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Chromosomes Powered By Docstoc
					Major types of mutations
              Major types of mutations
Basis of             Mutation type      Major features
Origin               Spontaneous        Absence of known mutagen
                     Induced            Presence of known mutagen
cell type            Somatic            Non-reproductive cells
                     Germ-line          Reproductive cells
expression           Conditional        Under restrictive conditions
                     Unconditional      Under permissive conditions

Effect on function   Loss-of-function   Eliminating normal function
                     Hypomorphic        Reducing normal function
                     Hypermorphic       Increasing normal function
                     Gain-of-function   Expressed at incorrect time
                                        or in inappropriate cell type
              Major types of mutations
Basis of           Major types    Major features

Molecular change   Transition     Pyrimidine (T,C) to Pyrimidine
                   Transversion   Pyrimidine to purine (or opposite)
                   Insertion      One or more nucleotides present
                   Deletion       One or more missing nucleotides

Effect on          Silent         No change in amino acid
translation        Missence       Change in amino acid
                   Nonesense      Creating stop codon
                   Frameshift     Shift reading of codon incorrectly
• Example for conditional mutation is temperature-sensitive
Usually the restrictive temperature is high (29°indrosophila)
and the phenotype is mutant above it. The permissive
temperature is low (18° in drosophila) and the phenotype is
wildtype under it.
• loss-of-function is also called knock out or null mutation.
• A common type of frameshift mutation is a single-base
   addition or deletion.
             CUG CUG CUG
              Leu Leu Leu
             CUG CAU GCU G
             Leu His Ala
        Example (PAH)
    phenylalanine hydroxylase
     CGG = codon on mRNA for Argenine (AA)
   UGG = codon on mRNA for Tryptophan (AA)
             wild type              mutant
 DNA: 5’ ……CGG……            .…..TGG……3’

DNA(T): 3’……GCC……                .…..ACC……5’

 mRNA: 5’……CGG……                  ……UGG......3’
So the Aa (Argenine) will be replaced by the Aa (
      Tryptophan), that means mutant protein.
         Fragile-X syndrome
• X-linked form of mental retardation shows
  dynamic mutations. The chromosome X
  tends to fracture near the end of the long
  arm and it is called fragile-X and the
  disease is called fragile-X syndrome.
• This syndrome affects 1 in 2500 children
  and is second only to Down syndrome as
  a cause of inherited mental impairment.
• Males are more affected than females.
• The molecular basis is trinucleotide
  repeat of the form CGG (or CCG on the
  other strand) in the part where the
  breakage takes place.
• Normal X-chromosome has 6 to 54
  repeating unit. Affected persons have
  230 to 2300.
• The premutation has an intermediate
  number of copies ranging from 54 to
• The premutation increases in copy number
  when transmitted through only females and
  is called trinucleotide expansion. This occurs
  not in germ line but in somatic cells of the
  early embryo.
• Amplification to 230 copies or more causes
  silence to the gene FMR1 (fragile-mental
  retardation-1) which is expressed usually in
  brain and testes.
• Different extent of amplification in different
  somatic cells accounts for the variation in
  severity of this syndrome among affected
• The molecular mechanism of trinucleotide
  expansion is a process called replication
  slippage (also called slipped-strand
• The mechanism of inactivation of FMR1 is
  Methylation of Cytosine in the full mutation.
• FMR1 protein regulates translation of some
  mRNA related to development of facial
  bones and the nervous system. FMRP also
  functions in learning and memory.
   DNA damage can be repaired
• Mismach-repair system fixes incorrectly matched
  base pairs.
• AP endonuclease system repairs nucleotide sites at
  which the base has been lost (Apyrimidine site and
  apurine site).
• Special enzymes repair damage caused to DNA by
  ultraviolet light.
• Postreplication repair skips over damaged bases.
               Cell cycle
The cell cycle is divided into three-part
   interphase composed of G1 (gap1), S
   (DNA synthesis), and G2 (gap2),
   followed by M (mitosis).
          Mitosis functions
The essential functions of mitosis are:
1) Replication of DNA once per cycle.
2) Distribution of the replicas (the sister
   chromatids) equally to the two daughter
Methods to study cell cycle control
• Genetic control has been approached via
  the methods of biochemistry, cell biology,
  and genetics.
• Budding yeast, Saccharomyces cerevisiae
  (S. cerevisiae) is good example for
  experiments on cell cycle control.
• It has 5538 genes and we can analyze the
  transcription pattern by gene microarray.
Cell division cycle (cdc) mutants
• Cdc mutant is a mutant whose phenotype
  is to arrest the cell cycle at a specific point.
• Cdc mutants are wildtype at 23° ( the
  permissive temperature) and unable to
  complete cell cycle at 36° ( the restrictive
• Cdc13 causes arrest at the G2/M
  boundary because of a defect of telomere
          Cdc13 mechanism
• So after the temperature shift, single cells
  or cells with small buds will give a pair of
  large cells configuration.
• Cells which were nearing the end of
  division and had large buds will give
  quartets after the temperature shift.
      Cyclins and cyclin-CDK
• In the early stages of cell cycle,
  progression from one phase to the next is
  controlled by characteristic protein
  complexes that are called cyclin-CDK
• They are composed of cyclin subunits
  combined with cyclin-dependent protein
  kinase (CDK) subunits.
        Routes of regulation
1) Phosphorylation:
• When cyclin subunit binds to the protein
  substrate, the CDK component
  phosphorylates the substrate.
• Phosphorylation activates some proteins
  and inactivate others.
• Phosphorylation of different site might
  inactivate the same activated protein.
       2) Dephosphorylation
Phosphatase enzymes dephosphorylate
proteins that CDKs have phosphorylated,
reversing the effects of CDKs.
  Some important Cyclin-Cdks
Cyclin D-Cdk4 and cyclin D-Cdk6complexes
appear in the early or middle part of G1.
They promote entry into S phase.
Cyclin A-Cdk2 and cyclin E-Cdk2 appear
later in G1.
Cyclin B-Cdk2 complex carries the cell
through the G2/M transition.
  Retinoblastoma (RB) protein
The normal role of RB is to maintain cycling
cells at a point in G1 called the G1
restriction point or start until the cell has
attained proper size.

RB binds to the transcription factor E2F
which is needed for further progression in
the cell cycle.
RB is phosphorylated by D-Cdk4,6, E-Cdk2
kinase as cells approach G1/S transition.

RB phosphorylation eliminates its ability to
bind the E2F transcription factor.

Release of E2F results in transcription of
enzymes responsible for DNA synthesis
and E2F itself (as positive autoregulation).
Through S-phase, E2F is phosphorylated by
cyclin A-Cdk2 and inhibits it binding to
DNA, so inactivating its function as a
transcription factor.

Cyclin B-Cdk2 complex (also called
maturation promoting factor) controls the
progression from G2 to M transition.
 Protein degradation (proteolysis)
Cell cycle go forward by protein degradation
that complements the periodic activation
of cyclin-CDK complexes.
Progression of cell cycle needs destruction
of previous proteins.
Protein degradation entails:
-Sister chromatids to separate.
-Cell to exit from mitosis.-
 Checkpoints allow repair or death
A DNA damage checkpoint. (G1/S
A centrosome duplication checkpoint. (G2/M
A spindle checkpoint. (Anaphase
 P53 for DNA damage checkpoint

DNA damage checkpoint acts via three
stages in the cell cycle:
G1/S transition
S period
G2/M boundary
      P53 transcription factor
It is responsible to stress in general, and to
DNA damage in particular.
In normal cells, activated p53 protein level is
very low. However, p53 mRNA and p53
protein are present.
P53 must go phosphorylation and
acetylation to be active .
            Mdm2 and p53
The protein Mdm2 inactivates p53 by
preventing it from phosphorylation and
subsequent steps of its activation.

Activated P53 triggers the transcription of a
number of genes and the repression of
      Genes activated by p53
14-3-3 σ which plays a role to arrest cell at
G2/M boundary.
P21 which plays a role at G1/S transition
and S checkpoint.
Apaf1 and Bax which promote apoptosis.
Maspin which inhibits angiogenesis
(formation of blood vessels) and
Means programmed cell death. (suicide)
DNA damage also triggers activation of the
pathway of apoptosis.
A cascade of proteins involved in the lysis of
the cell initiate suicide.
They are called caspases.
Oncogene is a gain-of-function mutation in a
cellular gene, called proto-oncogene,
whose normal function is to promote
proliferation or inhibit apoptosis;
oncogenes are often associated with
tumor progression.
       Oncogenes and Bcl2
Oncogenes can increase the level of
activated (phosphorylated) Bcl2, which
prevents apoptosis and allows the affected
cells to grow and divide indefinitely.
    Tumor suppressor genes
Tumor suppressor gene is a loss-of-function
mutation in a cellular gene, whose normal
function is to inhibit cell division or to
activate apoptosis.
      Tumor suppressor genes
Tumor suppressor alteration    consequence
p53              mutation Loss of G1/S, S, and
                            G2/M checkpoint
p21              mutation Loss of G1/S and S
RB               mutation Promote proliferation;
                            E2F uninhibited
Bax              mutation Failure of apoptosis
Centrosome duplication checkpoint
   and the spindle checkpoint
Centrosome is the organelle around which
the bipolar spindle is organized.
Failure in centrosome duplication checkpoint
might result in polyploidy.
Failure in spindle checkpoint might cause
APC is the anaphase-promoting complex
needed for entry into anaphase.
Correct attachment to the spindle causes
inactive spindle checkpoint (inactive Bub,
Mad, and Mps) and so, active APC.
Incorrect or unbalanced attachment to the
spindle activates spindle checkpoint and
so, inactive APC.
       Loss of heterozygosity
Loss of the presence of the wildtype allele,
or loss of its function, in a heterozygous
cell, enabling the phenotype of a recessive
mutant allele to be expressed.

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