Genomics of Ferns and Lycophytes
Chapter 6: Structure and evolution of fern plastid genomes
Paul G. Wolf and Jessie M. Roper
Question:
What is the inheritance of the
chloroplast genome in ferns?
Marchantia cp genome
• ca. 150 kb, circular molecule
• large and small single copy regions separated by inverted repeat
• gene number and order +/- conserved across land plants
Generally in land plants: maternal (via the egg, excluded via sperm)
• maternal with some biparental in Angiosperms
• paternal in Gymnosperms
• ferns?
Phyllitis (Aspleniaceae) – biparental
Osmunda (Osmundaceae) – maternal
Polystichum (Drypoteridaceae) – maternal
Pteridium Dennstaedtiaceae) – maternal
Pellaea (Pteridaceae) – maternal
“During insemination in Ceratopteris richardii [Pteridaceae], the
sperm cytoskeleton and flagella rearrange, and the coils of the
cell extend while entering the neck canal. . . . All cellular
components, except plastids, enter the egg cytoplasm”
Lopez-Smith and Renzagalia, 2008 (Sexual Plant Reproduction)
1992
Marchantia
30kb inversion
tobacco
Marchantia cp genome
• ca. 150 kb
• large and small single copy regions separated by inverted repeat
• gene number and order +/- conserved across land plants
1992
Lycopodium
30kb inversion
Equisetum
Psilotum
Lycopodium = Marchantia order
Osmunda
ferns = tobacco order
Fern and lycophyte total chloroplast genomes
sequenced
• Huperzia
• Isoetes
• Selaginella
• Equisetum (basal fern)
• Psilotum (basal fern)
• Angiopteris (basal fern)
• Adiantum (polypod)
• Alsophila (polypod
- 2009 paper)*
Gao et al. (2009) Complete chloroplast genome sequence of a tree fern Alsophila spinulosa
Fern and lycophyte total chloroplast genomes
sequenced
• few advanced ferns sequenced
• but, Fern Tree of Life project will do
many more
Rearrangements in fern chloroplast genomes
1. loss of some tRNA and other
protein coding genes
Gao et al. 2009
Rearrangements in fern chloroplast genomes
1. loss of some tRNA and other
protein coding genes
1. 2 inversions in the Inverted
Repeat (IR) of some ferns
Gao et al. 2009
Rearrangements in fern chloroplast genomes
IR inversion 2
1. loss of some tRNA and other
protein coding genes
IR inversion 1
1. 2 inversions in the Inverted
Repeat (IR) of some ferns
[also using PCR assays for
these inversions in other
genera]
30kb inversion
?
Chapter 7: Evolution of the nuclear genome of ferns and lycophytes
Takuya Nakazato, Michael S. Barker, Loren H. Rieseberg, and Gerald J.
Gastony
Unfurling fern biology in the genomics age (BioScience, 2010)
Michael S. Barker and Paul G. Wolf
Academic family tree of
Gerald J. Gastony
Rolla and Alice Tryon 1950s and 1990s
Is there an “Alice Tryon Women in Science” bequest for Botany Department?
Academic family tree of
Gerald J. Gastony
Rieseberg
Nakazato
Barker
The neglected fern and lycophyte nuclear genomes
- or the “crying ferns”
1. 1 genetic linkage map Ceratopteris
1. 4 EST libraries – Selaginella (2),
Ceratopteris, Adiantum
2. 3 BAC libraries - Selaginella (2),
Ceratopteris
3. 1 nuclear genome sequencing
project in the works - Selaginella
The neglected fern and lycophyte nuclear genomes
- or the “crying ferns”
Why?
1. large genome size (>2X)
1. lack of funding for low
economically important plants
The neglected fern and lycophyte nuclear genomes
- or the “crying ferns”
Why?
1. large genome size (>2X)
1. lack of funding for low
economically important plants
But !
1. 2nd largest land plant group
2. sister to seed plants
3. diverse land plant lineages need
to be compared
4. homologs of important seed
plant genes occur in ferns
A short history of the study of the fern genome
Haploid chromosome number
• 57 in ferns vs. 16 in angiosperms
[ > 14 = polyploid (Grant, 1981) ]
Ophioglossum (adder’s-tongue fern) 2n = 1440 (96 ploid) in O. reticulatum
A short history of the study of the fern genome
Haploid chromosome number
• 57 in ferns vs. 16 in angiosperms
[ > 14 = polyploid (Grant, 1981) ]
Questions:
How does this fern choreograph
meiosis with an n > 600? Has it ever
been observed? Do large n's lead to
more aborted or nonviable spores?
A short history of the study of the fern genome
Haploid chromosome number
• 57 in ferns vs. 16 in angiosperms
[ > 14 = polyploid (Grant, 1981) ]
• 13.6 in heterosporous ferns is exception
• heterosporous lycophytes << homosporous lycophytes
• heterosporous seed plants << homosporous ferns & allies
Therefore, homosporous ferns acquire high chromosome
number to select for increased heterozygosity via polyploidy
Hypothesis of Klekowski & Baker (1966)
A short history of the study of the fern genome
Two lines of evidence did not support this
hypothesis
1. Isozyme analysis indicated widespread
silencings of genes – diploid numbers
of copies
1. nn
2. Most homosporous ferns are
outcrossing
Therefore, homosporous ferns acquire high chromosome
number to select for increased heterozygosity via polyploidy
Hypothesis of Klekowski & Baker (1966)
A short history of the study of the fern genome
Homosporous ferns acquired high chromosome numbers with
diploid gene expression via repeated cycles of polyploidization
and subsequent gene silencing without chromosome loss
Hypothesis of Chris Haufler (1987)
A short history of the study of the fern genome
Many lines of evidence support this as the working
hypothesis in ferns
1. Pseudogenes in nuclear genes in Polystichum
1. FISH detection of multiple dispersed
chromosomal locations of rDNA in
Ceratopteris
1. +/- Genetic linkage map analysis in
Ceratopteris
Homosporous ferns acquired high chromosome numbers with
diploid gene expression via repeated cycles of polyploidization
and subsequent gene silencing without chromosome loss
Hypothesis of Chris Haufler (1987)
The future of fern genomics?
Ceratopteris has emerged as the “model” organism
for fern genomics
Study of the origin of polyploidy (neo- and paleo-)
Correlating genomic changes to speciation and
development
Two examples using Ceratopteris
1. Nakazato et al. (2006) genetic linkage
analysis
1. Barker (2010) EST analysis
The future of fern genomics?
Ceratopteris genetic linkage analysis
•
700 genetic markers
•
85% multiple copies
•
24% single copy – low!
•
large numbers of duplicate genes on
different chromosomes
The future of fern genomics?
Ceratopteris genetic linkage analysis surprises!
Maize linkage map
• Expect clusters of linked duplicate genes on
different chromosomes in recent (neo-) polyploids
Oxford plot of polyploid
cotton’s A & D
genomes
Rong et al. 2004
• Expect clusters of linked duplicate genes on
different chromosomes in recent (neo-) polyploids
Duplicated gene copies are
hyper-dispersed across the
genome of Ceratopteris
Indicates ancient polyploid
event and many
subsequent chromosomal
changes
• Expect clusters of linked duplicate genes on
different chromosomes in recent polyploids
The future of fern genomics?
Ceratopteris EST analysis
•
expressed sequence tags
•
examines transcriptome
•
mRNA is extracted
The future of fern genomics?
Ceratopteris EST analysis
•
cDNA is made with reverse
transcriptase
•
ds cDNA is cloned into
vector – library formed
•
cDNA sequenced from 5’
and 3’ ends (= Tags)
•
400-800 bp ESTs can be
contiged
The future of fern genomics?
Ceratopteris EST analysis
•
synonymous substitution
(silent) rate – Ks – obtained
for duplicate genes
•
most duplications young
and placed in ‘zero’ class
•
peak in duplications at 0.96
– 1.84 Ks or showing
paleopolyploidy
The future of fern genomics?
Ceratopteris EST analysis
•
synonymous substitution
(silent) rate – Ks – obtained
for duplicate genes
•
most duplications young
and placed in ‘zero’ class
•
peak in duplications at 0.96
– 1.84 Ks or showing
paleopolyploidy
•
using molecular clocks and
phylogenetic trees,
paleopolyploidy linked to
early polypod diversification
Question Set 1
1. Ferns and fern allies are diverse and old;
is it really appropriate to expect that all
have their nuclear genomes evolving by
same “rules”?
Question Set 2
1. What are the justifications for selecting
Ceratopteris richardii as a model
organism for ferns? Do the
“idiosyncratic” features of its genome
affect generalization to ferns?
1. You have been given a blank check to
sequence the fern genome of your
2. Could maintaining large amounts of
choice. Which would you choose and why?
physical genetic material be
What methods would you use?
disadvantageous for fern evolution?
Could it be related to slow speciation
2. Why is the fate of most duplicate genes to
rates, compared to angiosperms? Or, on
eventually become silenced? Could
the other hand, could the silenced genes
mutations accumulate in both copies at the
hold the key to the long history of fern
same rate causing subfunctionalization,
evolution?
where mutations cause the two copies to
functionally be diminished to one over
1. Can high chromosome numbers in ferns
time?
and lycophytes simply be an outcome of
the ‘stringent bivalent pairing’ that is
3. If you are really interested in
known in the group? How might that idea
understanding the process of speciation,
be further examined or tested?
would ferns be the better choice relative to
angiosperms?