Gene regulation

1. GENE
REGULATION

2. Pt. Ravishankar Shukla University
Center for Basic Sciences
Presentation on
Session: 2022-23
Presentation by-
Meenal Meshram
5th semester
Guided by-
Dr. Jipsi Chandra

3. Contents
 Prokaryotic gene regulation
1) Lac operon
2) Tryptophan operon
 Eukaryotic gene regulation
1) Chromatin structure on transcription
2) DNA methylation/gene regulation
3) Post-transcriptional gene reguation

4. PROKARYOTIC GENE
REGULATION

5. OPERON

6.  POLYCISTRONIC
 Polycistronic mRNA is a mRNA that encodes several proteins and is
characteristic of many bacterial and chloroplast mRNAs.
 Polycistronic mRNAs consist of a leader sequence which precedes the first
gene.
 MONOCISTRONIC
 Monocistronic mRNA is a mRNA that encodes only one protein and all
eukaryotic mRNAs are monocistronic

7. BACTRIAL METABOLISM
 Bacteria need to respond quickly to changes in their environment
- if they have enough of a product, need to stop production
 why? waste of energy to produce more
 how? stop production of enzymes for synthesis
- if they find new food/energy source, need to utilize it quickly
 why? metabolism, growth, reproduction
 how? start production of enzymes for digestion

8. TYPES OF REGULATED GENES
 CONSTITUTIVE GENES - Constitutive genes are those that are always active. Genes
for ribosomes are an example. They are constantly being transcribed because
ribosomes are constantly needed for protein synthesis.
 INDUCIBLE GENES - Inducible genes are those that have variable activity,
depending on the needs of the cell. For example, the glucose transporter proteins that
muscle cells produce in response to insulin are the product of inducible genes.
 REPRESSIBLE GENES - Repressible genes are those in which the presence of a
substance (a co-repressor) in the environment turns off the expression of those
genes (structural genes) involved in the metabolism of that substance. e.g., Tryptophan
represses the expression of the trp genes.

9. OPERON
operon, genetic regulatory system
found in bacteria and their viruses
in which genes coding for
functionally related proteins are
clustered along the DNA.
This feature allows protein
synthesis to be controlled
coordinately in response to the
needs of the cell.

10. ORGANIZATION OF BACTERIAL OPERON

11. ORGANIZATION OF BACTERIAL OPERON
 Structural genes: code for the enzymes themselves. Lie adjacent to one another, RNA pol
moves from one structural gene to the next, transcribing all genes into a single RNA
 Promoter: RNA polymerase binding site
 single promoter controls transcription of all genes in operon
 transcribed as one unit & a single mRNA is made
 Operator: resides adjacent to or overlapping with the promoter, serves as the binding site for a
protein, called the repressor (gene regulatory protein)
 Regulatory gene: encodes the repressor protein

12. Lac operon
 The lac operon is an operon, or group of genes with a single promoter (transcribed as a
single mRNA).
 The genes in the operon encode proteins that allow the bacteria to use lactose as an
energy source.
 The lactose operon (lac operon) is an operon required for the transport and metabolism
of lactose in E. coli and many other enteric bacteria.
 The gene product of lacZ is β-galactosidase which cleaves lactose, a disaccharide, into
glucose and galactose.

14. FOUR SITUATIONS ARE POSSIBLE
1. When glucose is present and lactose is absent the E. coli does not produce ß-
galactosidase.
2. When glucose is present and lactose is present the E. coli does not produce ß-
galactosidase.
3. When glucose is absent and lactose is absent the E.coli does not produce B-
galactosidase.
4. When glucose is absent and lactose is present the E. coli does produce B-
galactosidase.

15. SUMMARY

16. The Lac Operon
 It consist of:
Promoter
Operator
Structural genes
Lac Z (B-galactosidase)
Lac Y(permease)
Lac A(transacetylase)
Polycistronic m-RNA

17. OPERATORS ARE CIS- ACTING

18. REPRESSOR ARE TRANS- ACTING

19. Lac mutant

20. MUTANTS

21. LACTOSE ANALOG
Isopropyl-B-D-thiogalactoside (IPTG)
 IPTG binds to repressor and inactivates
 IPTG cannot be metabolized by E. coli
 Its concentration remains constant
Phenyl-ß-D-galactose (phenyl-Gal) phenyl-Gal)
 It is a substrate for B-galactosidase, but does not inactivate repressor and so is not an inducer.
 Since wild type cells produce very little ß-galactosidase, they cannot grow on phenyl Gal as a carbon and
energy source.
 Mutants lacking repressor are able to grow on phenyl-Gal.
 Minimal medium containing only phenyl-Gal as a source of carbon and energy is79 selective for repressor
mutants and operator mutants.it, but is not a substrate for ß-galactosidase.

22. POSITIVE GENE REQULATION

25. Tryptophan operon
 The trp operon, found in E. coli bacteria, is a group of genes that encode
biosynthetic enzymes for the amino acid tryptophan.
 The trp operon is expressed (turned "on") when tryptophan levels are low and
repressed (turned "off") when they are high.
 The trp operon is regulated by the trp repressor. When bound to tryptophan,
the trp repressor blocks expression of the operon.
 Tryptophan biosynthesis is also regulated by attenuation (a mechanism based
on coupling of transcription and translation).

26. General organization of theTryptophan operon of E.coli

27. Regulation of the trp operon:
 Two mechanisms regulate the trp operon:
 1. Repressor/operator interaction
 2. Termination of initiated transcripts

28. Repressor/operator interaction
 When tryptophan is present, tryptophan binds to trpR gene product.
 trpR protein binds to the trp operator and prevents transcription.
 Repression reduces transcription of the trp operon~70-fold.

31. Termination of initiated transcripts
 Transcription also is controlled by attenuation, process of translating a short,
incomplete polypeptide.
 When cells are starved for tryptophan, trp genes are expressed maximally..
Under less severe tryptophan starvation, trp genes are expressed at lower than
maximum levels.
 Attenuation can regulate transcription level by a factor of 8 to 10, and
combined with the repression mechanism, 560-700 fold.

32. Leader sequence

33. The tryptophans are important because:
If there is plenty of tryptophan, the
ribosome won't have to wait long
for a tryptophan-carrying tRNA,
and will rapidly finish the leader
polypeptide.
If there is little tryptophan, the
ribosome will stall at the Trp
codons (waiting for a Trp-carrying
tRNA) and will be slow to finish
translation of the leader.

34. EUKARYOTIC GENE
REGULATION

35. Levels Of Gene Regulation In Eukaryotes
Transcription
Post-
transcriptionally

36. Influence Of Chromatin
Structure On Transcription
1. Covalent Histone Modification
2. Nucleosome Remodeling

37. Covalent Histone Modifications
 Structural changes in histone proteins that are associated at the time of
replication and transcription
 Mediated by chromatin modifiaction
 Types of post transcriptional modifications of histone-
I. Acetylation
II. Methylation
III. Phosphorylation

38. Acetylation
 Enzyme histone acetyltransferase (HATs) adds the acetyl group to the lysine
amino acid residues in the histone tail.
 Neutrilize the positive charge.
 Reduces the binding between histones and DNA, which provides sites for
effector proteins that can change the chromatin in active state.
 More accessible for transcription
 Reversible process.
 Deacetylated by enzyme histone deacetylase (HDACs).

40. Methylation
 Many lysine and arginine amino acid residues at the N-terminal histone tails
undergo methylation.
 Methylation is catalyzes by enzyme histone methyltransferases (HMTs) and
demethylation is by histone demethylases (HDMs).
 Methylation of different parts of the N-terminal tails of H3 and H4 histones is
associated with both activation and repression.
 Transcription activation – methylation of lysine at 4th position of H3
Repression – methylation of lysine at 9th poation of H3
 Fundamental role in heterochromatin formation, chromosome X-inactivation,
genomic imprinting and transcription regulation.

42. Phosphorylation
 It takes place in on serines, threonines and tryosines in the N-terminal of
histone protein.
 Controlled by kinases and phosphatases.
 These modifications leads to chromatin condensation and the regulation of
charomatin structure during mitosis. Also serves as a recruiting point for DNA
damage repair protein.

43. Different classes of histones modifications and their role
Type of modification Residues modified Function regulated
Acetylation Lysine
Transcription, Repair,
Condensation
Methylation Lysine and Arginine Transcription, Repair
Phosphorylation
Serine, Threonine and
Tyrosine
Transcription, Repair,
Condensation

44. Nucleosome Remodeling
 Involes-
1) Sliding
2) Histone exchange
3) Nucleosome eviction
 ATP dependent chromatin remodeling complexes (CRC) responsible for
nucleosome remodeling.
 CRCs are two types swi/snf and ISWI

46. DNA Methylation And Gene
Regulation

47.  DNA methylation occurs predominantly at CpG dinucleotides sequence.
 Enzyme DNA methyltransferase mediates the transfer of a methyl group to cytosine, generating
5-methylcytosine.
 The regions of the genome with high number of methylated cytosine are usually
transcriptionally inactive.
 Sliencing is done by either inhibition of transcription factors through methylayed cytosine or
binding of other protein at CpG methylated site.

49.  There are two types of methylation process known in eukaryotic cells-
• Addition of methyl groups occurs in a
newly synthesized strand of DNA at
positions opposite to the parent strand
Maintenance
methylation
• Addition of methyl groups occurs at
totally new potions of daughter strand
De novo
methylation

51. Regulation Of Transcription By Methylation
 DNA methylation switches off the
eukaryotic gene expression, particularly
when it occur in the promoter region of
upstream.
 Protein containing methyl-CpG binding
domain (MBD) recognize the DNA
methylation.
 These proteins binds to 5-methyl cytosine
residues and inhibit the transcription by
recruiting the protein complexes that
induce the alteration of the chromosome
structure.

52. Genomic Imprinting
 Epigenetic phenomenon that causes genes to be expressed in a parent-specific
manner.
 There are few imprinted genes whose expression depends on weather they are
inherited from the mother or from the father.
Ex. The gene encoding for insulin-like growth factor 2 (Igf2) is only expressed
from the allele inherited from father.
 In germline cells the imprint is erased and then re-established according to the
sex of the individual, i.e., in the developing sperm (during spermatogenesis), a
paternal imprint is established whereas in developing oocytes (during
oogenesis), a maternal imprint is established.

54. Post-transcriptional Gene
Regulation

55. Post-transcriptional Gene Regulation
 Post-transcriptional regulation may occurs at the level of translation.
 Here, regulatory proteins may bind to DNA and either promote or repress
transcription.
Ex. The response of mRNA to iron supply for translational represion

57. Multiple Choice Question
1.Which of the following is the most appropriate definition of an operator?
a) A non-coding, regulatory DNA sequence that is bound by RNA polymerase
b) A non-coding, regulatory DNA sequence that is bound by a repressor protein
c) A DNA-binding protein that regulates gene expression
d) A cluster of genes that are regulated by a single promote
2. Which of the following statements about lac operon in e.coli is true?
a) Promoter is the binding site for the lac repressor
b) Operon is only switched on in the absence of lactose in the growth medium
c) β-galactosidase is only produced in large quantities when the lac repressor is bound to the
operator
d) Lac operon mRNA is a polycistronic mRNA

58. Multiple Choice Question
3.Which of the following is not part of the lac operon?
(a)I
(b)O
(c)P
(d)none of these
4. In the presence of high levels of tryptophan
(a)attenuator allows transcription of trp structural genes
(b)attenuator propogates transcription
(c)attenuator terminates transcription
(d)none of the above

59. Multiple Choice Question
5. Which of the following occur in the presence of glucose?
(a)lac Z gene expression is increased
(b)cAMP increases
(c)Binding of CAP-cAMP complex to the promoter area decreases
(d)none of the above
6.In eukaryotes, transcriptional gene control is mediated through
(a) metabolites that bind to cis-acting elements
(b) failure of trans-acting factors to bind to cis-acting elements
(c) binding of trans-acting factors to cis-acting components
(d) proteins that attach to operator sites and act as repressors

60. Multiple Choice Question
7.The most frequent type is in eukaryotes and bacteria.It is necessary to regulate.
(a) regulation of the promoter
(b) regulation of translation
(c) regulation of the repressor
(d) transcriptional regulation
Answers
1.(b), 2.(d), 3.(a), 4.(c), 5.(c), 6.(c), 7.(d)

61. References
• Watson J. D., Hopkins, N. H.,Roberts, J. W., Steitz, J. A. andWeiner, A. M.
Molecular biology of the gene, 4th edition, TheBenjamin/Cummings publishing companies,
• Stryer L
Biochemistry, 4 th edition