and Gene Expression
Firman Heru K
Alfiani Umi Farkha
• The nuclear genom contains most of the gene
required for the plant’s physiological functions.
• The first plant genome to be fully sequenced,
in 2000 was that of a small dicotyledonous
angiosperm called thale cress or Arabidopsis
• The genom of A. thaliana is made up of only
about 157 million base pairs (157 Mbp) which
are distributed over five chromosome
• Within its nuclear genome, A. thaliana holds
some 27.400 protein-coding genes and almost
another 5000 genes that are either
pseudogenes (nonfunctional genes) or parts of
The nuclear genome is packaged
• The nuclear genom consists of DNA molecules
that are wrapped around histone proteins to
form beadlike structure called nucleosomes.
• DNA and histone, together with other proteins
that bind to the DNA, are reffered to as
• Two types of chromatin can be distinguished :
euchromatin and heterochromatin
• Heterochromatin is usually more tightly
packaged and thus appears darker than the less
repeats : blocks of nucleotide motifs of about
150 to 180 bp that are repeatd over and over
.• Most genes that are actively transcribed in a
plant are located within the euchromatic
regions of a chromosome. several socalled knobs. while many genes
located in heterochromatic regions are not
transcribed. or chromosome
ends.these genes are inactive or silent
• Compared with euchromatin. Heterochromatic
regions include the centromeres. heterochromatin
is relatively gene poor. known as the subtelomeric
• Heterochromatic structure often consist of
highly repetitive DNA sequence. and the regions immmadiately
adjacent to the telomers.
• A second class of repeats is the
dispered repeats. One types of
dispersed repeats is known as simple
sequence repeats ( SSR) or
• These repeats consist of sequence
motifs as short as two nucleotides
that are repeated hundreds or even
thousands of times.
• Another dominant group of dispersed
repeat found in heterochromatin is the
“jumping genes” or transposons
• Centomers are constrictions of the chromosomes
where sister chromatids adhere to each other and
where spindle fibers attach during cell division
• The attachment of fibers to the centromeres is
mediated by the kinetochore. telomeres. and nucleolar
organizers contain repetitive sequence
• The most prominent structural landmark on
chromosomes are centromeres. and
nucleolar organizers.Centromeres. a protein complex
surrounding the centromere
• Centromere consist of highly repetitive DNA
regions and inactive transposable elements
it is not surprising that
Nos contain hundreds of repeated copies of each rRNA gene
• Depending on the plants species.• Telomeres are sequence located at the ends of each
chromosome. forms a prominent
nuclear structure called the nucleolus
. along with proteins that
transcripts for assembly into ribosomes. Telomeres act as caps on the chromosome ends
and prevents loss of DNA during DNA replication
• The DNA molecules that make up ribosomes (rRNA) are
transcribed from nucleolar organizer (NO) regions. Because
ribosomes are needed for translation. one or several nucleolar
organizers are presents within the genome
• Due to their repetitive nature and their high GC content. Nos
can be seen through a light microscope and thus can serve as
• The rDNA of the nucleolar organizer.
then reserve-transcribed into DNA before it is inserted
elsewhere in the genom
.Transposons are mobile sequence
within the genome
• One dominant type of repetitive DNA within the
heterochromatic regions of the genome is the transposon.
• Transposon or transposable elements are alson known as
“jumping genes” because some of them have the ability to
insert a copy of themselves in a new location within genome
• There are two large classes of transposons : the
retroelements or retrotransposon and the DNA transposons
• These two classes are distinguished by their mode of
replication and insertion into a new location
• Retransposons make an RNA copy of themselves.
splices out the transposon and
insertsit elsewhere in the genome. by contrast. If a
transposon lands within a coding region. move from one position to
another using a cut-and-paste mechanism catalyzed by an
enzyme that is encoded within the transposon sequence
• This enzymes. in most cases keeping
the total transposon copy number the same
• Transposition into a gene can result in mutations. Insertion od a transposon close to a gene can
also alter that gene’s expression pattern
. the gene may be
inactivated. transposase.• DNA transposons.
• Plants and other organisms seem to be able to regulate the
activity of transposons through the methylation of DNA and
• As more genomic noticed large numbers of highly methylated
transposons in heterochromatic region.
• Studies of mutants that are unable to maintain genome
methylation have shown that slow loss of proper methylation
over generations can activate dormant transposons and increase
the frequency of transpositional mutations
• Therefore. methylation and the formationof hererochromatin
appear to play important roles in the stability of the genom
diploid (2n) genome can undergo one or more additional
rounds of replication without undergoing cytokinesis to
• Two forms of polyploidy are distinguished : autopolyploidy
and allopolyploidy. while allopolypoids contain multiple
complete genomes derived from two or more separate
.the number of copies of the entire genome in
a cell-is another important aspect of genome structure that
may have implications for both physiology and evolution
• In many organism.Polyploids contain multiple copies of
the entire genome
• Ploidy level.
• Autopolyploids contain multiple complete genomes of a
single species. but especially in palnts.
resulting zygote contains four copies of each
chromosome and is said to be autotetraploid
if a diploid egg is fertilized by diploid sperm. the chromosomes of a diploid
germ cell undergo DNA replication followed by
two rounds of division
• If chromosome duplication is not followed by cell
devision during meiosis. diploid gametes result
• Within a species or in a self-fertilizing individual.• Both types of polyploidy can result from
incomplete meiosis during gametogenesis.
Allopolyploids usually form in one of
two ways :
1.A haploid sperm from one species
and a haploid egg from another
species may form a diploid
2.Diploid gametes from two different
species may join to form a tetraploid
• Diploid interspecies hybrids occur naturally, but they are
frequently sterile because their chromosome cannot pair
properly during prophase 1 of meiosis
• The lack of fertility in interspecies hybrids is in stark
contast to the phenomenon known as hybrid vigor or
heterosis : the increase vigor often observed in the
offspring of crosses between two inbred varieties of the
same plants species
• Heterosis can contributed to larger plants, greater
biomass, and higher yields in agricultural crops
• Polyploidy can also be induced artificially by treatment
with the natural cell toxin colchicine, which is derived
from the autumn cronus (Colchicum autumnale)
coffee.Phenotypic and physiological responses to polyploidy
Allopolyploids differ from their parental diploid
progenitor species in two major ways :
1. cotton. and
. wheat. like those of autopolyploids. oat.They are hybrid between two different species
Allopolyploids are frequently more vigorous or
higher yielding than their parent species and are
very common among agriculturally important
plantssuch allopolyploids includecanola and
2. rye.Their genomes.
including loss of DNA sequence
• Changes in epigenetic modifications
• Changes in gene transcriptional
• Activation of previously dormant
transposable elements through loss
of gene silencing
.Some of the genetic changes that have been
observed in newly formed allopolyploids compared
with their parent species are the following :
• Restructing of the chromosome.
• Aneuploids are organisms whose genomes
contain more or fewer individual
chromosomes (not entire chromosome
sets) than normal
• Such states are known as trisomies if one
type of chromosome is tripled or
monosomies if only one chromosome of a
given type is presents.
.• Polyploidy is in striking contrast to a
condition called aneuploidy.
• Cytoplasmic genome are probably the evolutionary
remnants the genomes of bacterial cells that were
engulfed by another cell. plant cells contain
two additional genome. postulates that the original
mitochondrion was an oxygen-using bacterium that
was absorbed by another prokaryotic organism.
• The endosymbiotic theory. and the chloroplast genome. which they share with animal
.Plant Cytoplasmic Genomes :
Mitochondria and Chloroplasts
• In addition to the nuclear genome.
like those of procaryotes are
not enclosed in a nuclear envelope and are called
nucleoids. Both mitochondria and chloroplasts are enclosed by
an outer and an inner membrane. and the inner
membrane is continuous with additional membranebound compartments inside the organelle.
2. Both organella genomes show sequence similarity to
procaryotic genomes.• Two main lines of evidence are often cited in support
of the endocymbiotic theory :
. The organelar genomes.
• Organellar genomes consist mostly of linear
similar to the circular plasmids of bacteria.
– Organelar genomes are complex in structure and they
usually consist of multiple copies of the genome on the
same DNA molecul
– Most of the DNA in both plant mitochondria and
chloroplasts is found in linear molecules that my contain
more than one copy of the genome.
– For many years organelar chromosomes had been thought
to contain a genome-sized DNA molecule in circular form.
. but usually
show uniparental inheritance and vegetative segragation. for Angisperms from
maternal parent. Among the
Gymnosperms from paternal parent.
1. Organellar genetics do not obey Mendelian laws. Both mitochondria and plastid generally show uniparental
inheritance ( sexual offspring (via pollen and eggs) only
inherit organelles from one parent).
– The genetics of organellar gene are governed by two
principles that distinguish them from Mendelian genetics.• Organellar genetics do not obey Mendelian laws. Both chloroplasts and mitochondria can segregate
vegetatively ( vegetative cell (as opposed to a gamete) can
give rise to another vegetative cell via mitosis that is
– Regulation of the first step.
• Each step is subject to regulation by the plant to control
the amount of protein that is produced by each gene. includes the control of
trancription initiation. and termination.
– This level of regulation. maintenance. determines
when and whether an mRNA is made. which is referred to as
transcriptional regulation. transcription.Transcriptional Regulation of Nuclear Gene
• The path from gene to protein is a multistep process
catalyzed by many enzymes.
translation efficiency and degradation.
known as posttranscriptional regulation. protein stability (posttranslational regulation)
plays an important role in the overall activity of a gene
or its product.– The next level in the regulation of gene expression. occurs after
transcription includes controls on mRNA stability.
– The region of the gene that binds RNA polymerase is
called the promotor. Core promoter or minimum promoter. which control the activity of the
core promoter. which binds to the DNA to the DNA to be
transcribed and makes an mRNA transcript complementary
to the DNA sequence. consisting of the
minimum upstream sequence required for gene
– Gene transcription is facilitated by an enzyme called RNA
– The structure of the eukaryotic promoter into two part :
2. Regulatory sequence.• RNA polymerase II binds to the promoter region of most
• In addition to RNA polymerase and the general transcription
• These regulatory proteins bind to the DNA and become part of
the transcription initiation complex.
. require specific transcription factors ( also
often called gene regulatory proteins) for RNA polymerase to
become active. most genes. especially those that play important roles
a process mediated by covalent modifications of
both DNA and histones.Epigenetic modifications help determine gene
• Transcription can be initiated only if the DNA is accessible to
the RNA polymerase and other required binding protein.
. its packaging has to be
• Because these modifications can change a gene’s behavior
without changing the DNA sequence of the gene its self.
• To make the DNA accessible. they
are referred to as epigenetic modifications.
determine gene activity. such us methylation of DNA and
methylation and acetylation of histone proteins.• Epigenetic modifications.
• RNA stability can be influenced by cis-elements.
. In order to remain useful as a specific respone to a
– One mechanism by which mRNA stability is regulated
depents on the presence of certain sequence within the
mRNA molecule it sel.Posttranscriptional Regulation of Nuclear Gene
• An organism often produceds mRNA in response to a specific
situation. individual mRNAs must have a finite
– These cis-elements can be bound by RNA-binding
proteins. called cis-elements. which may either stabilize the mRNA or promote
its degradation by nuclease.
dsRNA usually occur as a result of one of
three types of events
– In plant cells.
– Recall that mRNA is usually a single-stranded molecule
– Another mechanism for regulating mRNA stability is the
RNA Interference (RNAi) pathway.
– The RNAi pathway is a set of cellular reactions to the
presence of double-stranded RNA (dsRNA) molecules.• Noncoding RNAs regulate mRNA activity via the RNA
interference (RNAi) pathway.
The presence of microRNAs (miRNAs).
.1. which are involved
innormal developmental processes.
silence certain genes
. The production of short interfering RNAs (siRNAs).2.
The introduction of foreign RNAs. either by viral infection or
via transformation by foreign gene
• The cytoplasmic pathways of protein turnover
involves the ATP-dependent formation of a covalent
bond between the protein that is to be degraded and a
. mRNA stability plays an important
role in the ability of the gene to produce a functional
protein. We turn next to the stability of proteins an the
mechanisms that regulate a protein’s life span.Posttranslational regulation determines the life
span of proteins
• As we have seen. 76-amino acid polypeptide called ubiquitin.
adenylylated ubiquitin is then transferred to a
cysteine residue on a second enzyme. the ubiquitin
conjugating enzyme (E2).• Ubiquitination is initiated when the ubiquitin
activating enzyme (E1) catalyzes the ATP dependent
adenylylation of the C terminus of ubiquitin. a
ubiquitin ligase (E3)
. Proteins destined for
degradation are bound by a third type of protein.
analysis of mutants is an extremely powerful
tool that can help scientists infer the function
of a gene or map its location on the
chromosoms.Tools for Studying Gene Function
• Individuals that contain specific changes in
their DNA sequence are called mutants.
so the change in the
mutant’s nucleotide sequence must result in an
altered phenotype.Mutant analysis can help to elucidate gene
• The use of mutants for gene identification
relies on the ability to distinguish a mutant
from a normal individual.
explain a method called map
based cloning.• There are several ways to map a mutation to
its chromosome and ultimately clone the
affected gene. which uses crosses between a
mutant and a wild type palnt and genetic
analysis of the offspring to narrow down the
location of the mutation to a short segment of
. which is then sequenced.
a gene may be expressed only in
reproductive tissues. such as
a glass slide. onto which DNA sequences are spotted
that are representative of single genes of a given
• All microarray techniques use a solid support. or only I vegetative ones.Molecular techniques can measure the
activity of genes
• Once a gene of interest has been identified.
• For example. scientists
are usually interested in where and when the gene is
Gene fusions can introduse reporter
• A gene fusions is an artificial
construct that combines part of the
gene of interest with another gene
that produces a readily detectable
• A gene’s expression is regulated by
transcription factors that fine-tune its
activity and allow it to be trancsribed
only where and when its needed.
bioengineering allows the
transfer of specific gene ora gened
between spiceis that cannot be
.Genetic modification of crop plants
• In contrast to classical selective
• Plants produced in this way are
commonly referred to as genetically
modified organisms (GMOs)
.• Biotechnological tools circumvent
this problem by allowing insertion of
only the desired genes into the
recipient plant. most often either by
transformation or by biolistics.
not just those with which the
recipient can be succesfully crossed
• GMOs may carry gene constructs that are the
product of splicing a variety of genetic
components together to produce genes with
altogether new functions (for example.There are three essesntial differences between
GMOs and conventionally bred varieties of crops
• Gene transfer into GMOs occurs in the
laboratory and does not require crossbreeding
• The donor genes of GMOs can be derived from
any organism. the
promoter-GFP fusion genes we described
Transgenes can confer resistance to
herbicides or plant pets
• Any gene articially tranferred into an
organism is referred to as a
• Plants carrying a transgene for
glyphosate resistance will survive a
field application of glyphosate (the
commercial herbicide Roundup).
which kills weeds but does not harm
resistant crop plants.
• Another commonly used transgene
encodes an inseticidal toxin from th
soil bacterium Bacillus thuringiensis
. look on GMOs with
suspicion and cocern. In spite of
their enoumous humanitarian
potential. many individuals.Genetically modified organisms are
• The development of GMOs has not
been greeted with universal
enthusiasm and support . as well
as the government of some