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Foundations To Medicine 2 > Gene Regulation > Flashcards

Gene Regulation Flashcards

1
Q

Briefly outline how gene regulation is started

A
  1. RNA polymerase must find the promoter sequence in DNA before it can start transcribing . The polymerase can bind the DNA directly (in bacteria)

Or

  1. Seek out transcription factors that bind to the promoter (in eukaryotes)
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2
Q

Outline general transcription in bacteria

A

General transcription in bacteria occurs by binding of RNA polymerase to the promoter region of a gene, the subsequent unwinding of the DNA double helix to reveal a single stranded DNA template, and the synthesis of an RNA transcript

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3
Q

Contrast regulated and constitutive

A

Constitutive
-Some genes (constitutive genes) are always expressed within a cell

-Constitutive genes are often refferred to as “housekeeping” gene

Regulated
-Other genes only need to be expressed at certain times

-Therefore, to reduce wasted effort, many genes are regulated, and only expressed under certain conditions

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4
Q

How is gene expression regulated?

A

This regulation can occur at many differentiated steps of gene expression, including:

  • Transcription
  • mRNA processing
  • translation
  • Regulation of protein half life

However, the majority of regulation is in bacteria takes place at the transcriptional level

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5
Q

What is an operator in prokaryotes?

A

In bacteria, regulated genes have an upstream region adjacent to promoter called the operator

The operator is a sequence of DNA which is a binding site for specific proteins that help to regulate gene expression

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6
Q

What are repressors and activators?

A

There are two main types of regulatory protein in bacteria:

  1. Repressors: bind to an operator region and prevent RNA polymerase from initiating transcription
  2. Activators bind to or near an operator region and allow RNA polymerase to initiate transcription

A given gene (or group of genes ) may use either or both types of protein for regulation

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7
Q

Who discovered the lac Operon?

A

Jacob and Mood studied the regulation of genes required for the metabolism of lactose in bacteria (coordinate regulation)

Won the Nobel prize in physiology or medicine, 1965

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8
Q

What is the responsibility of each gene in the Lac operon?

A

LacZ- B-galactosidase
(Lactose> glucose + galactose)
(Lactose > allolactose)

LacY- permease

(Active transport of lactose across cell membrane )

LacA-Transacetylase
(Galactose > acetylgalactose)

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9
Q

Outline the map of the Lac Operon

A

All three genes share the same promoter (lacP), the same operator (lacO) and are transcribed as a single mRNA(polycistronic )

The gene for a regulatory protein (lacI) is found nearby, but separately from the Operon

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10
Q

Explain Lac operon gene regulation

A

Since the role of Lac is to break down the sugar lactose, the structural (protein coding) genes lacZ, lacY and lacA should only to be expresssed when lactose is present in the cell

-lacI gene encodes a repressor protein that shuts the system down when lactose is not present

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11
Q

Why is the Lac operon inducible ?

A
  • If lactose is added to a cell, the Lac operon will be ‘turned on’
  • Therefore, we say that the system is inducible
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12
Q

Why is the Lac operon inducible?

A

Repressor (LacI) bound to operator, RNA polymerase cannot initiate transcription at the promoter

No lactose- system turned OFF

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13
Q

How is lactose induced?

A

If lactose is added, there will be formation of the isomer, allolactose, which will bind to the repressor

The allolactose-bound repressor undergoes a conformational change and dissociates from the operator sequence. RNA polymerase is then free to initiate transcription.

Lactose present- system turned ON

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14
Q

What are the effects of structural genes of Lac operon having mutations?

A

LacZ-, LacY-, LacA-: structural gene mutations lead to non-functional proteins

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15
Q

What is the effect of lacP- mutations?

A

Non-functional promoter, RNA pol cannot bind so genes will not be expressed

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16
Q

What is the effect of lacO- mutations?

A

Non-functional operator, repressor cannot bind.

Since the system cannot be shut off, this is a constitutive mutation

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17
Q

What is the effect of lacI- mutation ?

A

Non-functional repressor, unable to bind the operator to shut off transcription

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18
Q

What the effect of lacI^S mutations?

A

Super-repressor, unable to dissociate from operator. System always off

19
Q

Contrast Lac operon positive and negative control

A

When a repressor is used to turn the system off, that system is under negative control. The LacI repressor is an example of this

The Lac operon is also under positive control, where an activator protein is used to increase the efficiency of transcription

20
Q

E. coli’s favorite carbon source is glucose…

A
  • While E. coli can metabolize lactose with the products of the Lac operon, it actually ‘prefers’ glucose as a carbon source
  • Therefore, when glucose is present, there is no need to express the genes in Lac operon
21
Q

Summarize positive control of Lac operon

A
  • Genetic expression of Lac operon occurs only if a regulator molecule directly stimulates RNA production (that molecule is cyclic AMP)
  • If a cell runs out of glucose, a small molecule (cyclic AMP or cAMP for short) is produced by active Adenylyl cyclase (converts ATP to cAMP)

This molecule is a ‘hunger signal’ that permits the expression of genes that break down other sugars, including lactose

-cAMP binds the activator protein CRP( cAMP receptor protein) or CAP (catabolite activator protein), which can then bind lacP to help activate transcription

22
Q

Explain the effect of the hunger signal in the Lac operon

A

Unless the hunger signal is given, Lac operon is not expressed even in the presence of lactose

Inactive CRP
no cAMP around means that gluc9se is in the environment

When glucose is present, no cAMP is produced so CRP is inactive

RNA pol cannot EFFICIENTLY initiate transcription

The hunger signal, in the presence of lactose

If no glucose is present, cAMP is produced, which binds to and activates CRP

Activated CRP can then bind to the promoter

23
Q

Where do s allolactose become bound during Lac operon expression?

A

Bound to the repressor so lactose is in the environment

24
Q

What happens after CRP binds to the promoter?

A

When activated CRP binds to the promoter, RNA pol can enter and initiate transcription (IF lactose is present)

If the hunger signal is given, but lactose is absent, the Lac operon isn’t expressed

Repressor bound to operator (lactose absent)—> blocks RNA pol from transcribing the operon

25
Q

Summarize the Lac operon

A

The Lac operon will only be transcribed when lactose is present (repressor unbound the operator) and glucose is absent (CAP/cAMP is bound to CAP binding site)

26
Q

In what circumstances is the Lac operon on?

A

Glucose absent, lactose present

No repressor bound, activator bound - it’s ON

All other circumstances, it’s off

27
Q

Give an overview of gene regulation in eukaryotes

A

Regulation of gene expression is a complex, multi-layered process that is crucial to correctly drive and maintain cell identity during development and adult life

All of these cells contain the exact same genetic information

Different expression of the genes, I.e. Gene regulation results in this variety of cell types. Thus, regulation is more complex in eukaryotes than in bacteria

28
Q

Why is transcriptional control more complex in eukaryotes?

A

Because of nuclear compartmentalization, chromatin remodeling, recruitment of transcription machinery, transcription initiation, elongation, termination

29
Q

What are the key concepts of Cis elements?

A

DNA sites where proteins and trans regulatory elements will bind

  • Latin root, means same side as
  • DNA motifs which are binding sites for transcription factors enhancer protein binding sites
  • These are the DNA sequence itself, the consensus sites mentioned when we discuss transcription
30
Q

What are the key concepts of trans regulatory elements?

A
  • Latin roots which means across or opposite side
  • These proteins are encoded for by different genes in the genome
  • These are typically proteins such as transcription factors/ enhancer binding proteins
  • May have positive or negative effects
31
Q

Where are trans acting elements synthesized?

A

Synthesized from genes that are different from the genes targeted from regulation. Trans-acting factors bind to cis-acting elements of DNA

32
Q

Explain Cis elements

A

May be classsified by location and function

Typically they have a consensus sequence
Sequence among many genes isn’t exact, it is similar

Each of these is a Cis regulatory element:
1. Basal promoter sequence
These bind the general transcription factor proteins which are associated with RNA Pol

  1. Proximal control regions
    These bind transcription factor proteins, and are found near the promoter
  2. Enhancer sequence
    Are found far away from the promoter
33
Q

Why bother with basal and upstream promoters?

A

Multiple levels of control allow for tight regulation

34
Q

What are the regulatory elements?

A

Distal- enhancer element (variety of STF)

Proximal - CAAT box(CTF(STF)) and GC box(Sp1)(STF)

35
Q

What are the core promoter of elements?

A

TATA Box (-25), Inr (+1), and DPE(+25)

All bind to TFIID(GTF)

36
Q

Contrast Basal transcription vs enhancer elements

A

A) Basal transcription machinery allows a low level of expression

B) Binding of trans factors to the proximal promoters (here CTF and Sp1) regulate frequency of transcription initiation

Enhancer trans factors binding to the enhancer element mediate the response to signals such as hormones and regulate which genes are expressed at a given point in time. Mediator proteins may also bind the enhancer binding proteins +/-

DNA bending can cause an enhancer element that is far from the promoter in the linear DNA molecule to interact with the transcription-initiation complex, stimulating transcription

37
Q

What are the enhancer regions?

A

Bending (or looping) of DNA allows distant regions to contact physically
-Enhancer regions can be located before or after a gene (or even in in an intron)

  • Enhancers can be quite from the gene (thousands, or many thousands of base pairs)
  • May be more then 200 bp away and still function as an enhancer
38
Q

Where can enhancer regions be found?

A

Can be found upstream from the promoter region

  • Thousands of base pairs can separate the enhancer sequence from the gene it regulates
  • An enhancer sequence can be downstream from the promoter region
39
Q

What is the DNA Binding domain?

A

Transcription factors have DNA binding domains that only bind to certain DNA sequences

40
Q

What is the dimerization domain?

A

Two transcription factors bind together to form a functional DNA binding unit called a dimer. Formation of a dimer adds an extra element of complexity and versatility

41
Q

What is the activation/repression domain?

A

Binds to the enhancer binding proteins or other transcription factors and modulates their function

42
Q

What are the parts of a transcription factor?

A
  • Activation and/or repression domain
  • dimerization domain
  • DNA binding domain
43
Q

What are 3 examples of eukaryotic transcription factors that must dimerization to function?

A

Zinc finger

Helix-turn-helix

Leucine zipper

44
Q

Explain how different genes can possess the same response element

A
  • Provides a mechanism to COORDINATE gene regulation
  • Allows multiple genes to be regulated together
  • Allow different genes to be turned on and off at the same time
    • Stage specific expression during embryo development
    • Tissue specific expression
    • Allow response to external stimuli-Many examples… including:
  • hypoxia (hypoxia response element)
  • hormonal (hormonal response element)
  • stress and heat (heat shock element)

Foundations To Medicine 2 (37 decks)

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