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Lecture 31 Flashcards

1
Q

What kind of genes are under constitutive gene experssion?

A

House keeping genes

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2
Q

How are genes regulated in prokaryotes?

A

Mainly use transcriptional control

amount of mRNA and transcription taking place dictates how much protein is produced

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3
Q

Role of operator in prokaryote

A

Adjacent to promoter, binding site for specific protein that help to regulate gene expression

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4
Q

What is the preferred energy source of Ecoli

A

glucose

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5
Q

Which sugar does E.coli use when there is absence of glucose

A

Lactose, it is converted into glucose

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6
Q

How is gene organization in bacteria?

A

Polycistronic operon in which several proteins are coded by one mRNA

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7
Q

3 proteins that are produced in absence of glucose and presence of lactose?

A

Beta galactosidase
Permease
Transacetylase

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8
Q

Role of Beta galactosidase

A

breaks 1-4 glycosidic link in lactose disaccharide to make D-glucose and D-galactose
(Cytoplasmic galactosidase turn lactose to allolactose)

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9
Q

Role of permease

A

helps lactose move through the cell membrane

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10
Q

Transacetylase

A

help in detoxification of beta galactosides

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11
Q

Lac Z

A

Beta galactosidase

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12
Q

Lac Y

A

Lactose permease

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13
Q

Lac A

A

Transacetylase

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14
Q

Lac O

A

Operator where binds repressor protein

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15
Q

Lac P

A

Promoter when binds RNA pol

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16
Q

Lac I

A

repressor which turns gene off (always on unless there is lack of glucose and presence of lactose)

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17
Q

Negative regulation of Lac operon

A

When glucose is present, Lac I makes repressor which binds to LacO sequence, thus physically blocking RNA polymerase at promoter initiating transcription.&raquo_space;System turned off

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18
Q

Induction of operon

A

allolactose (produced by cytosolic beta galactosidase) binds to repressor at Lac O, leading to dissociation from operator seq. Then, RNA pol can initiate transcription.
Thus induction by lactose turns operon on

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19
Q

Inducer of lac operon regulation

A

Allolactose

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20
Q

Activation of lac operon

A

cAMP (hunger signal), produced by adenylate cyclase, binds to Catabolite Activator Protein (CAP), forming cAMP regulatory protein (CRP). Active cAMP-CAP binds lac P and helps activate RNA pol for transcription in presence of lactose.

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21
Q

Hunger signal for lac operon

A

cAMP

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22
Q

Role of catabolite activator protein (CAP)

A

Bind with cAMP and binds to lacP, activating RNA pol at promoter seq of lac operon

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23
Q

Activator of lac operon

A

CAP

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24
Q

What happens to lac operon when there are glucose and lactose present

A

Allolactose is bound to repressor thus leading to dissociation of repressor, but adenylyl cyclase is inactive in the presence of glucose and CAP is not bound to cAMP, so RNA pol can’t initiate transcription&raquo_space; operon off

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25
Q

What is LacIs

A

mutation in lac I, leading to high affinity to operator, Super-repressor, which will not dissociate from operator despite the presence of allolactose, leading to system being always off

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26
Q

What is LacI-

A

Mutation in Lac I, leading to non-functional repressor. It is unable to bind to operator. System always on!&raquo_space; constitutive expression of genes

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27
Q

What is Lac Oc

A

Mutation in Lac O, A non functional operator, thus repressor cannot bind. System always ON&raquo_space; Constitutive expression of genes

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28
Q

Main mechanisms of gene regulation in Eukaryote

A 
Transcriptional control
mRNA processing
mRNA Transport
mRNA stability
Translation
Epigenetic modification
29
Q

What is Cis regulatory elements?

A

DNA binding sites, such as transcription factor vinding sites or enhancer binding sites

30
Q

What is Trans regulatory elements

A

proteins like transcription factors or enhancer binding proteins
- transcribed at a different location on the genome, translated in cytosol, then bind DNA at the cis elements or bind RNA pol II activating transcription

31
Q

Control of gene transcription in eukaryote (4)

A
  1. Cis regulatory sequence
  2. Transcription factor complexes (Trans)
  3. Epigenetics (regulate chromatin conformation)
  4. Small nuclear RNA = miRNA, siRNA
32
Q

Epigenetics (2)

A

gene methylation / histone acetylation

33
Q

What is miRNA & siRNA

A

They never leave nucleolus or get translated, but regulate transcription

34
Q

3 Example of cis regulatory element

A
  1. Basal promoter sequence - TATA or CAAT box or GC box that are upstream of start codon in promoter region
  2. Proximal control region - found near promoter
  3. Enhancer sequence (far away from promoter)
35
Q

Role of enhancer binding proteins

A

When it binds to the enhancer, it bends DNA so that the enhancer binding proteins that is far from the promoter can interact with transcription factors and enhance transcription by RNA pol II, leading to upregulation of transcription

36
Q

Location of enhancer

A

It can be located before or after a gene or even in an intron or distant from the gene (thousand of base pairs)

37
Q

Basal and upstream promoters

A

Basal transcription machinery maintains a low level of expression, but upregulation by upstream or downstream enhancer allows tight regulation.

38
Q

Structure of trans element (3)

A
  1. DNA binding domain
  2. Dimerization domain
  3. Activation/repression domain
39
Q

Dimerization domain of trans element

A

2 TFs bind together to form a functional DNA binding unit. Dimer formation adds an extra element of complexity and versatility

  • Zinc finger
  • Helix-turn-helix
  • leucine zipper
40
Q

What happens when activator binds to enhancer sequence?

A

it stabilizes the basal transcription apparatus and transcription level increase.

41
Q

3 modes of action via which TFs act as repressor

A
  1. Competition - repressor compete with activator to bind the enhancer region which reduce transcription levels
  2. Quenching - repressor binds to DNA-binding domain of an activator protein, disabling activator from binding to enhancer seq.»reduce transcription level
  3. Blocking - repressor bind to activation domain of activator protein, preventing activator from interacting with basal transcriptional machinery.
42
Q

Difference between response element in eukaryote and polycistronic gene regulation in prokaryote

A

In prokaryote, several genes are regulated by a single common regulatory element, but in eukaryote, the same response element allows multiple genes to be regulated together (turned on or off at the same time), but not polycistronic

43
Q

Role of hypoxia inducible factor (HIF)

A

Regulate gene responses to low oxygen
- HIF1alpha and HIF1beta TFs must dimerize, then bind to hypoxia response element which upregulate genes that are involved in many aspects of cell survival, glucose metabolism, angiogenesis, cancer progression and invasion.

44
Q

Sequence of hypoxia reponse element

A

5’-TACGTG-3’

45
Q

HIF-1 Regulation in high oxygen

A

proline residue of PEST amino acid sequence of HIF-1alpha is hydroxylated by prolyl hydroxylase. As a result, oxygen dependent degradation domain signals for ubiquitin-proteosome mediated degradation of HIF-1alpha&raquo_space; no dimerization&raquo_space; no binding to HRE

46
Q

HIF-1 Regulation in low / no oxygen

A

In no oxygen, HIF-1alpha is stabilized translocate into nucleus and hetero-dimerizes with HIF-1beta. The dimer binds to HRE. Then, dimer binds to basal TF machinery with its activating domain which upregulates gene transcription.

47
Q

Difference between HIF-1 alpha and beta

A

alpha is induced by oxygen while beta is constituitive. Alpha has different transactivation domain to beta (alpha has PEST seq and Oxygen dependent degradation domain (ODD).

48
Q

Relationship between HIF-1 and cancer

A

Solid cancer are often hypoxic, so knowing how to inhibit HIF1alpha and beta provide a way to kill tumors.

49
Q

What type of TF is glucocorticoid receptor?

A

Zinc-finger type TF

50
Q

How is glucocorticoid receptor upregulated?

A

Cortisol binds to GR which cause dissociation of receptor from regulatory complex. GR forms a homodimer, then moves into nucleus, binding to hormone response element(HRE).

  • GR need coactivator protein to interact with basal transcription machinery. Coactivator binds to homodimer and interact with basal transcription complex.
  • However, repressor can also bind to GR homodimer thus inhibiting interaction with basal transcription machinery.
51
Q

Special feature of hormone response element (HRE)

A
  • CIS element : found in promoter and regulatory seq of many genes
  • Inverted repeat of 5’>3’ and 3’>5’ like palindrome
52
Q

Role of activated GR complex physiologically

A

Upregulation of expression of anti-inflammatory genes in nucleus
Repression of expression of pro-inflammatory proteins in cytosol.

53
Q

Upregulation of Myc/max system

A

Myc dimerize with Max. Heterodimer bind to DNA domain (CACGTG), which upregulate cell cycle progression.

54
Q

Downregulation of Myc/max system

A

There is no Myc, so max forms a homodimer. Since max doesn’t have activating domain, the homodimer can’t bind to basal transcription machinery even though it can bind to enhancer DNA, which repress gene transcription&raquo_space; cell cycle inhibition

55
Q

Which system exhibit regulation of gene expression by control of dimerization?

A

Myc and Max system

56
Q

What happens if Myc is over expressed?

A

it leads to disruption of equilibrium between activation and repression of genes leading to impaired differentiation and promotion of cell differentiation > Cancer

57
Q

3 types of hemoglobin

A
  1. Fetal hemoglobin : tetramer of two alpha chain and two gamma chain
  2. Hemoglobin A : tetramer of two alpha chains and two beta chains
  3. Hemoglobin A2 : tetramer of two alpha chain and two delta chains
58
Q

trend of fetal hemoglobin (HbF) expression

A

Expression begin at conception with gamma chain expression exponentially decreasing soon after birth

59
Q

trend of hemoglobin A (HbA) expression

A

Expression of beta begins at conception with exponential increase to plateau at 6month

60
Q

trend of hemoglobin A2 (HbA2) expression

A

Expression of delta chain at low levels after birth

61
Q

Which chromosomes are responsible for globin gene?

A

Chromosome 16 - 2 alpha globin genes

Chromosome 11 - beta, gamma, delta genes

62
Q

What is transcription factor code?

A

gene sequence in all cells are same, but combination of TFs expressed by a cell define the cell. Combination of TFs differs in different cells or tissue.

63
Q

At what level does miRNA and siRNA regulate gene expression?

A

at translational level

64
Q

what is miRNA?

A

Derived from specific ds-pre-miRNA species that is endogenous. It repress mRNA translation

65
Q

What is siRNA

A

Derived from long dsRNAs that is exogenous, regulates expression by mRNA degradation.

66
Q

Steps of miRNA inhibition of translation

A
  1. Long pri-miRNA are processed to pre miRNA hairpin structures by Drosha (hairpin loop has few mismatch pairs)
  2. pre-miRNA is transported to cytoplasm by exportin5
  3. Then dicer process them to single stranded RNA by removing loop part of pre-miRNA and unwinding it, and initiates formation of assymetric RNA-induced silencing complex (RISC).
  4. RISC binds imperfectly to the 3’UTR of target mRNA, which inhibit translation by preventing ribosome from interacting with 5’cap structure.
67
Q

Steps of siRNA degradation of mRNA

A

Occur outside nucleus

  1. Dicer also process long dsRNA into siRNA, which has perfect complementarity.
  2. siRNA forms RISC.
  3. It perfectly binds to 3’UTR of target mRNA
  4. Perfect pairing of siRNA with mRNA within RISC complex activates RISC endonuclease AGO2 activity, resulting in cleavage of mRNA, leading to loss of poly A tail. This leads to degradation of mRNA transcript
68
Q

Relationship between miRNA and cancer

A

Some miRNA are involved in regulation of cell proliferation. And genes that encode these miRNA are amplified in tumor cells leading to duplication of DNA seq. This leads to loss of cell growth regulation leading to uncontrolled cell proliferation (cancer)

69
Q

miRNA code & TF code for gene regulation

A

The combination of TFs AND miRNAs expressed by a cel define the cell.

BPM DLA1 (47 decks)

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