Endosymbiotic gene transfer: organelle genomes forge eukaryotic chromosomes'

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Endosymbiotic gene transfer organelle genomes
forge eukaryotic chromosomes.

• J.N. Timmis, M.A. Ayliffe, C.Y. Huang and W.
  Marting, 2004. Nature Reviews, 5 123-135.

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Eukaryotic nuclear genomes have been the
recipient of mitochondrial and chloroplast DNA
donations.
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-non-Mendelian mode of inheritance -multiple
circular haploid (single chromosome or set of
chromosomes, single allele) genomes
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Organelle genomes prokaryotic remnants

• 1970s Notion that organelles originated from
  endosymbiotic prokaryotes gained acceptace.
• Later Sequence comparisons unequivocally
  identified proteobacteria and cyanobacteria as
  ancestors of mitochondria and chloroplasts.

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• Incorporation through endosymbiosis
• Host that acquired plastids probably had two
  flagella
• Host that acquired mitochondrion is debated.
  Generally accepted to have an affinity to
  archaebacteria, but beyond that cannot agree on
  its intracellular organization (prokaryotic,
  eukaryotic, or intermediate), its age,
  biochemical lifestyle, or how many and what kind
  of genes it possessed.

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• Many proteins that are encoded by the nuclear
  genome are essential to chloroplasts and
  mitochondria.
• Suggested that genes had been relocated from the
  ancestral organelles to the nucleus during
  evolution. This hypothesis has proved to be
  robust.

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• Gene content
• Organelles contain a miniscule set of genes
  compared with the nuclear genome.
• Sequenced plastids encode from 20-200 proteins.
• Mitochondrial genomes encode from 3-67 proteins.
• Unusual mitochondrial genomes contain many linear
  chromosomes with one gene each or the
  mitochondrial genome is missing altogether.

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• Magnitude of organelle genome reduction
• Close cousins of mitochondrial and chloroplast
  genomes are free-living alpha-proteobacteria and
  cyanobacteria.
• Modern alpha-proteobacterium Mesorhizobium loti
  7Mb of DNA encodes for more than 6,700 proteins.
• Bradyrhizobium japonicum contains a 9.1 Mb genome
  with more than 8,300 proteins.
• Cyanobactierum Nostoc PCC 7,120 has a 6.4 Mb
  genome, which encodes 5,400 proteins.
• Nostoc punctiforme is gt9 Mb, coding for 7,200
  proteins.

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• Nuclear copies of organelle genes now did the
  functions of the genes in those organelles

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• Two main categories of comparative analyses
• Studies that identify copies of genes that are
  still present in an organelle genome but that are
  also in other compartments of the same cell
  indicate the process of DNA movement and identify
  evolutionarily recent transfer events.
• Studies that analyze nuclear genomes, organelle
  genomes and the genomes of candidate prokaryotic
  ancestors to identify genes in the nucleus that
  are no longer present in the organelle show more
  ancient transfer events.

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• DNA can move among cell compartments.
• Fragments of cpDNA were found in the maize
  mitochondrial genome.
• Reports of mtDNA sequences and chloroplast
  sequences in nuclear DNA followed.
• Promiscuous DNA
• DNA mobility among the genetic compartments of
  eukaryotic cells.

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Black arrows Gene transferWhite arrows
Organelles import more than 90 of their
proteins from the cytoplasm.Dotted arrow DNA
still being transferred to the nucleus.Chloropla
st and nuclear sequences found in mitochondrial
genome, but little/no promiscuous DNA found in
the chloroplast
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Evidence

• Yeast genome contains tracts with 80-100
  similarity to mtDNA that range in size from 22 to
  230 base pairs integrated at 34 sites. Indicates
  recurrent transfer events, from ancient to
  contemporary.
• Human genome has at least 296 NUMTs of between
  206 bp and 14,654 bp (90 of the mitochondrial
  genome) that cover the entire mtDNA circle.
• Older NUMTs are more abundant in the human genome
  than recent integrants, indicating mtDNA can be
  amplified once inserted and many are organized as
  tandem repeats.

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Plant evidence

• Plant evidence of endosymbiotic DNA transfer.
• Arabidopsis contains a large (620 kb) insert of
  mtDNA and 7kb that is distributed among 13 small
  integrants. Also contains 17 insertions totaling
  11kb of cpDNA.
• Rice in just chromosome 10 contains 28 cpDNA
  fragments gt80bp long, including two very long
  insertions. Rice also shows many plastid
  insertions.

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• Recurrent transfers and convergent gene losses
• The deletion and functional replacement of
  mitochondrial genes by nuclear copies has
  effectively stopped in higher animals
  mitochondria encode 12 to 13 proteins.

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• Recurrent transfers and convergent gene losses
• Process is still active in higher plants, which
  have larger numbers of mitochondrially encoded
  proteins.
• mitochondrial rps10 gene has been independently
  transferred to the nucleus many times.
• The chloroplast translation initiation factor 1
  gene also shows striking evidence of mobility.
  Nuclear relocation accompanied by mutational
  decay and/or deletion of the corresponding
  chloroplast sequence.

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Similarity in gene content among contemporary
plastid genomes is the result of immensely
convergent evolution (parallel losses in
independent lineage).

• Same core set of genes (for photosynthesis and
  translation) has been retained.

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What do acquired nuclear genes do?

• Always targeted to the cell compartment form
  which the genes that encoded them originated?
  NO!
• In Arabidopsis, fewer than half the proteins that
  are identifiable as acquisitions from
  cyanobacteria are predicted to be targeted to
  chloroplasts. Many are targeted to the cytosol,
  the secretory pathway or the mitochondrion. A
  similar proportion of proteins that are targeted
  to the plastid do not seem to be acquired from
  cyanobacteria.
• Eukaryotes, 630 nuclear-encoded proteins were
  identified that originated form mitochondria, of
  which lt30 were predicted to be targeted to
  mitochondria in yeast and humans.

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• Many genes have relocated from the ancestral
  organellar genomes to the nucleus. Many have
  become competent nuclear copies that
• drive biogenesis of mitochondria and chloroplasts
  
• others have evolved to control further essential
  cellular processes

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• When genes are donated from organelles to the
  nucleus, there is no homing device that
  automatically re-routes the protein product back
  to the donor organelle.
• Rather, chance, natural selection and lineage
  diversification seem to govern the intracellular
  targeting fate of genes that organelles donate to
  the chromosomes of their host.
• In this sense, gene donations from organelles are
  important starting material for the evolution of
  new genes that are specific to the eukaryotic
  lineage.

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Questions of Empiricism Real time transfer and
research directions

• Chloroplast ? Nucleus gene transfer gametic vs.
  somatic cells
• Implications of Uniparental inheritance in this
  process
• Resulting questions for future research
• Example Coral-algal symbiosis

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Construct
Exclusive expression in chloroplast Confers
resistance to spectinomycin
Exculsive expression in nucleus, confers
resistance to kanamyacin
Homologous Recomb.
Transplantome Identified via growth of plant
cells in spectinomycin medium.
Huang, C. Y., Ayliffe, M. A. Timmis, J. N.
Direct measurement of the transfer rate of
chloroplast DNA into the nucleus. Nature 422,
7276 (2003).The initial experimental measurement
of the frequency of integrative transfer of DNA
between chloroplast and nucleus.
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X
Transplastomic plants
Female WT
WT Chloroplasts Nucleus with Transposition (?)
Screen for kanamyacin resistance
Huang, C. Y., Ayliffe, M. A. Timmis, J. N.
Direct measurement of the transfer rate of
chloroplast DNA into the nucleus. Nature 422,
7276 (2003).The initial experimental measurement
of the frequency of integrative transfer of DNA
between chloroplast and nucleus.
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Results

• 1 in 16,000 male tobacco gametes contained a
  newly integrated segment of chloroplast DNA.
• 1 transfer / 16,000 individuals / generation
• Is estimate high or low?
• May be low,
• The screen only identified whole, intact
  transfers, not partial transfers.
• The screen only identified the transfer of one
  intact gene, not the rate of all cholorplast gene
  transfer. Other regions integrated but were not
  detected

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• Locations of Insertions
• Highly variable, found each time (N 16 nuclear
  inserts/250,000) in a different location.
• What does this say about the timing of plastid
  DNA transfer?
• During meitoic or post-meiotic events in the
  formation of male gametes.
• Formation of GametesAt this specific rate.

• Somatic insertions
• A similar experiment was done measuring
  chloroplast to nucleus insertions in the somatic
  cells of tobacco.
• Estimate of transfer rate One event in every 5
  million somatic cells

Stegemann, S., Hartmann, S., Ruf, S. Bock, R.
Proc. Natl Acad. Sci. USA 100, 88288833 (2003).
High frequency gene transfer from the chloroplast
genome to the nucleus.
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Why is there a difference in gametic vs. somatic
insertion rates???
Programmed degeneration of plastids in pollen,
prior to fertilization..
This event is the essential step that all
eukaryotic cells that show in uniparental
inheritance of cytoplasmic genes. The
degradation of cytoplasmic organelle genomes may
increase the availability of cytoplasmic genes
that are escaping to the nucleus? These findings
generate many questions about interstitial life
and symbiosis..
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Is nuclear inheritance of cytoplasmic genes an
inherent design within uniparental
inherentance. If so, who benefits? The host or
the symbiont? Some direct lines of
question Evolutionary relationships Symbiotic
interactions Interstitial genomes The effect
of genes on multiple levels of life ONE QUICK
EXAMPLE
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Coral-algal symbiosis Thermal limits of symbiosis
determined by activity of Rubisco II
Bacterial form of RUBISCO II
Reduction in Symbiodinium genome
3 event of endosymbiosis
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Questions and Comments