Chapter 11 Genetic Organization in Higher Organisms

(1)

Chapter 11 Genetic Organization in

Higher Organisms

(2)

그림 11-1

Mitochondria

Cytoplasm (cytosol)

Endoplasmic reticulum (ER) Cell membrane

Golgi body (apparatus)

Cytoskeleton Nucleolus

Nuclear membrane Lysosome

Centrosome

(3)

그림 11-2 Mitochondrion

(4)

그림 11-3 Chloroplast

(5)

Number of genes

Eukaryotes have 10 times more genes than bacteria

(6)

Centromere

Telomere : Greek, telos (end) + meres (part) - located at the ends of eukaryotic chromosomes

consisting of a six base pair sequence repeated about 2000 times - protects the end of chromosome from destruction

- prevents rearrangement of chromosomes

Telomere General eukaryotic chromosome structure

Chromosome arm

Replication origin

(7)

(8)

그림 11-5

During each replication cycle the chromosomes are shortened due to loss of the RNA primer

? replication

replication

replication replication

If cells divides without telomeres, they would lose the end of the chromosome

(9)

그림 11-6

Telomerase

: add a few of the six base

pair to the end of chromosome , or telomere

(10)

However, telomerase is not active in most cells.

It certainly is active in stem cells( 줄기세포 ), germ cells( 생식세포 ), hair follicles( 모낭 ) and (worryingly) in 90% of cancer cells.

Aging and Cancer

“ 복제양 돌리는 같이 태어난 다른 양들보다 텔로미어가 짧았다 !!! ”

(11)

Haploid ( 반수체 )

: possessing only a single set of chromosomes : prokaryotes (bacteria)

Diploid ( 이배체 )

: possessing duplicate chromosomes : most eukaryotes

Back up your files!!

(12)

Nucleosome : DNA + histone

: 200bp DNA + 9 histones

: core particle = 140 bp + 8 histones : remaining 60bp + 9^th histone

Histones are positively charged.

(13)

그림 11-8 Solenoid

: 6 nucleosomes

nucleosome

Each loop has

~50 solenoids

Miniband: 18 loops

Chromatid

: 1,000,000 minibands

(14)

상자 1

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그림 11-9

(16)

그림 11-10

(17)

그림 11-11

Gene structure in Eukaryotes : exon & intron

Coding DNA Non-coding DNA

Single-celled eukaryotes: intron << exon Higher eukaryotes: intron >> exon

cf) pseudogene: defective copy of a genuine gene

(18)

RNA Polymerase No. Genes transcribed:

I genes for large rRNAs (housekeeping)

II genes which code for proteins III genes for tRNA, 5S rRNA and

some other tiny RNAs (housekeeping)

Transcription in Eukaryotes

RNA polymerase II requires accessory proteins, called transcription factors for its activity.

Transcription factors bind to & recognize specific sequences on the DNA (promoter & enhancer)

(19)

그림 11-12

 Transcription apparatus ( 전사기구 )

RNA polymerase II + transcription factors + TATA box factor + other factors

upstream element

TATA box

promoter

initiator box transcription apparatus

(20)

그림 11-13

Upstream

element TATA box Enhancer sequence

3 regions of promoter

1. Initiator box 2. TATA box

3. Upstream element

Enhancer:

- Bind to specific transcription factors &

enhance the rate of transcription

- away from the gene (upstream or downstream from the promoter)

Initiator box

(21)

Role

: regulate gene expression by controlling transcription 4 domains

1) Binding to a specific sequence on the DNA 2) Binding to the RNA polymerase II complex 3) Getting into the nucleus where the genes are

kept

4) Responding to a stimulus of some sort which signals that the gene should be turned on

Transcription Factor

(22)

그림 11-14

Transcription factor MyoD

(23)

Nrf2

 Nuclear factor E2 - related factor 2

 Play a key role in cellular defense mechanism against carcinogens/toxins

 Bind to

antioxidant response element (ARE)

in the promoter region of

phase II detoxifying/antioxidant enzymes

such as glutathione

S-transferase (GST), NAD(P)H:quinone oxidoreductase (NQO1), heme oxygenase (HO-1)

 Nrf2 K/O mice: reduced level of phase II enzyme, susceptible to carcinogens

 Regulated by

Keap1

(for nuclear translocation & proteasomal degradation)



Inducible by toxins as well as certain antioxidants & chemopreventive agents

(Kong et al 2001, Itoh et al 1997, Fahey et al 2002)

(24)

Cellular membrane

Cytosol

Nucleus

Chemopreventive agents

Electrophiles, ROS

Nrf2

Nrf2

ARE

Maf

CBP/p300

PI3K PKC MAPK

PERK ER stress

Nrf2 P

P

Phase II genes Antioxidant genes Degradation Nrf2

degradation degradation

Nrf2 P

degradation

Keap1 -S-S- Nrf2 -S-S-

Nrf2 pathway Nrf2 pathway

RNA pol II

(25)

NF-B

 Nuclear factor kappa B

 Consists of homo- and heterodimeric complexes formed from the Rel family of proteins

 Five members; p65 (Rel A), p50/p105, p52/p100, c-Rel, and Rel B

 Most common NF-B is a heterodimer of p65 and p50

 Activated by TNF-, IL-1, LPS, viruses, UV, oxidative stress, etc

 Sequestered in the cytosol by IBs

 Involved in chronic inflammatory diseases and cancers

 In most cells, its activation results in cell survival

(Baeuerle and Baltimore 1996, Siebenlist et al 1994, Karin and Ben-Neriah 2000)

(26)

TNF

P P

IB

p50 p65

P P

IB ^p50

p65

IKK

IKK IKK

IKK

NIK

Ub Ub

IB

Proteasomal degradation Proteolysis

p50 p65

NF-B binding site

Ubiquitination Phosphorylation

Cellular membrane

Cytosol

Nucleus

Transcription MEKK1/3

Inflammatory cytokines Oxidative Stress, UV

TNFR

TRADD FADD

TRAF2 RIP

CBP/p300 TFIIB

Ras

PI3K AKT

P

p50 p65

P

Growth factors

p38 Chemopreventive

agents

NF-B

Cell survival (or death ?)

Translation

NF- NF- B pathwayB pathway

P

IB

p50 p65

(27)

그림 11-15

Splicing

: removal of introns

and joining exons 5’ 3’

Guanosine monophosphate (GMP)

 methylation

Poly(A) polymerase 100~200 adenines

AAUAAA primary transcript

(28)

그림 11-16

Guanosine monophosphate (GMP)

 methylation on guanine

(29)

그림 11-17

Poly A tail

nuclease

(30)

그림 11-18

snRNP “snurps”

: small nuclear ribonucleoproteins : proteins + snRNA

: U1, U2, U4, U5, U6

RNA splicing

: removal of introns : by spliceosome

Spliceosome

: macromolecular complex

which removes introns from RNA : proteins + snRNA

(small nuclear RNA)

(31)

그림 11-19

Spliceosome recognizes both ends of the intron and binds to them.

 formation of loop

 Cutting and sticking Recognition : RNA of snRNP

Cutting and sticking: Protein of snRNP

(32)

그림 11-20

formation of loop Splice site

Branch site

(33)

그림 11-21

Alternative splicing : variations in processing mRNA which allow more than on possible protein to be made from a single gene

1. alternative promoter

(34)

그림 11-23

Alternative splicing

2. alternative tail site selection

(35)

그림 11-24

Alternative splicing 3. exon cassette selection

(36)

그림 11-25

Alternative splicing 4. trans-splicing

(37)

그림 11-26

RNA editing

(38)

그림 11-27

Nuclear pore

(39)

그림 11-28

(40)

그림 11-29

40S eukaryotic pre-initiation complex

Branch site

Initiator tRNA

(41)

(42)

그림 11-31

(43)

Regulation of gene expression

• The different cell types of a multicellular organism contain the same DNA.

• Neuron vs lymphocyte

• Cell differentiation depends on changes in gene

expression rather than on changes in the nucleotide sequence of the cell’s genome.

• examples : pancreatic β-cell – insulin hormone,

pancreactic �-cell – glucagon hormone, lymphocyte – antibodies, red blood cell – hemoglobin

• An individual cell specifies its many thousands of genes to express : differentiation

• By regulation of gene expression.

(44)

Gene expression can be regulated at many of the steps in the pathway from DNA to RNA to protein.

(1) controlling when and how often a given gene is transcribed,

(2) controlling how an RNA transcript is spliced or otherwise processed, (3) selecting which mRNAs are exported from the nucleus to the cytosol, (4) selectively degrading certain mRNA molecules,

(5) selecting which mRNAs are translated by ribosomes,

(6) selectively activating or inactivating proteins after they have been made