Functional Evolution of Genes Involved in Stress Responses of Plants

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: • To what degree is the cold stress responsiveness of genes conserved • 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 Arabidopsis duplication speciation

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

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

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

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 12 Mya Arabidopsis Poplar Tomato Rice Potato 112 ~156 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. Gene responsiveness under 16 stress conditions from ATgeneExpress Treatment >> control (% 5 FDR ) Up (1) Treatment << control ( %5 FDR ) Down (-1) Not significant non responsive (0)

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

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

17. General proportion of function change 0->0 Ancestral Current \ switch gain \ lost \ Maintenance \

18. Evolution of stress responsiveness over Ks

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

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

21. Summary the branch based observations • Maintenance > loss > gain > switch • loss of stress responsivenessrate: 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 0 -1 Switch ~ loss -1 Switch<<Loss

23. stress responsiveness evolution in duplicated pairs

24. stress responsiveness evolution in duplicated pairs cond4 cond3 cond1 cond2 cond5 Both lost Both maintain sub neo Both maintain Neo-functionalization Sub_functionalization Both lost

25. Sub-functionalization of stress responsiveness • Partition of ancestral function

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

34. Cold responsiveness conservation in the fine OGs of S.t and S.c S.l_up>0 S.l_up_0 S.t. S.t_up_0 P(S.l_up|S.t_up) = 69% S.t_up>0 S.l S.t. P(S.l_up|S.t_up) = 72.5% S.l.

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

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 gene2 gene1 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. Gene in cold are divergent between A.t and S.t in fine OGs A_up_0 A_up_>0 S.t_up_0 P(A.t_up|S.t_up) = 33.6% P(S.t_up|A.t_up) = 36.8% S.t_up>0 1854 P(A.t_up|S.t_up) = 22.9% P(S.t_up|A.t_up) = 42.2% 402 P(S.t_up|A.t_up) = 35% P(A.t_up|S.t_up) = 41% 775 P(A.t_up|S.t_up) = 36% P(S.t_up|A.t_up) = 42% 201 2

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 S.l_up>0 S.l_up_0 S.t. S.t_up_0 P(S.l_up|S.t_up) = 69% S.t_up>0 S.l S.t. P(S.l_up|S.t_up) = 72.5% S.l.

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 NS NG NG NS NM

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

50. Sub-functionalization of stress responsiveness • Partition of ancestral function