Gene expression,Regulation of gene expression by dr.Tasnim

gene expression regulation of gene expression ,human genom project

1. Gene Expression,Regulation of gene
Expression,Human genom project
DR.TASNIM ARA JHILKYDR.TASNIM ARA JHILKY
MD PART IIMD PART II
Phase APhase A
DEPARTMENT OF BIOCHEMISTRYDEPARTMENT OF BIOCHEMISTRY
SIR SALIMULLAH MEDICAL COLLEGE MIDFORD,DHAKASIR SALIMULLAH MEDICAL COLLEGE MIDFORD,DHAKA

1.

2. Gene Expression
•It is the process by which information from a
gene is used in the synthesis of a functional
gene product.
• These products are often proteins, but in
non-protein coding genes such as rRNA genes
or tRNA genes, the product is a functional RNA
.

3. Gene expression regulation:
Both of these cells
contain the same
genome, but they
express different RNAs
and proteins.

4. Classification of gene with respect to their
Expression:
•Constitutive ( house keeping) genes:
• 1- Are expressed at a fixed rate, irrespective to the
cell condition.
• 2- Their structure is simpler
•Controllable genes:
• 1- Are expressed only as needed. Their amount may
increase or decrease with respect to their basal level
in different condition.
• 2- Their structure is relatively complicated with some
response elements

5. Levels of regulation of gene Levels of regulation of gene
expressionexpression
5.

6. 1.positive regulation:
•When the expression of genetic is quantitatively
increased by the presence of specific regulatory
element is known as positive regulation.
•Element modulating positive regulation is known
as activator or positive regulator.

7. 2.Negative regulation.
•when the expression of genetic information
diminished by the presence of specific
regulatory element.
•The element or molecule mediating the
negative regulation is said to be repressor.

8. Purpose of regulation of gene Purpose of regulation of gene
expressionexpression
Regulated expression of genes is required for
1) Adaptation- Cells of multicellular organisms
respond to varying conditions.
Such cells exposed to hormones and growth
factors change substantially in –
oshape,
ogrowth rate, and
oother characteristics
8.

9. Purpose of regulation of gene Purpose of regulation of gene
expressionexpression
2) Tissue specific differentiation and development
The genetic information present in each somatic cell of
a organism is practically identical.
Cells from muscle and nerve tissue show strikingly
different morphologies and other properties, yet they
contain exactly the same DNA.
These diverse properties are the result of differences in
gene expression.
Expression of the genetic information is regulated
during ontogeny and differentiation of the organism
and its cellular components.
9.

10. Mechanism of regulation of gene Mechanism of regulation of gene
expression-an overviewexpression-an overview
•Gene activity is controlled first and foremost
at the level of transcription.
•Much of this control is achieved through the
interplay between proteins that bind to
specific DNA sequences and their DNA binding
sites.
•This can have a positive or negative effect on
transcription.
10.

11. Mechanism of regulation of gene Mechanism of regulation of gene
expression- An overviewexpression- An overview
•Transcription control can result in tissue-
specific gene expression.
•In addition to transcription level controls,
gene expression can also be modulated by
•Gene rearrangement,
•Gene amplification,
•Posttranscriptional modifications, and
•RNA stabilization.
11.

12. Differences between gene expression Differences between gene expression
in prokaryotes and eukaryotesin prokaryotes and eukaryotes
Gene regulation is significantly more complex in
eukaryotes than in prokaryotes for a number of
reasons:
1) First, the genome being regulated is significantly
larger
•Human cell have larger genome
12.

13. 2) Different cell types2) Different cell types
•Different cell types are present in most
eukaryotes.
•Liver and pancreatic cells, for example, differ
dramatically in the genes that are highly
expressed.
•Different mechanisms are involved in the
regulation of such genes.
13.

14. 3) Absence of operons3) Absence of operons
•The eukaryotic genes are not generally
organized into operons as are there in
prokaryotes
•Instead, genes that encode proteins for steps
within a given pathway are often spread
widely across the genome.
14.

15. 4) Chromatin structure4) Chromatin structure
•The DNA in eukaryotic cells is extensively
folded and packed into the protein-DNA
complex called chromatin.
•Histones are an important part of this
complex since they both form the structures
known as nucleosomes and also contribute
significantly into gene regulatory mechanisms.
15.

17. Principles of Gene Regulation in prokaryote
Most prokaryotic genes are regulated in units
called operons.
Francois Jacob & Jacques Monod, 1961.
This is largely based on regulation of lactose metabolism. By
intestinal bact. E.coli.

18. Different ways for regulation of gene
expression in bacteria:
•1- Promoter recognition.
•2-Transcription elongation( Attenuation).

19. Lac Operon – Lac Operon –
•Bacteria such as E. coli usually rely on glucose
as their source of carbon and energy.
•However, when glucose is scarce, E. coli can
use lactose as their carbon source even
though this disaccharide does not lie on any
major metabolic pathways.
•An essential enzyme in the metabolism of
lactose is β-galactosidase, which hydrolyzes
lactose into galactose and glucose
06/05/17 19
.

20. Components of Lac OperonComponents of Lac Operon
•The genetic elements of the model are a regulator
gene, a regulatory DNA sequence called an
operator site, and a set of structural genes.
•The regulator gene encodes a repressor protein that
binds to the operator site.
•The binding of the repressor to the operator
prevents transcription of the structural genes.
•If the operator is not bound by a repressor
molecules ,RNA polymeraze passes over the
operator and reaches the protein coding genes
which is transcribes to mRNA
06/05/17 20.

21. Components of Lac OperonComponents of Lac Operon
•For the lactose (lac) Operon, the i gene encodes the
repressor, o is the operator site, and the z, y, and a
genes are the structural genes for β -galactosidase,
the permease, and the transacetylase, respectively.
•The Operon also contains a promoter region
(denoted by p), where RNA polymerase binds and
two additional sites operator and CAP site is bound
by cAMP and catabolite activator protein.
06/05/17 21.

22. Components of Lac OperonComponents of Lac Operon
06/05/17 22Biochemistry For Medics

23. Components of Lac OperonComponents of Lac Operon
06/05/17 23Biochemistry For Medics

24. •The z, y, and a genes are transcribed to give a
single mRNA molecule that encodes all three
proteins.
• An mRNA molecule encoding more than one
protein is known as a polygenic or polycistronic
Components of Lac OperonComponents of Lac Operon
06/05/17 24Biochemistry For Medics

25. Regulation of Expression of Lac OperonRegulation of Expression of Lac Operon
1)In the absence of lactose/When glucose is available- Lac
Operon remains repressed (turned off)due to the
presence of lac repressor at the operator site- (Negative
control).
2) In the presence of only Lactose- Lac Operon is turned on.
the structural genes are transcribed and the lactose
metabolizing enzymes are synthesized .A small amount of
lactose is converted to allolactose and binds with
repressor protein and change its conformation ,so
repressor can not binds with operator
In the absence of glucose adenylyl cyclase active ,made cyclic
AMP which binds with CAP causing RNA polymerase to
more efficiently initiate transcription.
06/05/17 25
.

26. Regulation of Expression of Lac OperonRegulation of Expression of Lac Operon
3) In the presence of both glucose and
lactose- CAP -cAMP complex is not
formed(due to inhibition of adenylyl
cyclase),RNA polymerase can not initiate the
transcription of structural genes despite the
fact that the operator site is vacant due to
the binding of lactose/allolactose with lac
repressor.
Lac Operon remains in the repressed state.
06/05/17 26.

27. Maximum Expression of Lac OperonMaximum Expression of Lac Operon
The lac Operon is controlled by two
distinct DNA binding factors;
 One that acts positively (cAMP-CRP
complex) and
The other that acts negatively (LacI
repressor).
Maximal activity of the lac Operon occurs
when glucose levels are low (high cAMP
with CAP activation) and lactose is presen,t
LacI is prevented from binding to the
operator).
06/05/17 27.

28. Lac I
Promoter
gene
Operator
gene
Lac ZLac YLac A
R
Translation &
Transcription
RNA polymerase
No Gene
Expression
06/05/17 28

29. 06/05/17 . 29

30. .
06/05/17 30.

31. Combinatory Regulation of Lac Operon:
CAP: catabolite activator protein; breakdown of lactose when glucose is
low and lactose is present

33. ……..
•The genetic constitutions of nearly all metazoan
somatic cells are identical.
•Tissue or cell specificity is dictated by differences in
gene expression of this complement of genes.
•Alterations in gene expression allow a cell to adapt
to environmental changes.
•Gene expression can be controlled at multiple levels
by chromatin modifications ,changes in transcription,
RNA processing, localization, and stability or
utilization.
•Gene amplification and rearrangements also
influence gene expression.
33.

34. Eukaryotic gene regulation occurs at
several levels:

35. Changes in DNA level
•Is done by
•Gene amplification
•Gene rearrangement

36. Gene AmplificationGene Amplification
•The gene product can be increased by increasing
the number of genes available for transcription of
specific molecules
•Among the repetitive DNA sequences are hundreds
of copies of ribosomal RNA genes and tRNA genes.
•During early development of metazoans, there is an
abrupt increase in the need for ribosomal RNA and
messenger RNA molecules for proteins that make
up such organs as the eggshell.
36,

37. Gene Amplification (contd.)Gene Amplification (contd.)
•Such requirements are fulfilled by amplification of
these specific genes.
• Subsequently, these amplified genes, presumably
generated by a process of repeated initiations
during DNA synthesis, provide multiple sites for
gene transcription.
37.s

38. 5) Gene Rearrangement 5) Gene Rearrangement
Gene rearrangement is observed during
immunoglobulins synthesis.
here segment of DNA move from one location to another in
the genom.These DNA coding changes are needed for
generating the required recognition diversity central to
appropriate immune function.
38.

39. Mechanism of regulation of gene Mechanism of regulation of gene
expression-Transcriptional level expression-Transcriptional level
1) 1) Chromatin RemodelingChromatin Remodeling
•Large regions of chromatin are
transcriptionally inactive /heterochromatin in
some cells while they are either active or
potentially active /euchromatin in other
specialized cells
•For example, the DNA containing the -globin
gene cluster is in "active" chromatin in the
reticulocytes but in "inactive" chromatin in
muscle cells.
39.

40. 40Biochemistry for medics-Lecture notes

41. Cont……
Use of alternative promoter
Some genes has series of promoter s showing
tissue specific expression.so in different tissue
different transcript and different protein
produced
sSSlternative promoter
•S

42. Acetylation is known to occur on lysine residues in
the amino terminal tails of histone molecules.
This modification reduces the positive charge of
these tails and decreases the binding affinity of
histone for the negatively charged DNA.
Accordingly, the acetylation of histones could
result in disruption of nucleosomal structure and
allow readier access of transcription factors to
cognate regulatory DNA elements.
42.
2)Histone acetylation and deacetylation

43. 3)DNA methylation:
is the addition or removal of a methyl group predominantely
where cytosine bases occur consecutively.make
bases occur consecutively.

44. Con….
•Individual gene regulation
•Negative regulation
•Positive regulation

45. Post transcriptional regulation of
gene expresstion
•Done by
•Alternative splicing
•Class switching
•Regulation of RNA stability
•RNA editing

46. Alternative RNA ProcessingAlternative RNA Processing
•Eukaryotic cells also employ alternative RNA
processing to control gene expression.
•This can result when alternative promoters,
intron-exon splice sites, or polyadenylation
sites are used.
•Occasionally, heterogeneity within a cell
results, but more commonly the same primary
transcript is processed differently in different
tissues.
46.

47. Alternative RNA Processing (contd.)Alternative RNA Processing (contd.)
Alternative splicing and processing, results in
the formation of seven unique -tropomyosin
mRNAs in seven different tissues.
47Biochemistry for medics-Lecture notes

48. Calcitonin
gene-related
peptide
61
c
e
ll
1
c
e
ll
2
(four exons)
1 2 3 4
1, 2 & 3
1, 2 & 4
32 amino acids
Reduces bone resorption
37 amino acids
Vasodilator

49. Class switchingClass switching
In this process one
gene is switched off
and a closely related
gene takes up the
function.
During intrauterine
life embryonic Hb is
the first Hb to be
formed.
49.s

50. mRNA stability mRNA stability
•Changes in the stability of a specific mRNA can
therefore have major effects on biologic processes.
•The stability of the m RNA can be influenced by
hormones and certain other effectors.
•The ends of mRNA molecules are involved in
mRNA stability.
•The 5' cap structure in eukaryotic mRNA prevents
attack by 5' exonucleases, and the poly(A) tail
prohibits the action of 3' exonucleases.
50.

51. RNA Editing RNA Editing
51Biochemistry for medics-Lecture notes

52. Translational regulation
•Regulating the activity of initiation factors
,particularly initiation factor-2
•Action of IF2 is inhibited when
phosphorylated,so by phosphprylation and
prevention of phosphorylation of
IF2,translation can be regulated.

53. Post translational regulation
•Protein activation
Some proteins are not active when first
formed.so to be active they undergo
modifications
Folding
Proteolysis
Engymatic cleavage
Regulation of protein degradation or turnover.

54. Specific motifs of regulatory proteinsSpecific motifs of regulatory proteins
•Certain DNA binding proteins having specific
motifs bind certain region of DNA to influence
the rate of transcription.
•The specificity involved in the control of
transcription requires that regulatory proteins
bind with high affinity to the correct region of
DNA.
54.

55. Specific motifs of regulatory proteins Specific motifs of regulatory proteins
(contd.)(contd.)
•Three unique motifs—the helix-turn-helix, the zinc
finger, and the leucine zipper—account for many of
these specific protein-DNA interactions.
•The motifs found in these proteins are unique; their
presence in a protein of unknown function suggests
that the protein may bind to DNA.
•The protein-DNA interactions are maintained
by hydrogen bonds and van der Waals forces.
55.

56. Three unique motifs of DNA binding Three unique motifs of DNA binding
proteinsproteins
Helix –turn- helix
Leucine zipper
Zinc finger
56Biochemistry for medics-Lecture notes

57. Method of quantification of gene
expression
•1)detection of mRNAlevel by
•Northern blotting
•Microarrays
•Reverse transcriptase PCR
•2)Analysis of protein level by
•ELISA
•Western blotting
•Photometry
•proteomics

58. HUMAN GENOME HUMAN GENOME
PROJECTPROJECT

59. arg GTbto Pgodbg The Human Genome
•The human genome is the complete set of genetic
information for humans (Homo sapiens).
•The human genome is by far the most complex and largest
genome.
•Its size spans a length of about 6 feet of DNA, containing
more than 30,000 genes.
•The DNA material is organized into a haploid chromosomal
set of 22 (autosome) and one sex chromosome (X or Y).
Male Female

60. Human Genome Sequencing 2/11/2001
22 autosome + 2 sex chromosomes

From NCBI

61. 5rte sti GTbto Pgodbg A,d6gpe1GPAv What was Human Genome Project(HGP)
•The Human Genome Project
was an international research
effort to determine the
sequence of the human
genome and identify the
genes that it contains.
•The US Human Genome
Project is a 13 year effort,
which is coordinated by the
–Department of Energy (DOE) and
–National Institutes of Health
(NIH).

62. 1.To identify all the genes in human DNA.
2.To develop a genetic linkage map of human genome.
3.To obtain a physical map of human genome.
4.To develop technology for the management of human
genome information.
5.To know the function of genes.
6.Determine the sequences of the 3 billion chemical
base pairs that make up human DNA.
7.Store this information in public databases.
8.Develop tools for data analysis.
9.Transfer related technologies to the private sectors. PdtIi dl GTbto Pgodbg A,d6gpe
Goals of Human Genome Project

63. THANK YOU.