Title:

Testing for natural selection in human exonic splicing regulators

associated with evolutionary rate shifts

Publicationtype:

ResearchArticle

Authors:

RodrigoF.Ramalho,SaharGelfman,JorgeS.deSouza,GilAst,SandroJ.deSouza,DiogoMeyer

SupplementaryMaterial

Exondatabase

Ourexondatabasewascreatedwithpublicgenomic(build36.1)andcDNA(mRNAsandESTs)sequencedatafromUCSCGenomeBrowser(

TheidentificationofsplicingeventswasobtainedbythealignmentofcDNAsonthehumangenomeaspreviouslydescribed(Galanteetal.2004;Galanteetal.2007).Insummary,thecoordinatesofexon/intronbordersforallcDNAsmappedontothehumangenomewerecomparedagainsteachothertoidentifyallsplicingvariantsforallhumangenes.WeusedthesoftwareSIM4(

SupplementaryTables

TableS1:Numberofidentical(orcompletelyaligned)motifsbetweenrate-shiftedhexamersandknownESRssets.

Motifset / SRP55
(133) / SRP40
(671) / SF2_IgM
(924) / SC35
(2585) / RESCUE
(238) / PESE
(2069) / ESS
(176)
Rate-shiftedSkipped
(145) / 1 / 25 / 32 / 85 / 12 / 103 / 9
Rate-shiftedConstitutive
(198) / 4 / 30 / 50 / 113 / 21 / 161 / 4

TableS2:ModifiedMKtestappliedforExonicSplicingSilencers(ESS)locatedwithin(a)Constitutiveexonsand(b)Skippedexons.TherespectivevaluesfortheNeutralityindex(NI)anditsconfidenceintervals(C.I)arepresentedbeloweachcontingencytable.

TableS2a:Constitutiveexons

DISRUPT-ESS / MAINTAIN-ESS
Substitutions / 2433 (+0,9%) / 925 (-2,3%)
Polymorphism / 711 (-3,0%) / 311 (+7,7%)
NI = 0,87
Chi-square p-value = 0.08
Genes / Pool / p-value-Pool / NI-Mantel-Haenszel (C.I) / NI-Tarone-Greenland (C.I)
1970 / 0,84 / 0,03 / 0.85 (0,70-1,05) / 0,85 (0,68-1,06)

TableS2b:Skippedexons

DISRUPT-ESS / MAINTAIN-ESS
Substitutions / 484 (+2,0%) / 185 (-4,9%)
Polymorphism / 165 (-5,4%) / 82 (+13.2%)
NI = 0,78
Chi-square p-value = 0.12
Genes / Pool / p-value-Pool / NI-Mantel-Haenszel (C.I) / NI-Tarone-Greenland (C.I)
495 / 0,77 / 0,14 / 0,49 (0,25-0,92) / 0,36 (0,18-0,79)

TableS3ModifiedMKtestappliedforrate-shiftedESRsidentifiedfromsequencesofhumanconstitutiveexons(rate-shiftedconstitutiveESRs)locatedwithin(a)Constitutiveexonsand(b)Skippedexons.TherespectivevaluesfortheNeutralityindex(NI)anditsconfidenceintervals(C.I)arepresentedbeloweachcontingencytable.

TableS3a:Constitutiveexons

CREATE-rate-shiftedconstitutive / MAINTAIN- rate-shiftedconstitutive
Substitutions / 5606 (-0,2%) / 2630 (+0,5%)
Polymorphism / 1915 (+0,7%) / 872 (-1,5%)
NI = 1,03
Chi-square p-value = 0.54
Genes / Pool / p-value-Pool / NI-Mantel-Haenszel (C.I) / NI-Tarone-Greenland (C.I)
2924 / 1,05 / 0,29 / 1,08 (0,95-1,20) / 1,15 (1,01-1,30)

TableS3b:Skippedexons

CREATE- rate-shiftedconstitutive / MAINTAIN- rate-shiftedconstitutive
Substitutions / 1101 (-0,4%) / 479 (+0,9%)
Polymorphism / 432 (+1,1%) / 178 (-2,5%)
NI = 1,05
Chi-square p-value = 0.64
Genes / Pool / p-value-Pool / NI-Mantel-Haenszel (C.I) / NI-Tarone-Greenland (C.I)
764 / 1,10 / 0,39 / 0,86 (0,64-1,14) / 0,89 (0,66-1,21)

TableS4:ModifiedMKtestappliedforrate-shiftedskippedESRslocatedinConstitutiveexonswith(a)strong3’ splicesite,(b)strong5’ splicesite,(c)weak3’ splicesiteand(d)weak5’ splicesite.TherespectivevaluesfortheNeutralityindex(NI)anditsconfidenceintervals(C.I)arepresentedbeloweachcontingencytable.

TableS4a:Strong3'ss

CREATE- rate-shiftedskipped / MAINTAIN- rate-shiftedskipped
Substitutions / 2287 (-0,7%) / 650 (+2,7%)
Polymorphism / 764 (+2,3%) / 187 (-8,4%)
NI = 1,16
Chi-square p-value =0.12
Genes / Pool / p-value-Pool / NI-Mantel-Haenszel (C.I) / NI-Tarone-Greenland (C.I)
671 / 1,11 / 0,25 / 2,89 (2,03-4,31) / 2,39 (1,60-3,73)

TableS4b:Strong5'ss

CREATE- rate-shiftedskipped / MAINTAIN- rate-shiftedskipped
Substitutions / 2689 (-0,4%) / 687 (+1,4%)
Polymorphism / 849 (+1,2%) / 200 (-4,6%)
NI = 1,08
Chi-square p-value =0.40
Genes / Pool / p-value-Pool / NI-Mantel-Haenszel (C.I) / NI-Tarone-Greenland (C.I)
1840 / 1,00 / 0,98 / 0,94 (0,73-1,22) / 1,03 (0,78-1,35)

TableS4c:weak3'ss

CREATE- rate-shiftedskipped / MAINTAIN- rate-shiftedskipped
Substitutions / 2328 (-0,9%) / 626 (+3,6%)
Polymorphism / 831 (+2,7%) / 186 (-10,3%)
NI = 1,20
Chi-square p-value = 0.05
Genes / Pool / p-value-Pool / NI-Mantel-Haenszel (C.I) / NI-Tarone-Greenland (C.I)
1559 / 1,14 / 0,2 / 1,14 (0,84-1,56) / 1,21 (0,88-1,70)

TableS4d:weak5'ss

CREATE- rate-shiftedskipped / MAINTAIN- rate-shiftedskipped
Substitutions / 1926 (-1,6%) / 589 (+5,5%)
Polymorphism / 746 (+4,3%) / 173 (-14,9%)
NI = 1,31
Chi-square p-value = 0.005
Genes / Pool / p-value-Pool / NI-Mantel-Haenszel (C.I) / NI-Tarone-Greenland (C.I)
1485 / 1,3 / 0,01 / 1,06 (0,75-1,46) / 1,05 (0,69-1,54)

Supplementary Figures

Figure S1