in the medium, the lac Z gene is
repressor
switched-off . That is, no mRNA is transcribed and no proteins
are made.
-In the presence of lactose in the medium, lac Z gene is
turned on. That is, mRNA is transcribed and the proteins are

Question:1
Repressor is a
protein: should
there be a
separate gene
for repressor?

INDUCIBLE
TRANSCRIPTION

Illustration of a NEGATIVE
REGULATION gene expression

Question:2
How will lac Z gene
be regulated if a
gene coding for
repressor is
mutated
Question:3
What will be the
consequence if
operator sequence is
altered?
Question:4
What will happen if
promoter sequence is
altered

Regulation of lac Z
gene

This is also called

an inducible
model of gene
regulation. WHY?

RNA POl
REPRESSOR
ACTIVE
INDUCER

REPRESSOR
INACTIVE
Repressor binding
site
OPERATOR

Concept
Mutations need always directly alter the target gene. There are
alternative ways of altering gene expression, other than
mutations within the gene

Further, lac Z gene is co-regulated along

with two other genes called lac A and lac Y

A set of gene co-regulated under one promoter are

called operon. Thus, an operon represents a group
of genes that are transcribed at the same time.
They usually control an important biochemical
process.
They are only found in prokaryotes.

The lac Operon

P

mRNA

lacZ

lacY

How many
messenger
RNA are made
by the lac
operon?

lacA

RIBSOSOME BINDING SITE

The lac Operon

P

mRNA

lacZ

lacY

lacA

Proteins
b-galactosidase

Permease

Why would you consider the lac operon a

smart system?

Transacetylase
How many
polypeptide
(proteins) are made
by the lac operon?

Regulation of the lac Operon by the

repressor
P

lacZ

lacY

NO mRNA

lactose

lacZ

lacY

lacA
Note that there are
two binding sites in
the repressor one
for the operator
and the other for
the inducer

lacA

NO mRNA

In the presence of lactose in the medium, lactose binds to the

repressor. The lactose-repressor complex is unable to bind to the
operator.

Regulation of the lac Operon by the repressor

lacZ

lacY

lacA

mRNA

Once Inducer-Repressor complex moves away from

the Operator (O) RNA polymerase can bind to the
promoter (P) and initiate transcription of the lac Operon

About the lac I gene that codes for the

repressor

lacZ

lacY

X
lac
repressor

lacA

NO mRNA
lac repressor

The lac repressor binds to the operator and

inhibits transcription of the lac operon.

Question:
Should the lac I gene be
always active (constitutive)
or sometimes active
(inducible)?

Question
Lac repressor has two binding sites: one
for binding with Operator and the other
for the Inducer. What will be the
consequence if a mutation alters its
inducer binding site

Eukaryotic Gene
Regulation:

Most eukaryotic genes are controlled individually

and have regulatory sequences that are much
more complex than those of the lac operon.

Why is gene regulation in eukaryotes more

complex than in prokaryotes?

All of the cells in a multi-cellular organism carry the complete

genetic code in their nucleus, but only a few of the available genes
can be expressed in the appropriate cells of different tissues.
Complex regulation allows for this specificity.

Gene regulation determined the

complexities of the eukaryotic
organisms

Increasing complexity of higher order structure

chromosome
chromatin

Eukaryotic DNA bind

with a variety of
protein.
DNA and a group of
proteins called
histones form the
primary chromatin
structure.
Chromatin in turn
complexes with a
large array of nonhistone proteins

Region upstream of the transcription

start site in higher eukaryotes

mRNA
GGGCGG
-200 bp

CCAAT
-100 bp

Promoter proximal elements

TATA
-30 bp

Promoter

Regulatory elements and promoter of an eukaryotic gene

Regulatory protein
Promoter
Promoter Transcribed region of a gene

DNA
Regulatory
elements
Promoter proximal elements

RNA polymerase

RNA transcript

Relative transcription
level

Functional role of the

regulatory elements and
promoter

Question:
Can changes
in the
sequence of
regulatory
sequences
too cause
gene
mutation?

The effect of specific point mutations in the

proximal regulatory elements and promoter on globin gene transcription. Each line represents
the transcription level of in a mutant relative to
that of a wild-type -globin gene. Red arrows
shows some the mutations that affect gene
transcription. The black dots represent nucleotides

Applications of the understanding

of the gene regulation

What if lacZ gene is fused with

the promoter of another gene?
A different promoter

lacZ

Background information
Genes expression is regulated during development the
colored stripes represent the areas where a certain gene
(named even-skipped) is expressed . Note a total of total
seven stripes

Normal 7 stripes of
even-skipped gene in
Drosophila embryo

Now a bacterial lacZ gene

fused to only stripe 2 module of
even skipped regulatory
element and injected back into
the fly embryo
What does
this finding
reveal?

Even skipped regulatory region

Stripe 2 module

Regulatory elements of a gene determine its expression in

specific locations

Promoter function seen in transgenic mice.

Recombinant
plasmid containing
rat growth hormone
fused to mouse
metallothionein
regulatory region in
a bacterial plasmid
vector

Mt-1 (metallothionein) gene is induced by heavy metal

This recombinant
plasmid,was injected
into the mouse
oocytes.

Key conclusions:
Genes regulate functionsGH Growth
Promoters drive gene
expression
Promoter of each gene
each gene is uniquely
regulated

Which one is
the mother?

A mouse derived from the eggs injected with

growth hormone gene (left) and a normal
littermate
(right).
The
expression
of a transgene' mostly depends on
the regulatory sequences which have been used in
the design of the gene construct. A classical
example is the transgenic giant mice in which the
expression of a growth hormone (GH) gene is driven