20. Control of gene expression operons

Question 1

why regulate gene expression?

Answer 1

• high metabolic energy to synthesise RNA & protein
• some genes active all the time because of products in constant demand
  – constitutive
• others turned off mostly, only switched on when products needed
  – induced

Question 2

how do prokaryotes regulate gene expression?

Answer 2

• control of transcriptional initiation

Question 3

how do eukaryotes control gene expression?

Answer 3

• initiation of transcription
• DNA folded chromatin control
  – dsDNA wrapped around histones
  – controls activity of genes
• epigenetic control
  – region in DNA, far from gene controls expression
• mRNA transcript processing
• RNA transport from nucleus to cytoplasm
• protein stability
  – more stable if required for longer
• protein transport
• post-translational modifications
  – protein gains more stability
  – can be earmarked for degradation

Question 4

what are bacterial promoters?

Answer 4

• consist of 2 short DNA sequences
  – separated by defined number of bases
• two primary sequences control expression level of promoter
  – (-10 & -35)
• these sequences are positioned roughly -10 &-35 in relation to start position of transcription they initiate
  – -10 box sometimes refferred to ‘Pribnow box’ or TATA box
• sequence of these positions are very important and consensus sites are shown

Question 5

how do eukaryotes regulate transcription?

Answer 5

• transcription factors
• proteins bind to DNA near start of transcription of gene
  – within promoter region
  – between promoter region or transcription start site
  – upstream the promoter
• either inhibit or assist RNA polymerase
  – in initiation
  – or maintenance of transcription
• prokaryotes do not have this level of control

Question 6

how do enhancers enable eukaryotic transcription regulation?

Answer 6

• epigenetic control
• stretches bases within DNA
  – about 50-150 base pairs in length
• elicits response near/far from promoter
• structure of DNA able to fold back on itself
  – elicits response at different spot within DNA
• activities greatly increased in promoters
  – can exert stimulatory actions over several thousand base pairs

Question 7

what is gene control in prokaryotes?

Answer 7

• group functionally related genes together
  – operon, for joint regulation to perform coordinated function
• clustered genes transcribed together from one promoter
  – gives polycistronic messenger
• genes of same operon have related functions in cell
  – expressed and repressed together

Question 8

what was the first operon discovered?

Answer 8

• lac operon
  – named because products involve in lactose breakdown
• genes needed for metabolism of lactose
• E. coli switches from glucose to lactose metabolism

Question 9

what does an operon consist of?

Answer 9

• a promoter
  – binding site for RNA polymerase
• repressor (operator)
  – overlaps promoter
  – when repressor protein binds to operator RNA, polymerase will not transcribe genes
• structural genes

Question 10

how does an operator work?

Answer 10

• repressor proteins encoded by repressor genes
  – synthesised to regulatre gene expression
• bind to operator site, blocking transcription by RNA polymerase

Question 11

how does a promoter work?

Answer 11

• promoter sequences recognised by RNA polymerase
• RNA polymerase binds to promoter
  – transcription occurs

Question 12

what are activators?

Answer 12

• RNA polymerase acticity
  – regulated by interaction with accessory proteins (activator)
• presence of acticator
  – removes repression, transcription occurs

Question 13

how are bacterial operons transcriptionally regulates?

Answer 13

• two major modes
  – repression
  – induction
• induction
  – in operons producing gene products needed forutilisation of energy
  – catabolic pathways
• repression
  – produce gene products necessary for synthesis of small biomolecules (eg. amino acids)
  – anabolic pathways

Question 14

give 5 repective points of induction and repression

Answer 14

• induction
  – turns on operon
  – starts transcription and translation
  – caused by new metabolite, needing enzymes to get metabolised
  – operates in catabolic pathway
  – prevents repressor by joining operator gene
• repression
  – turns operon off
  – stops transcription and translation
  – caused by excess existing metabolites
  – operates in anabolic pathway
  – aporepressor enabled by co-repressor to join operator gene

Question 15

what is gene repression?

Answer 15

• effector moleculte interacts with repressor protein
  – binds to operator, stopping transcription
• binding of effector molecule to repressor in repressible systems
  – greatly increases affinity of repressor for the operator
  – repressor binds and stops transcription by blocking RNA polymerase from binding to promoter
• negative control
  – mediated by repressor
• expression form repressible operon attenuated by sequences within transcribed RNA

Question 16

what is an example of a repressible operon

Answer 16

• trp operon
  – responsible for biosynthesis of tryptophan
• the addition of tryptophan (effector molecule) to E.coli environment
  – shuts off system because repressors bind to operator

Question 17

what is gene repression?

Answer 17

• regulator gene (Reg.)
  – synthesises inactive repressor protein
  — activated by co-repressor binding before it can bind to operator (O) to block transcription
• absense of co-repressor
  – repressor is inactive
  – can’t bind to operator and transcription occurs

Question 18

what is the Trp Operon?

Answer 18

• contains structural genes encoding for 5 enzymes in the biosynthetic pathway of tryptophan
• under control of repressor protein coded for by trpR gene
  – excess tryptophan inhibits transcription of operon genes by acting as co-repressor and activating repressor protein

Question 19

what is the processess of Trp Operon

Answer 19

• tryptophan present, repressor bound to operator, operon repressed
  – when complexed with tryptophan, repressor protein produced by trpR gene binds to trp operator, preventing RNA polymerase from transcribing operon genes
• tryptophan absent, repressor not bound to operator, operon derepressed
  – in absenseof tryptophan, free trp repressor cannot bind to operator site, RNA plymerase can then move past operator and transcribe trp operon genes. Giving cell capability to synthesis tryptophan.
• tryptophan plentiful, ribosome continues, allowing 3-4 transcription termination signal to form.
  – the moving ribosome completes translation of leader peptide, pausing at stop codon, blocking region 2. Thus, 2-4 structure forms and terminates transcription near end of leader sequence.

Question 20

what is an inducible system?

Answer 20

• effector molecule interacts with repressor protein such that can’t baind to operator
• binding of effector molecule to repressor
  – greatly reduces affinity of repressor for operator
  – repressor released and transcription goes ahead
• inducible operon under positive control
  – mediated by an activator

Question 21

what is an example of an inducible system?

Answer 21

• catabolite-mediated operon
  – lac operon
  – responsible for obtaining energy from galactosides (such as lactose)

Question 22

what is gene induction?

Answer 22

• regulator gene (Reg.)
  – synthesises active repressor that binds to operator (O)
  – blocks RNA polymerase binding to promoter (P) unless unducer inactivates it
  – repressor protein is inactive and transcription occurs

Question 23

explain E/coli’s lac operon

Answer 23

• obtains energy from galactosides
  – lactose
• B-galactosidase
  – in presence of lactose is inducible enzyme
  – each E.coli has 3000 molecules of B-gal
  – in absense of lactose, have <3 molecules of B-gal
• operon encodes genes facilitating lactose import and breakdown