Genes involved in plant stress

1. Functional Evolution of Genes Involved in
Stress Responses of Plants
Cheng Zou
Shiu Lab
Thomashow lab
2/19/2008

2. • Pattern of the loss
,maintanence and gain of
functions
• what is the process

3. Paralogs R1,
R2
T1 T2
A1
A2
Orthologs Group R1,R2 T1,T2,A1,A2

4. • Cross species -----Functional evolution in
orthologs groups
• Arabidopsis ------Functional evolution in
Paralogs

5. The stress response mechanism is Unique
in Plants
Soybean aphid, Aphis glycines (Photo Claudio Gratton)

6. Evolution of stress responsiveness cross
species
Questions:
1. To what degree is the cold stress
responsiveness of genes conserved
2. What kinds of genes tend to be
conserved

7. Cold responsiveness of Arabidopsis, potato
and tomato
• ATH 1 Gene Chips ----- Arabidopsis
(Colleen)
• TIGR Potato 10k cDNA Array ---------
potato and tomato (Marcela Carvallo )
Treatment >> control (% 5 FDR ) and > 2 fold change Up
(1)
Treatment << control ( %5 FDR ) and >2 fold change Down
(-1)
Not significant non
responsive (0)

8. Cold responsiveness in orthologs groups
(OGs)
Reconcile species
tree
with the gene tree
Rice tomato Arabidopsi
s
duplication
speciation

9. Identify OGs in potato and Arabidopsis
Poplar
Arabidopsis
Rice
potato
Anchor potato
onto
reconciled
gene tree
Arabidopsi
s Poplar Rice
Arabidopsis
potato
0:1 1:0 1:1 1:n 1:n n:n
moss

10. Cold responsiveness conservation in OGs of
Arabidopsis and potato
Arabidopsis
potato
1
1
0
0
0
0
0
0
0
0
1
1
0
0
0
conserved
# AT genes =680
#AT genes
=307
# potato genes =521
# potato genes =325
P(A.t_up|S.t_up) = 31% P(A.t_up|S.t_up) = 33.6%
P<10-5
P<10-5

11. Cold responsiveness conservation in
OGs of tomato and potato
tomato
potato
1
1
0
0
0
0
0
0
0
0
1
1
0
0
0
conserved
# tomato genes =277
#tomato genes
=744
# potato genes =249
# potato genes =749
P(S.l_up|S.t_up) = 73% P(S.t_up|S.l_up) = 69%

12. Summary of evolution stress responsiveness
cross species
• The cold
responsiveness
conservation is
dependent on the
divergence time.
• The two species that
diverged 112 ~156
Mya still have 30%
conservation.
• The two species that
diverged 12 Mya have
about 70%
conservation
Poplar Tomato
Arabidopsis Potato Rice
112 ~156 Mya
12 Mya

13. Evolution of stress responsiveness in one
species ---Arabidopsis
Questions
1. After gene duplication in one
species, what is the probability of
the daughter genes losing or
maintaining their function, and what
is the probability of gaining a new
function?
2. How do those probabilities
change over time?

14. Type of stress
Abiotic
stress
Osmotic
UV-B
Wounding
Cold4C
Heat
Genotoxic
Drought
Salt
Oxidative
Type of stress
Biotic stress
avrRpm1
DC3000
Flg22
GST-NPP1
HrcC-
HrpZ
LPS
P-infestans
Psph
Treatment >> control (% 5 FDR ) Up (1)
Treatment << control ( %5 FDR ) Down (-1)
Not significant non responsive (0)
Gene responsiveness under 16 stress
conditions from ATgeneExpress

15. Reconstruction of ancestral gene functions
Step1:construct
the phylogeny of
genes
Step2: map
current functions
Step3:reconstruct
the function of
ancestral genes

16. Evolution of stress responsiveness from
ancestral to current state------branch based
Ancestral
Current
switch
gain
lost
Maintenanc
e

17. 0>-0
change
General proportion of function change
Ancestral
Current
switch
gain
lost
Maintenanc
e
1->1
-1-> -1
-1->0
1->0
gain
switch

18. Evolution of stress responsiveness over Ks

19. loss of stress responsiveness over Ks
N Maintenance
NLoss
Ks Ks
Nswitch
Abiotic stress biotic stress
N = total number

20. NGain
gain of stress responsiveness over Ks
Ks Ks
Abiotic stress biotic stress
N Maintenance
Nswitch
N = total number

21. Summary the branch based observations
• Maintenance > loss > gain > switch
• loss of stress responsiveness rate:
Ks< 0.8 decreasing over time
Ks> 0.8 almost consistent
• gain of stress responsiveness rate:
Ks<0.8 increasing over time
Ks>0.8 almost consistent

22. stress responsiveness evolution and
regulatory network
• Maintenance > loss > gain > switch
1
-1
-1
0
Switch ~ loss
Switch<<Loss

23. stress responsiveness evolution in
duplicated pairs

24. stress responsiveness evolution in
duplicated pairs
0
10
20
30
40
50
60
70
80
90
100
lost redendunt sub neo
S
Both lost Both maintain sub neo

25. Sub-functionalization of stress responsiveness
• Partition of ancestral function
0
0.2
0.4
0.6
0.8
1
1 0~1 0
Sp
Frequency
(pair)
2-5
6-10
11-15
>15
Number of conditions
1 1 1 1 1
1 1 0 -1 0
0 1 1 1 0
Informative conditions
1 1 1 1
1 1 0 0
0 0 1 1
Sp = 1
1 1 1 1
1 1 1 1
0 0 0 0
Sp = 0

26. Number of up-regulated conditions per
gene
Abiotic stress biotic stress

27. Summary the pairwised based observations
• Sub-functionalization > both maintain > neo-
functionalization
• Sub-functionlization is extremely asymmetric

28. Acknowledgement
• Shinhan Shiu
• Kousuke Hanada
• Melissa Lehti-Shiu
• Gaurav Moghe
• Michael F. Thomashow
• Marcela Carvallo
• Colleen Doherty

29. Duplication mechanisms
+
• Whole genome duplication
• Tandem duplication
• Segmental duplication
• Replicative transposition

30. Families that enriched in up regulated genes
which arise from whole genome duplication

31. Families that enriched in up regulated genes
which arise from whole genome duplication

32. Families that enriched in up regulated genes
which arise from tandem duplication

33. Pipeline for identify OGs based on gene tree
topology
Blastx against proteins
in other 4 plant species
tblastn
EST contig
Best Match
Protein EST contig
EST peptide
APR orthlog
include Protein
A R
P P
A R
Align the sequences
and build a NJ-tree
S.t 70344
S.c 36781(77%)
S.t 49358(70%)
S.c 33953
S.t 45669
S.c 31171(66%)
S.t 42763(61%)

34. Cold responsiveness conservation in the fine OGs of S.t
and S.c
1871 221
209 476
349 40
66 174
S.l_up_0
S.t_up_0
S.t_up>0
S.l_up>0
S.t.
S.l
S.t.
S.l.
P(S.l_up|S.t_up) = 69%
P(S.l_up|S.t_up) = 72.5%

35. Function conservation in OGs of
tomato and potato
S .l
S.t
observation
P< 10 -3
P< 10 -5
221
209

36. Function evolution in paralogs ---Arabidopsis
Questions
1. After gene duplication in one species, what is the probability of
the daughter gene lose or maintain their function, and what is the
probalility of gain a new function?
2. How does those probabilities change along evolution?
3. How does the result inply the machenism of regulatory network
in one species?
Challenge
2.gene function of ancestral genes

37. Interpret the branch based
observations 2

38. The significance of the asymmetric partition
gene1
gene2
observed frequency
Log likelihood ratio = -----------------------
expected frequency

39. Breadth of the stress responsiveness in different
domain families

40. Genes involve in stress responds in RLKs tend to
located in Tandem clusters
Shiu et al., 2004 Plant Cell

41. Paralogs R1,
R2
T1 T2
A1
A2
Orthologs Group R1,R2 T1,T2,A1,A2

42. 201
1854
402
775
123
1
195
314 114
242 38
94 28
377 170
133 94
70 43
56 32
A_up_0 A_up_>0
S.t_up_0
S.t_up>0




 








N
j
N
i
j
i
N
i
i
N
j
j
obs
j
i
OG
j
i
OG
j
i
OG
j
i
OG
j
i
OG
j
i
OG
,
1
,
1
1 1
10
exp
10
)
,
(
)
,
(
)
,
(
)
,
(
log
)
,
(
)
,
(
log
P(A.t_up|S.t_up) = 22.9%
P(A.t_up|S.t_up) = 33.6%
P(A.t_up|S.t_up) = 41%
P(A.t_up|S.t_up) = 36%
Gene in cold are divergent between A.t and S.t
in fine OGs
2
P(S.t_up|A.t_up) = 36.8%
P(S.t_up|A.t_up) = 42.2%
P(S.t_up|A.t_up) = 35%
P(S.t_up|A.t_up) = 42%

43. Cold response of S.t paralogs are more conserved
than A.t paralogs
P(S.t_2_up| A.t_up& S.t_1_up ) = 57%
P(A.t_2_up| S.t_up& A.t_1_up ) =26%
?
?

44. Function
lost Function gain
Function
maintain
Function evolution in Paralogs Arabidopsis
Function evolution in orthologs Cross species

45. Cold responsiveness conservation in the fine OGs of S.t
and S.c
1871 221
209 476
349 40
66 174
S.l_up_0
S.t_up_0
S.t_up>0
S.l_up>0
S.t.
S.l
S.t.
S.l.
P(S.l_up|S.t_up) = 69%
P(S.l_up|S.t_up) = 72.5%

46. Summary of function evolution of OGs
• In OGs that the two species have evolved a long time
(more than 100 million years), the cold response of
orthologs diverged greatly. The number of conserved
OGs is slightly bigger than random expectation
• In OGs of two close related species, the cold response
of orthlogs are conserved. Generally, if one gene in OGs
in up regulted, the probability of one of its orthologs is up
regulated is around 70%.

47. Evolution of stress responsiveness
• How does or stress responsiveness change over
time?
• Ancestral to current state: maintenance, gain, loss,
and switch

48. Functional change over Ks
• Maintenance, gain, vs. loss
NM
NG
NS
NS
NG
NM
1
1
0
-1
-1
1
0
-1

49. Functional change over Ks
• Maintenance, loss, and swtich
NM
NL
NS
NS
NL
NM
N
NM
N
NL
N
NS
N
NM
N
NL
N
NS

50. Sub-functionalization of stress responsiveness
• Partition of ancestral function
0
0.2
0.4
0.6
0.8
1
1 0~1 0
Sp
Frequency
(pair)
2-5
6-10
11-15
>15
Number of conditions
1 1 1 1 1
1 1 0 -1 0
0 1 1 1 0
Informative conditions
1 1 1 1
1 1 0 0
0 0 1 1
Sp = 1
1 1 1 1
1 1 1 1
0 0 0 0
Sp = 0

51. The significance of the asymmetric partition
gene1
gene2
observed frequency
Log likelihood ratio = -----------------------
expected frequency

52. Summary
• TF family is over represented in up regulated genes which evolved from
whole genome duplication
• Kinase family and families involved in secondary metabolism are over
represented in up regulated genes which evolved from tandem duplication
• After duplication, daughter genes lost their and gain function gradually ,but it
become consistent
• When function partition happened in a pair of daughter genes, the partition
is extremely asymmetric.

53. Gene regulation under stress conditions
Type of stress
Abiotic stress
Osmotic
UV-B
Wounding
Cold4C
Heat
Genotoxic
Drought
Salt
Oxidative
Type of stress
Biotic stress
avrRpm1
DC3000
Flg22
GST-NPP1
HrcC-
HrpZ
LPS
P-infestans
Psph
Treatment >> control (% 5 FDR ) Up (1)
Treatment << control ( %5 FDR ) Down (-1)
Not significant Not regulated
(0)

55. Stefanie Hartmann
postdoc, Todd Vision’s lab

56. Paralogs R1,
R2
T1 T2
A1
A2
Orthologs Group R1,R2 T1,T2,A1,A2

57. • Cross species -----Function evolution in
orthologs
• Arabidopsis ------Function evolution in Paralogs