Download MBBS Biochemistry PPT 28 Regulation Of Gene Expression Lecture Notes

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Regulation of Gene Expression

1

Case report

? A 10 year old boy presented with signs and

symptoms and subsequent laboratory

investigation reports suggestive of Acute

lymphoblastic leukaemia. Patient was

receiving Methotrexate as a part of this

treatment. After several weeks of

chemotherapy with methotrexate, the boy

showed signs of resistance to treatment. What

could be the most likely mechanisms to

explain this resistance to methotrexate?

2
Objective

? What is Gene expression

? Necessity of regulation of gene expression

? Different types of gene regulation

? Gene regulation in prokaryotes

? Lac operon

? Tryptophan operon

? Gene regulation in Eukaryotes

? At transcription level

? At translation

? Modification of DNA

? DNA regulatory proteins

3

? What is gene expression?

? Formation of gene products

? RNA and Protein

? Necessity of Gene Expression and regulation

? Normal growth , development and functioning of

an organism

? Production of gene product according to the

requirement of the body

4
Type of gene expression

? Constitutive

? Occur all the time and at a relatively constant rate

? Regulated

? Transcribed only when needed

5

Regulation of gene expression

? Positive regulation---- Enhancer/Activator

? Negative regulation----Repressor/ Silencer

6
Temporal responses to regulatory signal

Type A

observed in prokaryotes in response to sudden changes of

the intracellular concentration of a nutrient

in higher organisms after exposure to inducers such as

hormones, nutrients, or growth factors

7

Type B

occurs during development of an organism, when only the

transient appearance of a specific gene product is required

8
TypeC

Occurs during the development of differentiated function

in a tissue or organ

9

Stages of regulation of gene

expression

? 1. Transcription
? 2. Post transcriptional modification
? 3. Translation
? 4. Post translational modification
? 5. RNA degradation
? 6. Protein degradation

10
Few specialized genetic and regulatory terms

Operon groups of genes sequentially arranged

on the chromosome along with the regulatory

elements that determine their transcription

Polycistronic mRNA

Constitutive /House keeping gene

Expressed at a constant level

Inducible gene/Regulated egne

expressed only under certain condition

Gratuitous inducer: Acts as inducer but not as substrate

An example is isopropylthiogalactoside (IPTG)

11

Cis acting

elements and

Trans acting

molecules

12
Regulation of Prokaryotic Gene

Expression

? Transcription is the primary site of regulation.

13

Role of Promoters

14
Prokaryotic promoters share two regions of highly

conserved nucleotide sequence

Promoter: A regulatory region o DNA that serves to bind RNA polymerase

I that in turn binds other substances that wil lead to initiation of transcription

15

Adapted from Harper's Biochemistry

Operon Model

Described by Jacob and Monod in 1961

Hypothesis: Based on the regulation of lactose metabolism by the

intestinal bacterium E coli

Led to the discovery of basic principles of Gene

transcription activation and Repression

16
Type of Operon

? Catabolic Operon

? Regulates the gene expression whose product have a

catabolic role

? e,g. Lac operon

? Anabolic Operon

? Genes of anabolic pathway such as synthesis of amino

acids are coordinately regulated

? e,g. Tryptophan operon

17

Lac Operon

18
LacI gene encoding repressor

CRE: cAMP response element

Regulatory region : Promoter : RNAP binds

Operator: Repressor binds

Structural genes:

lac Z: Galactosidase

lac Y: Permease

lac A:Thiogalactoside transacetylase

19

? The operator locus is a region of double-stranded DNA that

exhibits a twofold rotational symmetry and an inverted

palindrome (indicated by arrows about the

dotted axis) in a region that is 21 bp long, as shown below:

The binding occurs mostly in the major groove

The operator locus is between the promoter site and the

transcription initiation site of the lacZ gene

20
21

22
Lactose acts both as inducer and substrate

23

Tryptophan Operon

? Trp operon contains 5 structural genes

? Trp operon is subject to negative control

? Trp facilitates the binding of the repressor to the

operator

? Trp is a corepressor

? Trp operon also regulated by attenuation

? Transcription initiated but terminated wel before

completion

Transcriptional attenuation occur in Prokaryotes

but not in Eukaryotes

24
Attenuation of transcription of the trp

operon when tryptophan is plentiful.

25

Coordination of transcription and

translation in prokaryotes

? 1. Stringent response

? . 2. Regulatory ribosomal proteins

26
Regulation of

transcription by the

stringent response

to amino acid

starvation

27

Regulation of translation by an

excess of ribosomal proteins.

28
MCQ

? Which of the following mutations is most

likely to result in reduced expression of the lac

operon?

? A. i? (no repressor protein made)
? B. Oc (operator cannot bind repressor protein)
? C. Cya? (no adenylyl cyclase made)
? D. Functionally impaired glucose transporter

29

Regulation of Eukaryotic Gene

Expression

? Transcription is the primary site

of regulation

? Role of Promoter

30
Schematic showing the transcription control regions in

a hypothetical mRNA-producing eukaryotic gene

transcribed by RNA polymerase I

31

The eukaryotic basal transcription complex

32
Stages of Regulation of Eukaryotic

Gene Expression

? A. Coordinate regulation

? 1. Galactose circuit

? Coordinated expressin mediated by Gal4

? 2.Hormone response system

? Intracellular receptors
? Cell surface receptors

33

Regulation of Galactose circuit in yeast

34
Transcriptional regulation by intracellular steroid hormone

receptors.

GRE = glucocorticoid-response element

(an example of a hormone response element); GR = glucocorticoid

receptor.

35

Transcriptional regulation by

receptors located in the cell

membrane.

36
Regulation of Eukaryotic Gene

Expression

? B. Regulation by co- and posttranscriptional

processing of mRNA

? 1. Splice-site choice/ Alternate splicing

? Tissue-specific protein products

? tropomyosin (TM) regulates the functions of actin

in both muscle and nonmuscle cel s

? 2. Alternative polyadenylation

? 3. mRNA editing

37

RNA editing of apo B in the intestine and generation of the

apo B-48 protein needed for chylomicron synthesis

38
? 4. mRNA stability

? A. IRON Metabolism

? Regulation by mRNA degradation--Regulation of transferrin

receptor (TfR) synthesis

IRE = iron-responsive element; IRP = iron-responsive element binding protein.

39

? Ferritin synthesis is regulated at the level of

translation

? Ferritin mRNA contains IRE at 5' UTR
? Binding with IRP prevents translation because it overlaps the

translation initiation site

? Iron availability is more: Fe binds to IRP-allows translation of

free mRNA of ferritin

? Iron metabolism involves both mRNA stability and translation

level

40
3. mRNA stability

--b. RNA interference (RNAi) by

cleavage of target mRNA.

miRNA

si RNA

RNAi therapy:

Age-related macular degeneration (AMD) is triggered

by overproduction of vascular endothelial growth factor

(VEGF). The siRNA drug targets the mRNA of VEGF and

Promotes its degradation- injected in eyes.

41

C. Regulation at Translation level

Regulation of translation initiation in eukaryotes by phosphorylation

of eIF-2. RER = rough endoplasmic reticulum.

42
D. Regulation through modifications

to DNA

? 1. Access to DNA
? 2. Amount of DNA:
? 3. Arrangement of DNA
? 4. Mobile DNA elements

43

1. Access to DNA

? Histone acetylation/deacetylation by the

histone acetyltransferase and histone

deacetylase enzymes

? Extent of methylation of cytosine bases in CG

rich regions (CpG islands) in the promoter

region by methyltransferases that use S-

adenosyl - methionine as the methyl donor

44
2. Amount of DNA:

? Gene amplification : increase in copy

Number seen in response to particular

chemotherapeutic drugs such as methotrexate

inhibitor of the enzyme dihydrofolate

reductase (DHFR),

required for the synthesis of thymidine

triphosphate (TTP)

in the pyrimidine biosynthetic pathway

? Resistance to the drug

45

3. Arrangement of DNA

V=Variable, D= Diversity, J= Joining

46
4. Mobile DNA elements

? Transposons comprise 50% 0f human genome
? Movement is mediated by transposase
? Movement can be

? Direct
? Replicative

? Retrotransposon (90% of transposon)
? Hemophilia, Duchenne Muscular dystrophy
? Antibiotic resistance ?Exchange of Plasmids containing

Tn carrying antibiotic resistance genes

47

Summary

? Gene expression results in the production of a functional gene product

(either RNA or protein

? Genes can be either constitutive or regulated

? Regulation of gene expression occurs primarily at the level of

transcription in both prokaryotes and eukaryotes

? mediated through the binding of trans-acting proteins to cis-acting

regulatory elements on the DNA

? In eukaryotes, regulation also occurs through modifications to the DNA,

as well as through posttranscriptional and posttranslational events

? In prokaryotes regulation is achieved through operons

? Operon: groups of genes sequentially arranged on the chromosome along

with the regulatory elements that determine their transcription

? operon is induced by an isomer of lactose (al olactose)

? The trp operon is also regulated by attenuation

48
? Transcription of rRNA and tRNA is selectively inhibited in prokaryotes by

the stringent response to amino acid starvation

? Gene regulation is more complex in eukaryotes. Operons are not present,

but coordinate regulation of the transcription achieved through the

binding of trans-acting proteins to cis-acting elements

? In multicellular organisms coordinated regulation, achieved through

binding of hormone receptor?hormone complex to the DNA

? Co and posttranscriptional regulation is also seen in eukaryotes

? Regulation at the translational level can be caused by the

phosphorylation (inhibition) of eIF-2.

? Gene expression in eukaryotes is also influenced by availability of DNA to

the transcriptional apparatus

49

MCQ1

? Which of the following is the basis for the

intestinespecific expression of apoprotein B-

48?

? A. DNA rearrangement and loss
? B. DNA transposition
? C. RNA alternative splicing
? D. RNA editing
? E. RNA interference

50
MCQ 2

? Which of the following is best described as

being trans-acting?

? A. CAP site
? B. Operator
? C. Promoter
? D. Repressor

51

MCQ 3

? Which of the following is most likely to be true in

hemochromatosis, a disease of iron accumulation in the

body?

? A. The mRNA for the transferrin receptor (TfR) is stabilized

by the binding of IRPs to 3' stem-loop structures known as

IREs.

? B. The mRNA for the TfR is not bound by IRPs, and is rapidly

degraded.

? C. The mRNA for apoferritin is not bound by IRPs at its 5'

stem-loop IRE, and is translated.

? D. The mRNA for apoferritin is bound by IRPs, and is not

translated.

? E. Both B and C

52
MCQ 4

? After several weeks of chemotherapy with

methotrexate, a cancer patient 's tumor begins to show

signs of resistance to treatment. Which of the following

mechanisms is most likely to explain this resistance to

methotrexate?

? A. Deficiency of thymidylate synthase

? B. Overproduction of xanthine oxidase.

? C. Deficiency of PRPP synthase.

? D. Deficiency of thymidine kinase.

? E. Overproduction of dihydrofolate reductase

53

DNA regulatory protein

? Proteins that regulate gene expression:

Repressor, enhancer, silencer

? These proteins show several motifs which are

involved in these DNA protein interaction

? Only a small part of the protein has direct

contact with DNA

? Interactions are reversible and involve non

covalent bonds

? Some commonly seen structural motifs

54
? Helix turn helix motif
? Zinc finger motif
? Leucine Zipper motif

55

Helix turn helix motif

E,g. Lambda repressor, CAP protein, Tryptophan repressor, Homeobox protein in mam5m6 als
Zinc fingers are a series of repeated domains (two to nine) in which

each is centered on a tetrahedral coordination with zinc.

Steroid thyroid receptor protein and calcitriol (vitamin D) receptor protein

57

The leucine zipper motif

58
A schematic model of the DNA-binding domain of C/EBP

cfos.cjun or cjun.cjun

59

Transcription Factors That Contain DNA Binding Motifs

60
61

This post was last modified on 05 April 2022