Lewin Benjamin (ed.) Genes IX

Genes

Types

RNA-coding

165 rRNA

23S rRNA

4.5S rRNA

55

rRNA

tRNA

Gene Expression

r-proterns

RNA

polymerase

Others

1

30-32

20-21

z

Chloroplast

functions

Rubisco

and thylakoids

31-32

NADH

dehydrogenase

11

Total 105-113

Fil;"ift{::

i.

.i !:;

The chloroptast

genome

in

[and

ptants

codes

for

4

rRNAs,

30 tRNAs, and

-60

proteins.

i

iirL'

ii

lr

.1.

!

ij The mitochondriaI genome

of S. cerevisi

oe

contains

both interrupted

and

uninterrupted

protein-

coding

genes,

rRNA

genes,

and IRNA

genes

(positions

not indicated).

Anows indicate

direction

of transcription.

The

map shown in

:

I{ri-{{iti +.;

ii

accounts

for

the major RNA

and

protein

products

of the

yeast

mitochondrion. The

most notable

feature

is the

dispersion

of

loci

on the

map.

The

two most

prominent

loci

are

the

interrupted

genes

box

(coding

f or

cytochrome

bl and oxi3

(coding

for

subunit I

of cytochrome oxidase).

Together

these

two

genes

are almost as long

as the

entire mito-

chondrial

genome

in

mammals!

Many

of the

Iong introns in

these

genes

have

open read-

ing frames in

register with

the

preceding

exon

(see

Section 27.5.

Some Group

I Introns

Code

for Endonucleases

That

Sponsor Mobility).

This

adds several

proteins,

all

synthesized in

Iow

amounts. to

the complement

of the

yeast

mitochondrion.

The remaining genes

are uninterrupted.

They

correspond to

the other

two subunits

of

cytochrome

oxidase coded

by the mitochon-

drion, to the

subunit(s)

of the ATPase,

and

(in

the case of varl)

to a mitochondrial

ribosomal

protein.

The

total numtrer

of

yeast

mitochon-

drial

genes

is unlikely

to exceed

-25.

oxi2

8,4 kb

pal

fl

Exons

fft

Intr,rns

o/l

I

=

subunits ol

oliqomvcrn-sensitive

aapJ

ATPase

oxi

=

subunits of

cytochrome

c

Dox

=

cytochrome

b

par

unknown lunctions

var

=

small ribosome

subunit

orotein

The Chloroplast Genome

Codes

for Many Proteins

and RNAs

.

Chloroptast

genomes

vary in size, but are [arge

enough

to code

for

50

to 100

proteins

as

wetl

as

the rRNAs

and tRNAs.

What

genes

are carried by chloroplasts?

Chloro-

plast

DNAs vary in length from

120

to

190 kb.

The sequenced

chloroplast

genomes (>10

in

total) have

87 to 183

genes.

Flil*qi.

'r,-l*

summa-

rizes the functions coded by the chloroplast

genome

in land

plants.

There

is more

variation

in the chloroplast

genomes

of algae.

The

situation

is

generally

similar to that of

mitochondria,

except

that more

genes

are

involved.

The chloroplast

genome

codes for all

the rRNA

and IRNA species

needed

for

protein

slmthesis. The ribosome includes

two small rRNAs

in addition

to

the major species.

The IRNA set

may indude

all of the

necessary

genes.

The

chloro-

plast genome

codes for

-50

proteins,

including

RNA

polymerase

and ribosomal

proteins.

Again,

the rule is

that organelle

genes

are transcribed

and translated by the apparatus

of the organelie.

About half

of

the chloroplast

genes

code for

proteins

involved in

protein

synthesis.

The

endosymbiotic

origin

of the chloroplast

is empha-

sized by the relationships

between these

genes

and their

counterparts

in bacteria.

The organi-

zation of the rRNA

genes

in

particular

is closely

related to that of a cyanobacterium,

which

pins

down more

precisely

the

last common ancestor

between

chloroplasts

and bacteria.

4.1.2 The Chtoroplast

Genome Codes

for Many Proteins and RNAs

7t

Introns in

chloroplasts

fall into two

general

classes. Those in IRNA

genes

are usually

(although

not inevitably) located

in

the

anti-

codon loop,

like

the

introns found in

yeast

nuclear IRNA

genes

(see

Section

26.I4,

yeast

IRNA

Splicing

Involves Cutting and

Rejoining).

Those in

protein-coding genes

resemble the

introns

of

mitochondrial

genes

(see

Chapter

27,

Catalytic

RNA). This

places

the endosymbiotic

event at a time

in

evolution

before the separa-

tion of

prokaryotes

with uninterrupted

genes.

The role of the chloroplast is to undertake

photosynthesis.

Many of

its

genes

code

for

pro-

teins

of complexes

located in the thylakoid

membranes. The constitution of

these com-

plexes

shows a different balance from that

of

mitochondrial complexes. Although some com-

plexes

are

like mitochondrial complexes in hav-

ing some

subunits

coded by the organelle

genome

and some by the

nuclear

genome,

other

chloroplast complexes are coded entirely

by one

genome.

@

Mitochondria Evolved

by

Endosymbiosis

How did a

situation

evolve in which an organelle

contains

genetic

information for

some

of its

functions,

whereas others are coded

in the

nucleus?

f

l{-ii+:i.

.:;.=+

shows

the

endosymbiosis

model

for mitochondrial evolution,

in

which

primitive

cells captured bacteria that

provided

the functions that evolved into mitochondria

and chloroplasts. At this

point,

the

proto-

organelle

must have

contained all of the

genes

needed

to specify its functions.

Sequence

homologies

suggest that

mito-

chondria

and chloroplasts evolved separately,

from lineages

that are common with eubacte-

ria,

with mitochondria sharing an origin with

cr-purple bacteria and chloroplasts sharing an

origin with cyanobacteria. The closest known

relative

of mitochondria among the bacteria is

Rickettsia

(the

causative agent of typhus), which

is

an obligate intracellular

parasite

that is

prob-

ably descended from free-living

bacteria.

This

reinforces

the

idea

that

mitochondria

originated

in an endosymbiotic event involving

an ances-

tor that is

also common to Rickettsia.

Two

changes must have

occurred

as the

bacterium became

integrated

into the recipient

cell and

evolved

into

the mitochondrion

(or

chloroplast). The

organelles have far fewer

genes

than an

indenendent

bacterium and have lost

IItiJ*il +.i*

Mitochondria originated

by a endosymbi-

otjc event when a bacterium

was

captured by a eukary-

otic ce[t.

many of the

gene

functions that

are

necessary

for independent

life

(such

as metabolic

path-

ways).

The majority of

genes

coding

for

organelle functions are

in fact

now located in

the nucleus, thus these

genes

must have

been

transferred there

from

the organelle.

Transfer

of

DNA

between organelle and

nucleus has occurred over evolutionary time

periods

and still continues.

The rate

of transfer

can be measured directly by introducing into

an organelle a

gene

that can function

only in the

nucleus, for

example,

because it

contains a

nuclear intron,

or

because the

protein

must

function

in

the cytosol.

In

terms of

providing

the material for evolution, the

transfer rates

from organelle to nucleus are roughly

equiva-

lent

to

the rate

of single

gene

mutation. DNA

introduced into mitochondria is transferred

to

the

nucleus

at a

rate

of

2

x

l0-5

per

generation.

Experiments to measure transfer in the reverse

direction,

from nucleus

to mitochondrion,

sug-

gest

that the rate is a much lower

<I0-10.

When

Bacterium evolves into

mitochondrion,

losing

genes

that

are

necessary for independent

lite

72

CHAPTER 4 The

Content of the Genome

a nuclear-specific

antibiotic

resistance gene

is

introduced

into

chloroplasts,

its

transfer

to the

nucleus

and successful

expression

can

be

followed

by screening

seedlings

for

resistance

to

the

antibiotic.

This

shows

that transfer

occurs

atarate

of I in 16,000

seedlings,

or

6

x

l0-5.

Transfer

of a

gene

from

an

organelle

to the

nucleus

requires physical

movement

of

the

DNA,

of course,

but successful

expression

also

requires

changes

in

the

coding

sequence.

Organelle

proteins

that

are coded

by nuclear

genes

have

special

sequences

that

allow them

to be imported

into

the

organelle

after

they have

been

synthesized

in the

cytoplasm

(see

Section

10.16,

Posttranslational

Membrane

Insertion

Depends

on

Leader

Sequences).

These

sequences

are not required

by

proteins

that

are

synthesized

within

the

organelle.

perhaps

the

process

of effective gene

transfer

occurred

at

a

period

when

compartnlents

were less

rigidly

defined,

so that it

was easier

both

for the DNA

to

be relocated

and for

the

proteins

to be incor-

porated

into the

organelle

irrespective

of the

site of synthesis.

Phylogenetic

maps show

that

gene

transfers

have

occurred independently

in

many

differ-

ent lineages.

It appears

that

transfers

of mito-

chondrial

genes

to the

nucleus

occurred

only

early in

animal

cell evolution,

but it

is

possible

that

the

process

is

still continuing

in

plant

cells.

The

number

of transfers

can

be large;

there are

>800

nuclear

genes

in Arabidopsis

whose

sequences

are related

to

genes

in

the chloro-

plasts

of other

plants.

These

genes

are candi-

dates for evolution

frorn

genes

that originated

in

the chloroplast.

Sum mary

The

DNA sequences

composing

a eukaryotic

genome

can

be classified

in three

groups:

.

nonrepetitive

sequences

are

unique;

.

moderately

repetitive

sequences

are

dispersed

and repeated

a small

number

of

times

as

copies;

.

and highly

repetitive

sequences

are short

and usually repeated

as tandem

arrays.

The

proportions

of the types

of sequences

are

characteristic for

each

genome,

although

larger

genomes

tend to

have a

smaller

propor-

tion

of nonrepetitive

DNA.

Almost

50% of the

human

genome

consists

of repetitive

sequences,

the vast majority

corresponding

to

transposons

sequences. Most

structural

genes

are located in

nonrepetitive

DNA. The

amount of nonrepeti-

tive DNA

is a better reflection of the

complex-

ity

of the

organism than the total

genome

size;

the

greatest

amount of nonrepetitive DNA is

genomes

is

-2

X

l0e bp.

Non-Mendelian inheritance is

explained

by the

presence

of

DNA in

organelles

in

the

cytoplasm.

Mitochondria and chloroplasts are

membrane-bounded

systems in which some

proteins

are

synthesized

within the

organelle,

while

others are imported. The organelle

genome

is

usually a circular

DNA

that codes

for

all

the RNAs and some of the

proteins

required

by the organelle.

Mitochondrial

genomes

vary

greatly

in

size

from

the l6 kb minimalist mammalian

genome

to the 570 kb

genome

of

higher

plants.

The

Iarger

genomes

may code for additional func-

tions.

Chloroplast

genomes

range from 120-200

kb. Those

that have been sequenced have sim-

ilar

organization

and coding

functions. In both

mitochondria

and chloroplasts, many of the

major

proteins

contain some subunits synthe-

sized

in the organelle and some subunits

imported

from

the cytosol.

Rearrangements

occur

in mitochondrial

DNA rather

frequently in

yeast,

and

recombi-

nation

between mitochondrial

or between

chloroplast

genomes

has been found. Transfers

of DNA have

occurred

between chloroplasts or

mitochondria

and

nuclear

senomes.

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Genomes

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rc

h

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A.,

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J.,

and Donnelly,

P.

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@l

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@

Reviews

74 CHAPTER

4

The

Content

of the Genome

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Proteins

and

RNAs

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References

75

Genome

Sequences

and Gene

Numbers

CHAPTER OUTLINE

Introduction

Bacteria[

Gene

Numbers

Ranqe 0ver an

Order

of

Magnitude

Total

Gene

Number Is Known for

Several

Eukaryotes

.

There

are 6000

genes

in

yeasU

18.500

in

a

worm; 13,600 in

a

fty; 25,000 in

the sma[[

plant

Arabidopsrs,'

and

probabty

20,000

to 25,000 in

mouse

and

man.

How Many Different Types

of Genes

Are There?

The Human

Genome Has Fewer Genes Than

Expected

r

0nty 1% of the human

genome

consists

of coding

regions.

r

The exons

comprise

-5olo

of each

gene,

so

genes

(exons

ptus

introns)

comprise

-25olo

of the

genome.

.

The human

genome

has 20,000 to 25.000

genes.

o -60olo

of human

genes

are atternatively spticed.

.

Up to 800/o

of the atternative sptices change

protein

sequence, so the

proteome

has

-50,000

to 60,000 members.

How Are

Genes and 0ther Seouences

Distributed in the

Genome?

.

Repeated sequences

(present

in more than one copy)

account

for >50%

of the human

genome.

.

The

great

bulk of repeated sequences consist of copies of

nonfunctjonaI

transposons.

.

There are many

duplications of large chromosome

regions.

The Y

Chromosome Has

Several

Male-Soecific Genes

o

The Y chromosome has

-60

genes

that are expressed specifi-

catty

in

the testis.

.

The mate-specific

genes

are

present

in muttiple copies in

repeated chromosomaI

segments.

r

Gene conversion

between

muttipte

copies attows the active

genes

to be majntained during

evotution.

More

Comptex Species Evotve

by

Adding New

Gene

Fu nctio ns

.

Comparisons of djfferent

genomes

show a steady

increase in

gene

number

as additional

genes

are added to make eukary-

otes, mutticellutar

organisms, anima[s. and vertebrates.

o

Most

of

the

genes

that are

unique to vertebrates are

con-

cerned

with the

immune or nervous systems.

How

Many

Genes

Are

Essential?

.

Not a[[

genes

are

essential. In

yeast

and

fty. detetions of

<50% of the

genes

have detectable effects.

.

When two or

more

genes

are

redundant, a mutation'in any

one of them

may not have detectable

effects.

o

We

do

not futty understand

the survival

in

the

genome

of

genes

that

are apparently dispensable.

Genes

Are Expressed at Widel.y

Differing Levets

.

In any

given

ce[t,

most

genes

are expressed at

a low [eve[.

.

Onty

a smatl number of

genes,

whose

products

are speciat-

ized for

the

ce[[ type. are

high[y expressed.

How Many Genes

Are Expressed?

r

mRNAs

expressed

at low levets overlap

extensively when dif-

ferent

cetl

types are compared.

r

The abundantty

expressed

mRNAs

are

usuatly specific for the

cett type.

.

-L0,000

expressed

genes

may

be common to

most ce[[ types

of a

higher eukaryote.

Expressed Gene

Number Can

Be Measured En Masse

.

"Chip"

technology

altows a snapshot

to be taken of the

expression

of the entire

genome

in a

yeast

ce[[.

t

-75olo

(-4500

genes)

of the

yeast

genome

is expressed

under normaI

growth

conditions.

.

Chip

technology a[tows detailed

comparisons of retated ani-

mal

celts to

determine

(for

exampte) the differences

in

exoression between

a normal cetl and a cancer ce[t.

Summary

76

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40,000

o

30,000

o

zo,ooo

10,000

300

400 500

Genome size

(Mb)

homology

with known genes

in

other species.

These

genes

fall approximately

equally into

classes whose

produr:ts

are concerned

with

metabolism,

cell struclure

or

transport of

com-

ponents,

and

gene

expression

and its regulation.

In virtually

every

genome,

>25o/o

of the

genes

cannot

be ascribed any

function.

Many

of these

genes

can

be

found

in related

organisms, which

implies

that they have

a conserved

function.

There has

been sorne

emphasis

on sequenc-

ing the

genomes

of

pathogenic

bacteria,

given

their medical

importarLce.

An important

insight

into

the nature

of

pathogenicity

has

been

pro-

vided

by the demonst.ration

that

"pathogenic-

ity

islands" are

a characteristic

feature

of their

genomes.

These

are large

regions,

-10

to 200 kb,

which are

present

in the

genome

of a

patho-

genic

species

but absent

from the

genomes

of

nonpathogenic

variants

of the same

or related

species. Their

G-C

content often

differs from

that

of the

rest

of the

genome,

and it

is likely that

they migrate

between

bacteria

by a

process

of

horizontal

transfer.

For example,

the bacterium

that causes

anthrax

(Bacillus

anthracis)

has two

large

plasmids

(extrachromosomal

DNA),

one

of

which

has

a

pathoge:nicity

island

thar includes

the

gene

coding for

the anthrax

toxin.

i

ir,iiirli:

,

The number of

genes

in

a eukaryote

varjes from 6000

to 40,000

but does

not corretate

with

the

genome

size or the com-

ptexity

of the organism.

rirlL.:i'r:';r.'r

TheS.cerevisiaegenomeofl3.5Mbhas6000genes.almosta[[unin-

terrupted. Ihe 5.

ponbe

genome

of 12.5

Mb has

5000

genes,

almost

half having

introns.

Gene sizes and spacing

are

fairly

simjlar.

The

major difference

between them

is

that

only

5t/. of. S. cerevisiae

genes

have introns, compared

to 43o/o in

S.

pombe.

The density of

genes

is high;

organization is

generally

similar,

although the

spaces between

genes

are a bit shorter

in S. cere-

visiae.

About half of

the

genes

identified by

sequence were either

known

previously

or

known

genes. The remainder are

new

which

gives

some

indication of the

number of

new

types of

genes

that may be

discovered.

The

identification

of long

reading frames on

the basis of sequence

is

quite

accurate.

How-

ever, ORFs coding

for

<100

amino

acids cannot

be identified solely by sequence

because of

the

high

occurrence

of false

positives.

Analysis

of

gene

expression suggests

that

-300

of

600 such

ORFs in

S.

cerevisiae are

likely to be

genuine genes.

A

powerful

way to validate

gene

structure

is to

compare

sequences

in closely

gene

is active, it

is likely to be con-

served. Comparisons between

the sequences

of

four closely

species

suggest that

503 of the

genes

originally

identified

in

5%

oi S. cerevisiae

genes

have 1 intron on average

43"koI

S.

pombegenes

have

introns

Average

interrupted

gene

has 2 introns

\

Total

Gene Number

Is

Known for

Several

Eukaryotes

.

There

are 6000

genes

in

yeas|

18,500 in

a worm;

13,600 in

a

fty;

25,000 in

the smat[

ptant

Arobidopsis;

and

probably

-25,000

in

mouse

ano

man.

As

soon as we look

at eukaryotic

genomes,

the

relationship

between

genome

size

and

gene

number is lost. The

genomes

of unicellular

eukaryotes fall in

the same

size

range

as the

largest

bacterial

genornes.

Higher

eukaryotes

have more

genes,

but the number

does not cor-

relate with

genome

size,

as can be seen from

tlti.:-stl:

,

The most extensive

data for lower

eukary-

otes

are available frorn

the sequences

of the

genomes

of the

yeasts

Saccharomyces

cerevisiae

and Schizosaccharomyce,s

pombe. i'!r;rr*r

r,

:r sum-

marizes

the most important

features. The

yeast

genomes

ol12.5 Mb and 13.5

Mb have

-6000

and

-5000

genes,

respectively.

The

average open

reading frame

(ORF)

is

-1.4

kb, so that

-70o/o

of the

genome

is occupied

by coding regions.

5.3

Total Gene

Number

Is Known for Several

Eukaryotes 79

'

-20olo

of DrosophiLa

genes

code for

proteins

concerned with majntaining or expressing

genes,

-20%

code

for

enzymes, and

<100/o

code

for

proteins

concerned

with the

cet[ cycle or signal transductjon.

Ha[f

ofthe

genes

of DrosophiLa

code

for

products

of unknown

function.

S. cerevisiae do not have counterparts in the other

species and therefore should be deleted from

the catalog. This reduces the total

gene

number

f,or S. cerevisiae

to

5726.

The

genome

oI Caenorhabditis elegans DNA

varies between regions rich in

genes

and regions

in which

genes

are

more

sparsely organized.

The

total sequence contains

-18,500

genes.

Only

-42oh

of the

genes

have

putative

counter-

parts

outside the

Nematoda.

Although the fly

genome

is larger

than the

worm

genome,

there are

fewer

genes (13,600)

in D.

melanogaster.The number of different tran-

scripts is

slightly

larger

(

14,

100) as the result of

alternative splicing. We do not understand why

the fly-a much more complex organism-has

only

70o/o

of the number of

genes

in the worm.

This emphasizes forcefully

the

lack

of an exact

relationship

between

gene

number and com-

plexity

of the

organism.

The

plant

Arabidopsis thaliana

has a

genome

size intermediate

between the worm and the

fly, but has a larger

gene

number

(25,000)

than

either. This again

shows the

lack

of a clear rela-

tionship

and also emphasizes the

special

qual-

ity

of

plants,

which may have more

genes (due

to ancestral

duplications) than animal cells. A

majority

of. the Arabidopsis

genorr'e

is found in

duplicated

segments, suggesting that

there was

an

ancient doubling of the

genome (to give

a

tetraploid).

Only 35ok oI Arabidopsls

genes

are

present

as single copies.

The

genome

of rice

(Oryza

sativa)

is

-4larger

than Arabidopsis,

but the number

of

genes

is

only

-50%

larger,

probably

-40,000.

Repeti-

tive DNA

occupies 42o/o-45oh

of the

genome.

More than 80%

of the

genes

lotndinArabidop-

CHAPTER 5 Genome

Sequences and

Gene

Numbers

sls are represented

in rice. Of these

common

genes,

-8000

are found

in Arabidopsrs

and

rice

but not in any of

the

bacterial or animal

genomes

that have been sequenced. This is

prob-

ably

the

set

of

genes

that codes for

plant-

specific

functions, such as

photosynthesis.

From

the

fly

genome,

we can form an

impression of how

many

genes

are devoted to

each type of

function. i l,ili.iii:

:..r

breaks down

the

functions into different categories. Among

the

genes

that are

identified, we find 2500

enzymes,

-750

transcription

factors,

-700

trans-

porters

and ion channels, and

-700

proteins

involved with signal

transduction. Just

over

half

of

the

genes

code for

products

of unknown func-

tion. Approximately

2oo/o of the

proteins

reside

in membranes.

Protein size increases

from

prokaryotes

and

archaea

to eukaryotes. The archae a M.

jannaschi

and bacterium

E.

coli

have

average

protein

lengths

of

287 and 317 amino acids, respec-

tively, whereas S. cerevisiae and C elegans

have

average lengths oI 484 and

442

amino acids,

respectively. Large

proteins

(500

amino acids)

are rare

in

bacteria, but comprise a significant

component

(-l

/3) in eukaryotes. The increase

in length is due to the addition of extra domains,

with each domain typically constituting 100-300

amino acids. The increase in

protein

size, how-

ever, is responsible for only a very small

part

of

the

increase in

genome

size.

Another insight

into

gene

number is

obtained by counting

the number

of expressed

genes.

If we rely upon the estimates of the num-

ber of different mRNA species that can

be

counted in a cell, we would conclude that the

average vertebrate cell expresses

-I0,000

to

20,000

genes.

The existence of significant

over-

laps

between the

messenger

populations

in dif

-

ferent cell types would suggest that the total

expressed

gene

number for

the organism should

be within a

few fold

of this amount. The esti-

mate for the total human

genome

number

of

20

to 25,000

(see

Section

5.5, The Human

Genome

Has Fewer Genes Than Expected) would imply

that a

significant

proportion

of the total

gene

number is

actually expressed in any

given

cell.

Eukaryotic

genes

are transcribed individu-

ally, with each

gene producing

a monocistronic

messenger. There is

only one

general

exception

to this rule: in the

genome

of

C. elegans,

-l5oh

of the

genes

are organized into

polycistronic

units

(which

are associated with the

use oltrans-

splicing to allow expression of

the downstream

genes

in these units; see Section 26.I3,

trans-

splicing Reactions

Use Small RNAs).

ffi DNA manipulation

I

Transcription

n

Translation

!

Protein

structure

I

Cell

cycle/death

E

Cytoskeleton

I

Enzymes

tr

Signal transduction

I Cell adhesion

I

Transporters

/channels

tr

Unknown

80

Lewin Benjamin (ed.) Genes IX

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