DNA sequence evolution in Sunflower and Lettuce

1
DNA sequence evolution in Sunflower and Lettuce
Thesis capstone report

• Yi Zou

Advisor Dr. Loren Rieseberg Dr. Sum Kim
Major Bioinformatics 07/16/2004
2
Background

• Sunflower and lettuce represent two major
  subfamilies of the Compositae Family, which is
  one of the largest and most diverse families of
  flowering plants
• Sunflower is an important oil seed crop and
  domestication and breeding have focused on seed
  traits.
• Lettuce is an important leaf crop and
  domestication and breeding have focused on leaf
  traits.
• Extensive lettuce and sunflower EST database
  available (CGPDB)

3
Background

• Examination of DNA differences between closely
  related species of Compositae will provide
  insight into the nature of mutational rates and
  processes in this family
• Hypothesis genes associated with primary
  domestication traits (seeds in sunflower and
  leaves in lettuce) will evolve faster than genes
  expressed in other tissues.
• Hypothesis upstream enzymes in metabolic
  pathways will evolve less rapidly than downstream
  enzymes.

4
Goals

• Compare distribution of indels and base
  substitutions among closely related lettuce and
  sunflower EST sequences
• Compare rates of EST sequence divergence for
  genes from different tissue types
• Compare rates of EST sequence divergence from
  different pathway, and protein evolution among
  specific genes along major metabolic pathways

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Data
http//cgpdb.ucdavis.edu
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CGPDB

• contains about 112,000 individual ESTs sequenced
  from both sunflower and lettuce
• Sunflower about 44,000 individual ESTs,
  previously assembled into 4430 unique contigs,
  were sequenced from two Helianthus annuus
  cultivars RHA801(exotic) and RHA280(oil).
• Lettuce around 68,000 ESTs, previously assembled
  into 8179 unique contigs(genes), sequenced from
  two species Lactuca serriola (wild) and L.
  sativa (cultivated)

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Goals

• Compare distribution of indels and base
  substitutions among closely related lettuce and
  sunflower EST sequences
• Compare rates of EST sequence divergence for
  genes from different tissue types and metabolic
  pathways
• Compare rates of EST sequence divergence from
  different pathway, and protein evolution among
  specific genes along major metabolic pathways

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Data Analysis Example from sunflower
Genotype 1
Genotype 2
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Data Analysis comparison of complete EST
sequence
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Data Analysis comparison of coding region
only
sun_vs_ath_TIGR_unique lettuce_vs_ath_TIGR_unique

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Result - Sequences information for assembling
Contigs and conseneus
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Result Comparison of complete EST sequences
between two sunflower and two lettuce genotypes
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Result Comparison of coding region only
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• Conclusion1
• Substitutions are 3-6 times more frequent than
  indels in both sunflower and lettuce, regardless
  of whether coding regions or complete EST
  sequences are analyzed.

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Goals

• Compare distribution of indels and base
  substitutions among closely related lettuce and
  sunflower EST sequences
• Compare rates of EST sequence divergence for
  genes from different tissue types and metabolic
  pathways
• Compare rates of EST sequence divergence from
  different pathway, and protein evolution among
  specific genes along major metabolic pathways

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Data Analysis EST divergence for different
tissue types

• Lettuce
• TAG0 - callus - "cls"
• TAG1 - roots - "rot"
• TAG2 - none (leaf) - "not"
• TAG3 - flowers pre-fert - "flr"
• TAG4 - flowers post-fert - "flo"
• TAG5 - chemical induction - "chi"
• TAG6 - none - "nos"
• TAG7 - roots env stress - "rts"
• TAG8 - shoots env stress - "shs"
• TAG9 - germinating seeds - "gsd"
• TAG10 - flowers env stress - "fls"
• TAG11 - leaves dark grow - "lvd
• Tag_1_7 all root related contigs
• Tag_3_4_10 All flower related contigs
• Tag_7_8_10 All contigs related to environment
  stress

• Sunflower
• TAG0 - callus - "cls"
• TAG1 - roots - "rot"
• TAG2 - disk ray flowers - "drf"
• TAG3 - flowers pre-fert - "flr"
• TAG4 - developing kernel - "dkn"
• TAG5 - chemical induction - "chi"
• TAG6 - none - "nos"
• TAG7 - roots env stress - "rts"
• TAG8 - shoots env stress - "shs"
• TAG9 - germinating seeds - "gsd"
• TAG10 - flowers env stress - "fls"
• TAG11 - hulls - "hls
• Tag_1_7 all root related contigs
• Tag_3_10 All contigs related to flower
• Tag_7_8_10 All contigs related to environment
  stress

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Result number of tissue-specific contigs in
sunflower
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Result number of tissue-specific contigs in
lettuce
Lettuce TAG-specific contig information
800
738
700
600
500
contigs with coding region
found in both genotypes
400
336
278
300
140
200
85
78
69
60
100
30
29
18
14
11
0
0
0
TAG1(rot)
TAG3(flr)
TAG0(cls)
TAG2(no)
TAG4(flo)
TAG5(chi)
TAG7(rts)
TAG6(nos)
TAG8(shs)
TAG9(gsd)
TAG10(fls)
TAG11(lvd)
TAG_1_7(root)
TAG_7_8_10(stress)
TAG_3_4_10(flower)
Tissue and Treatment
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Result Rates of sequence divergence among
tissue-specific contigs in sunflower and
lettuce
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Result Comparison of rates of sequence
divergence for genes expressed in seeds
versus other
16.00
T-test SubRateKH vs SubRateNonKH P-value
0.0009414
14.00
indel rate
12.00
10.00
Content
8.00
Substitution
6.00
rate
4.00
2.00
0.00
DknHls
Non-DknHls
Other
Seeds
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Result Rates of sequence divergence among
treatment-specific contigs in sunflower and
lettuce
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• Conclusion2
• As predicted, sunflower genes expressed in seeds
  evolve significantly faster than genes expressed
  in other tissues. Artificial selection for large
  seeds and high seed oil content may contribute to
  these higher rates.
• For lettuce, there are no significant differences
  in rates of sequence evolution among different
  tissues
• No differences were found in sunflower or lettuce
  among biotic and abiotic stress treatments

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Goals

• Compare distribution of indels and base
  substitutions among closely related lettuce and
  sunflower EST sequences
• Compare rates of EST sequence divergence for
  genes from different tissue types and metabolic
  pathways
• Compare rates of EST sequence divergence from
  different pathway, and protein evolution among
  specific genes along major metabolic pathways

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Data Analysis EST divergence among metabolic
pathways

• To identify contigs for specific pathways, the
  metabolic pathway information from TAIR (The
  Arabidopsis Information Resource
  http//www.arabidopsis.org/) database was
  utilized.
• Each contig in the CGPDB was assigned to an
  Arabidopsis gene locus (or remained unassigned)
  based on the BLAST results.
• Genes (contigs) for different metabolic pathways
  were clustered and protein divergence was
  estimated.

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Data Analysis protein evolution along major
metabolic pathways

• Metabolic pathways
• Lipid metabolic pathways
• Phenylpropanoid biosynthetic pathways
• Cellulose, lignin, sucrose etc. metabolic
  pathways
• The nonsynonymous substitution rate (Ka) was
  calculated for enzymes in different positions
  along pathways
• Software DnaSP 4.0 was utilized for this
  calculation

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Result comparison of metabolic pathway genes
between CGPDB and TAIR

• Based on the blast results, the contigs in CGPDB
  were compared with genes in TAIR and assigned to
  appropriate pathways
• Currently there are 186 pathways with more than
  800 unique reactions in the TAIR database. For
  these reactions, 1144 unique locus_iDs were
  assigned to the enzymes involved.
• Among the TAIR loci, 72.1 match Contigs in the
  sunflower database and 83.15 match Contigs in
  the lettuce database.

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Result Rates of sequence evolution for
sunflower metabolic pathway-specific contigs
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Result Rates of sequence evolution for lettuce
metabolic pathway-specific contigs
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Result Nonsynonymous substitution rate (Ka)
for genes along four metabolic pathways in
sunflower and lettuce
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Conclusion3

• Rates of sequence divergence did not differ among
  metabolic pathways
• Rates of protein evolution (Ka) did not vary
  along metabolic pathways (i.e., upstream genes
  evolved at the same rate as downstream genes)

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Summary

• Substitutions are much more frequent than indels
  in both sunflower and lettuce
• Sunflower genes expressed in seeds evolve
  significantly faster than genes expressed in
  other tissues
• There are no significant differences in rates of
  sequence evolution among different tissues in
  lettuce
• Rates of sequence divergence did not vary
  significantly either among or along metabolic
  pathways in either sunflower or lettuce

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Acknowledge

• Thanks
• Dr. Loren Rieseberg
• Dr. Sun Kim
• Dr. Sheri Church
• Dr. Zhao Lai