The Molecular History of Eukaryotic Life

The Molecular History of Eukaryotic Life The Synapomorphies

David Nelson Dec. 10, 2000; modified Jan. 11, 2001; Last modified Jan. 19, 2001

Note: some data removed on April 18, 2001 to satisfy requirements on no prior publication
for Science. Once publication is in press, this data will be restored.

Given below is a list of current insertions used for making phylogenetic inferences. The list
is followed by some genereal and specific discussion about these insertions. The ARP3, enolase
and xanthine dehydrogenase/ aldehyde oxidase insertions are original with this website.

EF1 alpha insertion for fungal (including Microsporidia) + animal clade
actin related protein 3 (ARP3) insertion for animal + fungal + Amoebozoa clade
alanyl tRNA replacement from mitochondrial genome for Diplomonandida exclusive clade
This may include Parabasala also, but no sequence is available yet.
Ribosomal protein L24 insertion for Euglenozoa specific clade
xanthine dehydrogenase/ aldehyde oxidase insertion for plant specific clade
This may include Rhodophyta and Glaucophyta but no sequences are available yet.
Ribosomal protein L34 insertion for plant specific clade
This may include Rhodophyta and Glaucophyta but no sequences are available yet.

Rare insertions or deletions do occur in proteins resulting in length variations. If
the change in length is small and it is not in a critical region so the protein can tolerate the
change, these mutations become part of the history of the protein. They are passive markers
that are inherited in all future descendants of this gene. The thirteen eukaryotic branches
that represent all extant eukaryotic life each had a beginning. There was a time that each line
branched off from a common ancestor and existed for a while as a single species before splitting
again. The interval between the first branching and the second branching had a duration.
Certainly, there were mutations that occurred during this time and probably neutral insertions
or deletions occurred that have been preserved over time to the present day. These are the 12
synapomorphies that we are searching for to define the geneaology of the main eukaryote lines.

Twelve is a small number. Genomes on the other hand are large, with thousands of genes.
At the bare minimum there will be a few hundred of these genes that have orthologs in every
species of eukaryote. It is only a matter of looking at enough sequence data to find the
insertions/ deletions that occurred at the right time interval at the beginning of a branch. With
genome projects begun on a number of human pathogens like Plasmodium, Leishmania, Trypanosoma,
Giardia, Encephalitozoon cuniculi and other key non-pathogenic species on the eukaryotic tree like
Dictyostelium, it will not be long before massive amounts of data are available to search.

Already two of these 12 branchings are resolved. The first was the plant, animal, fungi
trichotomy. The molecular feature that supported the joining of animals and Fungi was an
insertion in the EF1 alpha protein sequence. This 12 amino acid insertion was present in
animals, fungi and no other eukaryote or prokaryote phyla with sequence data(1). The same was
also true for three short deletions in enolase. These are synapomorphies that define a clade.
The structure of the cells is not important, and in this case it was misleading.

Since the microsporidia have been moved into the fungi, I checked for the 12 amino acid
insertion from a microsporidian to see if it was there and I did find it as an 11 amino acid
insertion. See below. This is the data from Figure 1A of (1) with the sequence of Glugea
plecoglossi added and some relabeling of higher order taxa to fit the new results of Baldauf
et al. (2). EST sequences from Alveolates, Stramenopiles and Oxymonads (Parabasala) have been
added to make the set more complete. Note the KTL sequence after the gap in Eimeria. This is
most like the animal and fungi sequences. Two of the Rhodophyta sequences have an insert of
four amino acids seen in six ESTs from yezoensis and one complete mRNA from purpurea.

DEFINITION Glugea plecoglossi peptide elongation factor 1 alpha, complete cds.
ACCESSION D84253
MAEEKPILNVCFIGHVDSGKSTTVGNLAFQLGAIDARKMDKLKK
EAEERGRGTFSYAYVMDMSAAERERGITITTSLMKLETSKHMLNVIDCPGHQDFIKNM
VTGAAQADVGVVLVPCATGEFESCISGGTLKDHIMISGVLGCRKLIVCVNKVDTIDEK
NRISRFDEVAKEMKGIIAKSHPDKDPIIIPISGYLGINIVEKGDKFEWFKGWKPVSGA
GDSIFTLEGALNSQIPPPRPIDKPLRMPIDSIHKIPGIGMVYTGRVSTGAIKPGMVVS
SQPTGVVAEVKTLEIHKQSRAAVVSGENCGVALKAASQGNPALIKPGHVFSNTKDSPV
EIFEAARAKIVVVAHPKGIKPGYCPTMDLGTHHVPCQITKFISKRMPGIKEEIPSPDV
VQKGENVTCIIHPQKQVVMETLKEVPSLARFALRDAGRIVGIGAIEARYTKAEYETEV
PSTTGKGRKATRGPG

Animals Xenopus laevis GDNMLEPSPNM....PWFKGWKITRKEGSGSGTTLLEALDCILPPSRP
Animals Drosphila melanogaster GDNMLEASDRL....PWYKGWNIERKEGKADGKTLLDALDAILPPSRP
Animals Onchocerca volvulus GDNMLEPSANM....PWFKGWSVERKEGTMTGKTLLEALDSVVPPQRP
Fungi Saccharomyces cerevisiae GDNMIEATTNA....PWYKGWEKETKAGVVKGKTLLEAIDAIEQPSRP
Fungi Mucor racemosus GDNMLDESTNM....PWFKGWNKETKAGSKTGKTLLEAIDAIEPPVRP
Microsporidia Glugea plecoglossi GINIVEKGDKF....EWFKGWKPVSGAGD-SIFTLEGALNSQIPPPRP
Amoebozoa Dictyostelium discoideum GDNMLERSDKM....EWYKG............PTLLEALDAIVEPKRP
Amoebozoa Entamoeba histolytica GDNMIEPSTNM....PWYKG............PTLIGALDSVTPPERP
Plants Arabidopsis thaliana GDNMIERSTNL....DWYKG............PTLLEALDQINEPKRP
Plants Triticum aestivum (wheat) GDNMIERSTNL....DWYKG............PTLLEALDQINEPKRP
Rhodophyta Porphyra yezoensis GENLFERTDKTHALGKWYKG............PCLLEALDNCDPPKRP
Rhodophyta Porphyra purpurea tef-c GENLFERTGGDHALGKWYKG............PCLLEALDACDPPKRP
Rhodophyta Porphyra purpurea tef-s GDNMLEKSTNM....PWYTG............PTLFEVLDAMKPPKRP
Glaucophyta Cyanophora paradoxa GDNMLEPSSNL....GWYKG............PTLVEALDQVEEPKRP
Heterokonta Phytophthora infestans GDNMIDRSTNM....PWYKG............PFLLEALDNLNAPKRP
Heterokonta Laminaria digitata GDNMVDKSTNM....PWYKG............PYLMEALDTMKEPTRP
Alveolata Eimeria S5-2 GDNMVERSSNM....GWYKG............KTLVEALDSVEPPKRP
Alveolata Plasmodium falciparum GDNLIEKSDKT....PWYKG............RTLIEALDTMEPPKRP
Discicristata Euglena gracilis GDNMIEASENM....GWYKG............LTLIGALDNLEPPKRP
Discicristata Trypanosoma cruzi GDNMIDKSENM....PWYKG............PTLLEALDMLEPPVRP
Discicristata Acrasis rosea GDNMLEKSTNM....PWYKG............PTLLEALDALEPPKRP
Diplomonadida Giardia lamblia GDNIMEKSDKM....PWYEG............PCLIDAIDGLKAPKRP
Parabasala Dinenympha exilis GDNMLDRSTNM....PWYKD............PILFDALDLLEVPKRP
Parabasala Pyrsonympha grandis GDNMLERSPNM....SWYKD............PILFEALDLLEVPKRP
Parabasala Trichomonas vaginalis GDNMTEKSPNM....PWYNG............PYLLEALDSLQPPKRP
Parabasala Trichonympha agilis GDNMTEKSDKM....PWWKG............LTLLEALDTLEPPKRP
Archaea Sulfolobus acidocaldarius GDNVTHKSTKM....PWYNG............PTLEELLDQLEIPPKP
Archaea Halobacterium halobium GDNIAEESEHT....GWYDG............EILLEALNELPAPEPP

The second branch to be resolved is Giardia’s position as one of the most distant
members of the eukaryotes based on alanyl tRNA synthetases. There is a paper in the Oct. 24
PNAS on alanyl tRNA synthetases(3). Eukaryote ATSs appear to have come from the mitochondrial
ancestor by displacing the host ATS of archaeal origin. These two types of ATS are easy to
distinguish because of a fairly large insertion in the eukaryotes that is missing in the
archaea. When this gene was sequenced from Giardia (a diplomonad and thought to be a very
ancient eukaryote), the insert was not there. It looked like the archaeal ATS genes. Giardia
do not have mitochondria, but they do have mitochondrial like nuclear genes like HSP70s etc.
This suggests that they did have the mitochondrial ancestor and did transfer some genes from it
to the nucleus. However, the ATS gene did not replace its host gene counterpart in this line
(possibly the only line of eukaryotes where this failed to happen). The authors argue that the
diplomonads diverged very early before this key evolutionary event happened. This qualifies for
a very rare unique evolutionary landmark, placing Giardia outside all other eukaryote lineages.
The sequence of this gene needs to be done for other members of the Archezoa group like
Trichomonas vaginalis to see if they all have the archaeal sequence. If they do then the last
branch on the eukaryote tree is fixed and every other eukaryote species falls between the animal
fungal clade and the Archezoa clade.

The next branch to be fixed in place is probably the Mycetozoa. With most of the Dictyostelium
genome sequenced, there is a good chance a key synapomorphy could be found that unites this
clade with the animal fungal clade and excludes plants and protists.

[Jan. 16, 2001] An insertion has been found in ARP3 (actin related protein 3) that unites
animals, fungi and Amoebozoa (Mycetozoa + Lobosea). This supports the tree in Baldauf et
al.(2). ARP3 is part of a conserved eukaryotic complex called the ARP2/3 complex that organizes
microtubules. The alignment of the region of interest is shown below. Because this is an actin
related gene and there are hundreds of actin sequences and many isoforms, it is difficult to do
the comparison. Actin numbering in Arabidopsis is at least up to actin 12. The insertion that
seems relevant is the one shown in sequences 1-7 and 12 below and probably the one in the two
entamoeba histolytica sequences (14,15 Amoebozoa). The latter two have two additional amino
acids, that probably reflects a further modification of this insert. The plants have a five
amino acid insert that is conserved across monocots and dicots. There is an intron in the
Arabidopsis gene that occurs between SK and CEM. The three animals shown do not have an intron
in the insertion. The fungi do not have an intron here. Arabidopsis actin genes have a phase 1
intron in G of GIVLD (Actin 1, 2, 3, 4, 7, 8, 11, 12) The intron in the ARP3 Arab gene is phase
0 and much larger. Note that potatos have at least eight kinds of actin and none of those have
this insert. I speculate that the plant ARP3 insert is derived from a separate event related to
the modification of the intron seen in other plant actins. Note that the Arabidopsis ARP3 actin
does not have the intron seen in the other Arabidopsis actins mentioned above. I point out the
insertion in enolase that unites plants with heterokonts and alveolates to the exclusion of
animals and fungi as additional support for this interpretation. The fact that there are
Amoebozoa (and fungi and animals, not shown) actin sequences without the insert suggests it is
specific to ARP3. It is not clear if ARP3 exists in all the species shown, though the ARP2/3
complex seems to be well conserved among eukaryotes. The sequence fragment shown ends in GYVIGS
instead of GY[AS]LPH for the other actins and that sequence is not found yet in the heterokonts,
alveolates, discicristata, diplomonadida and parabasala.

1. PGLYIAVQAVLALAASWTSrqvgert..lTGTVIDSGDGVTHVIPVAEGYVIGS ANIMALS
2. PGLYIAVQAVLALAASWTSrqvgert..lTGTVIDSGDGVTHVIPVAEGYVIGS ANIMALS
3. PGLYIAVQAVLALAASWASrsaeert..lTGIVVDSGDGVTDVIPVAEGYVIGS ANIMALS
4. AGLYIAVQAVLALAASWTSskvqdrs..lTGTVIDSGDGVTHVIPVAEGYVIGS FUNGI
5. AGLYIAVQAVLALAASWTSskvtdrs..lTGTVVDSGDGVTHIIPVAEGYVIGS FUNGI
6. AGLYIAVQAVLALAASWTSskvtdrs..lTGTVIDSGDGVTHVIPVAEGYVIGS FUNGI
7. PGLYIAVQAVLALAASWTSkna.ekt..lTGTVIDSGDGVTHVIPISEGYVIGS AMOEBOZOA
8. PAMYVAIQAVLSLYASGRT..........TGIVMDSGDGVSHTVPIYEGYALPH AMOEBOZOA
9. PAMYVAIQAVLSLYASGRT..........TGIVMDSGDGVSHTVPIYEGYALPH AMOEBOZOA
10. PAMYVAIQAALSLYASGRT..........TGIVMDSGDGVSHTVPIYEGYALPH AMOEBOZOA
11. PAMYVAIQAVLSLYASGRT..........TGIVMDSGDGVSHTVPIYEGYSLPH AMOEBOZOA
12 PGLYIAVQAVLALAASWTSkqvtekt..lTGTVIDSGDGVTHVIPVAEGYVIGS AMOEBOZOA
13. PAMYVAIQAVLSLYASGRT..........TGIVLDSGDGVTHTVPIYEGYALPH AMOEBOZOA
14. PGMYIAVQAVLAIVASWSRkdnnpanarlTGTVIDSGDGVTHIIPVADGYVIGS entamoeba hist.
15. PGMYIAVQAVLAMCNHGSRkdnnpanaelTGTVIDSGDGVTHIIPVADGYVIGS entamoeba hist.
16. PGLYIAVNSVLALAAGYTTsk.....cemTGVVVDVGDGATHVVPVAEGYVIGS Arabidopsis intron in gap
17. PGLYIAVNSVLALXAGYTTsk.....cxmTGVVXDIGDGAT Medicago truncatula EST
18. VLALAAGYTTtk.....cemTGVVVDVGDGATHIVPAADGYVIGS Zea mays EST
19. PGLYIAVXSVLALAAGYTTsk.....cemTGIVVDGGDGATHVVPGADRYVIGS Glycine max EST
20. PGLYIAVQPVLALAAGYTAsk.....cemTGVVVDIGDGATHVVPVAEGYVIGS Lycopersicon (tomato) EST
21. PAMYVAIQAVLSLYASGRT..........TGIVLDSGDGVSHTVPIYEGYALPH PLANTS
22. PAMYVAIQAVLSLYASGRT..........TGIVLDSGDGVSHTVPIYEGYALPH PLANTS
23. PAMYVAIQAVLSLYASGRT..........TGIVLDSGDGVSHTVPIYEGYALPH PLANTS
24. PAMYVAIQAVLSLYASGRT..........TGIVLDSGDGVSHTVPIYEGYALPH PLANTS
25. PAMYVAIQAVLSLYASGRT..........TGIVLDSGDGVSHTVPIYEGYALPH PLANTS
26. PAMYVAIQAVLSLYASGRT..........TGIVLDSGDGVSHTVPIYEGYALPH PLANTS
27. PAMYVAIQAVLSLYASGRT..........TGIVLDSGDGVSHTVPIYEGYALPH PLANTS
28. PAMYVAIQAVLSLYASGRT..........TGIVLDSGDGVSHTVPIYEGYALPH PLANTS
29. PAMYVAIQAVLSLYASGRT..........TGIVLDSGDGVSHTVPIYEGYALPH PLANTS
30. PAMYVAIQAVLSLYASGRT..........TGIVLDSGDGVSHTVPIYEGYALPH PLANTS
31. PAMYVAIQAVLSLYASGRT..........TGIVLDSGDGVSHTVPIYEGYALPH PLANTS
32. PAMYVAIQAVLSLYASGRT..........TGIVMDSGDGVSHTVPIYEGYALPH PLANTS
33. PAMYVAIQAVLSLYASGRT..........TGIVLDSGDGVSHTVPIYEGYALPH PLANTS
34. PAFYVAIQAVLSLYASGRT..........SGIVVDSGDGVTHTVPIYEGYSLPH RHODOPHYTA
35. PAMYVAIQAVLSLYASGRT..........TGIVLDSGDGVTHTVPIYEGXALPH GLAUCOPHYTA
36. PAMYVNIQAVLSLYASGRT..........TGCVLDSGDGVSHTVPIYEGYALPH HETEROKONTA
37. PAMYVNIQAVLSLYASGRT..........TGCVLDSGDGVSHTVPIYEGYALPH HETEROKONTA
38. PAMYVNIQAVLSLYASGRT..........TGCVLDSGDGVSHTVPIYEGYALPH HETEROKONTA
39. PAMYVNIQAVLSLYASGRT..........TGCVLDSGDGVSHTVPIYEGYALPH HETEROKONTA
40. PAMYVNIQAVLSLYASGRT..........TGCVLDSGDGVSHTVPIYEGYALPH HETEROKONTA
41. LAMYVNIQAVLSLYASGST..........TGCVLDSGDGVSHTVPIYEGYALPH HETEROKONTA
42. PAMYVAIQAVLSLYSSGRT..........TGIVLDSGDGVSHTVPIYEGYALPH ALVEOLATA
43. PAMYVSIQAILSLYASGRT..........TGIVLDSGDGVSHTVPIYEGYVLPH ALVEOLATA
44. PAMYVAIQAVLSLYSSGRT..........TGIVLDSGDGVSHTVPIYEGYALPH ALVEOLATA
45. PAMYVNIQAVLSLYASGRT..........TGIVLDSGDGVSHTVPIYEGYAIPH ALVEOLATA
46. PALYVSIQAVLSLYSSGRT..........TGIVLDCGDGVSHTVPIYEGYSLPH DISCICRISTATA
47. PAMYVNIQAVLSLYASGRT..........TGCVLDSGDGVSHTVPIYEGYALPH DISCICRISTATA
48. PAMYVAIQAVLSLYASGRT..........TGIVLDSGDGVSHTVPIYEGYALPH DISCICRISTATA
49. PAMYVAIQAVLSLYASGRT..........TGIVLDSGDGVSHTVPIYEGYALPH DISCICRISTATA
50. PAFYVQVQAVLALYSSGRT..........TGIVIDTGDGVTHTVPVYEGYSLPH DIPLOMONADIDA
51. PSFYVGIQAVLSLYSSGRT..........TGIVFDAGDGVSHTVPIYEGYSLPH PARABASALA
52. PSFYVGIQAVLSLYSSGRT..........TGIVFDAGDGVSHTVPIYEGYSLPH PARABASALA
53. PSFYVGIQAVLSLYSSGRT..........TGIVFDAGDGVSHTVPIYEGYSLPH PARABASALA
54. PSFYVGIQAVLSLYSSGRT..........TGIVFDAGDGVSHTVPIYEGYSLPH PARABASALA
55. PSFYVGIQAVLSLYSSGRT..........TGIVFDAGDGVSHTVPIFEGYSLPH PARABASALA

1. = Fugu rubipes pufferfish AF034581.1 actin-related protein (ARP3) gene
2. = ARP3_HUMA human AC027120.4 Homo sapiens chromosome 3 clone RP11-141B14
3. = ARP3_DROME Drosophila melaonogaster AE003556.2 genomic scaffold
4. = ARP3_NEUCR NEUROSPORA CRASSA
5. = ARP3_SCHPO SCIZOSACCHAROMYCES POMBE
6. = ARP3_YEAST SACCHAROMYCES CEREVISIAE
7. = ARP3_DICDI Dictyostelium dicoideum
8. = ACT1_DICDI Dictyostelium dicoideum
9. = ACT8_DICDI Dictyostelium dicoideum
10. = ACT2_DICDI Dictyostelium dicoideum
11. = ACT3_DICDI Dictyostelium dicoideum
12. = ARP3_ACACA Acanthamoeba castellanii (Amoeba)
13. = ACT1_ACACA Acanthamoeba castellanii (Amoeba)
14. = AZ680014.1 ENTIP12TF Entamoeba histolytica
15. = AZ527984.1 ENTCL76TF Entamoeba histolytica
16. = AC007357 gene= F3F19.20 Strong similarity to gb|U29610 Actin-like protein (Arp3)
17. = BF638396.1 NF057B03PL1F1027 Phosphate starved leaf Medicago truncatula
18. = BE025380.1|BE025380 945026A07.Y1 945 - Mixed adult tissues Zea mays
19. = AW202092.1 sf11a07.y1 Gm-c1027 Glycine max
20. = BE441091.1 EST408361 tomato immature green fruit Lycopersicon esculentum
21. = ACTB_ARATH ARABIDOPSIS
22. = ACT1_ARATH ARABIDOPSIS
23. = ACT3_ARATH ARABIDOPSIS
24. = ACT4_ARATH ARABIDOPSIS
25. = ACTC_ARATH ARABIDOPSIS
26. = ACTD_SOLTU SOLANUM TUBEROSUM (POTATO)
27. = ACTC_SOLTU SOLANUM TUBEROSUM (POTATO)
28. = ACT8_SOLTU SOLANUM TUBEROSUM (POTATO)
29. = ACTA_SOLTU SOLANUM TUBEROSUM (POTATO)
30. = ACT5_SOLTU SOLANUM TUBEROSUM (POTATO)
31. = ACT3_SOLTU SOLANUM TUBEROSUM (POTATO)
32. = ACT9_SOLTU SOLANUM TUBEROSUM (POTATO)
33. = ACTB_SOLTU SOLANUM TUBEROSUM (POTATO)
34. = AV435141.1 Porphyra yezoensis
35. = U90325.1 Cyanophora paradoxa actin gene
36. = ACT1_PHYIN Phytophthora infestans
37. = ACT2_PHYIN Phytophthora infestans
38. = ACT_PHYME Phytophthora megasperma (Potato pink rot fungus)
39. = ACT_COSCS Costaria costata
40. = ACT_FUCVE Fucus vesiculosus
41. = ACT_FUCDI Fucus distichus
42. = ACT1_PLAFA Plasmodium falciparum
43. = ACT2_PLAFA Plasmodium falciparum
44. = ACT_TOXGO Toxoplasma gondii
45. = ACT_CRYPV Cryptosporidium parvum
46. = EUGLENA GRACILIS O65204
47. = ACT_ACHBI Achlya bisexualis
48. = ACT1_NAEFO Naegleria fowleri
49. = ACT2_NAEFO Naegleria fowleri
50. = ACT_GIALA GIARDIA LAMBLIA
51. = U63122.1 Trichomonas vaginalis actin gene
52. = U63124.1 Trichomonas vaginalis Type2 actin mRNA
53. = U63126.1 Trichomonas vaginalis Type3 actin mRNA
54. = U63125.1 Trichomonas vaginalis Type4 actin mRNA
55. = U63123.1 Trichomonas vaginalis Type5 actin mRNA

XANTHINE DEHYDROGENASE / ALDEHYDE OXIDASE PLANT SPECIFIC INSERTION
The xanthine dehydrogenase and aldehyde oxidase genes code for molybdenum containing proteins
with FeS centers and flavins. They share a common ancestor(4). Only the plants so far show the
8 amino acid insert below. Arabidopsis has at least three aldehyde oxidase genes and one
xanthine dehydrogenase(5). The sampling in the lower eukaryotes is not present yet. Depending
on the time of this insertion, the question of Plantae as a monophyletic group may be answered,
if the insertion is found in Rhodophyta and Glaucophyta but no other lower eukaryote groups. At
the present, the sequences are not available from these species.

ANIMALS 1 GQGLHTKMVQVASRALK........IPTSKIYISETSTNTVPNTSPTAA HUMAN XDH
ANIMALS 2 GQGVHTKMIQVVSRELR........MPMSNVHLRGTSTETVPNANISGG HUMAN AOX
ANIMALS 3 GQGLNQKMLQVCSEALK........RPIDTITIVDCSTDKITNAPETGA C. ELEGANS AF038614
ANIMALS 4 GQGLHTKILQIAARCLE........IPIERIHIHDTSTDKVPNASATAA C. ELEGANS Z83318
ANIMALS 5 GQGLNTKMIQCAARALG........IPSELIHISETATDKVPNTSPTAA D. MELANOGASTER
ANIMALS GQGMNTKISQVAAHTLG........IPMEQVRIEASDTINGANSMVTGG D. MELANOGASTER
ANIMALS 6 GQGVNTKVAQVAAHILG........IPMTKISIKTMSSLTSPNASVSGG CULEX MOSQUITO
FUNGI 7 GQGLHTKMTMIAAEALG........VPLSDVFISETATNTVANTSSTAA ASPERGILLUS
FUNGI 8 GQGLHTKMTQIAAQALN........VPLENVFISETATNTVANASATAA NEUROSPORA
PLANTS GQGLHTKVAQVAASAFN........IPLSSVFVSETSTDKVPNASPTAA Arab XDH AL079347
PLANTS 9 GQGLWTKVQQMVAYGLGmikcegsdDLLERIRLLQTDTLSMSQSSYTAG ARABIDOPSIS
PLANTS 10 GQGLWTKVKQMTAFGLGqlcpgggeSLLDKVRVIQADTLSMIQGGVTGG ZEA MAYS
PLANTS 11 AQGLWTKVRQMTAYALGsiesswaeDLVEKVRVIQADTLSVVQGGLTAG TOMATO
PLANTS 12 GQGLWTKVKQMTAYALSliqcagneELLEKVRVIQADTLSLIQGGFTSG COTTON
PLANTS 13 SQGLMTKVKQMAAFALGavqcdridSLLDKVRVVQTDTVSLIQGGLTAG GLYCINE MAX
PLANTS 14 GQGLWTKVQQMTAFGLGelcpdggeSLLDKVRVIQADTLSMIQGGFTGG SORGHUM BICOLOR
PLANTS 15 GQGLWTKVKQMAAFGLGqlwadrsqDLLERVRVIQADTFIVVQGGWTTG WHEAT
PLANTS 16 GQGLWTKVRQMTAYALGsiesswaeDLVEKVRVIQADTLSVVQGGPTNG POTATO
BACTERIA 17 GQGLHAKMVQVAAAVLG........IDPVQVRITATDTSKVPNTSATAA RHODOBACTER CAPSULATUS

1 = NM_000379.2 Homo sapiens xanthine dehydrogenase (XDH)
2 = L11005.1 Human aldehyde oxidase (hAOX) mRNA
3 = U53333.1 Caenorhabditis elegans cosmid F36A4 THIS SEQ HAS AN INTRON IN THE GAP
4 = Z83318.1 C. elegans cosmid F55B11 Aldehyde oxidase/ xanthine dehydrogenase
5 = AE003698.2 Drosophila melanogaster
6 = AF202953 Culex pipiens quinquefasciatus aldehyde oxidase (AO1)
7 = X82827 hxA gene; xanthine dehydrogenase. Aspergillus nidulellus.
8 = AL391572 Neurospora crassa probable xanthine dehydrogenase
9 = AB037271 Arabidopsis thaliana AAO4 mRNA for aldehyde oxidase
10 = D88452 Zea mays mRNA for aldehyde oxidase-2
11 = U82559.1 Lycopersicon esculentum aldehyde oxidase 1 homolog (TAO1)
12 = AI728545 BNLGHi11006 Six-day Cotton fiber Gossypium hirsutum
13 = BE801522 sr15f03.y1 Gm-c1050 Glycine max + BF066257 st28c11.y1 Gm-c1067 Glycine max THESE
TWO ESTS ARE NOT FROM IDENTICAL GENES AA 1-9 FROM BF066257
14 = BE596841 PI1_59_C08.b1_A002 Pathogen induced 1 (PI1) Sorghum bicolor
15 = BE497404 WHE0751_A10_A19ZS heat-stressed seedling cDNA library Triticum aestivum
16 = BE921778 EST425547 potato leaves and petioles Solanum tuberosum
17 = AJ001013.1|RCXDHAB Rhodobacter capsulatus xdhB (XANTHINE DEHYDROGENASE B)

TREE IS A UPGMA TREE WITHOUT THE INSERT INCLUDED BASED ON 41 AMINO ACID ALIGNMENT ONLY
XDH genes cluster at the top of the tree. Animal genes below the Bacterial XDH gene are
Assumed to be Aldehyde oxidase genes, though only human is known to be AOX

+---------ANIMALS 1 HUM XDH
+--6
! +---------ANIMALS 5 DROS XDH
+-10
! ! +----FUNGI 7 ASPER XDH
! ! +------3
+-11 +--8 +----FUNGI 8 NEUROS XDH
! ! !
! ! +-----------PLANTS ARAB XDH
+------13 !
! ! +--------------ANIMALS 4 C ELEGANS XDH
+-15 !
! ! +----------------BACTERIA 17 XDH
! !
+-16 +------------------------ANIMALS 2 HUM AOX
! !
! ! +-----------------ANIMALS DROS AOX
+------------17 +------14
! ! +-----------------ANIMALS 6 CULEX AOX
! !
! +----------------------------ANIMALS 3 C ELEGANS AOX
!
-18 +--------------PLANTS 9
! !
! ! +-PLANTS 10
! ! +------1
! AOX GENES ! ! +-PLANTS 14
+--------------------------12 +--5
! ! ! +--PLANTS 11
! ! ! +---2
! +--7 +--4 +--PLANTS 16
! ! ! !
! ! ! +------PLANTS 12
+--9 !
! +----------PLANTS 15

The EF1 alpha sequences have a two amino acid deletion that appears in several groups
so it may have occurred multiple times. It may not be too useful as a cladistic
marker. It is interesting to note that Entamoeba histolytica (seq. 15) clustered with
Euplotes (seqs. 31,32 in Alveolata) in a 1997 paper by Baldauf and Doolittle(6)
using EF1 alpha sequences to look at eukaryote evolution. Perhaps the Entamoeba
sequence does belong with the Alveolates and not with the Amoebozoa, though the text
of that paper claims this is an artefact. The close association between Euplotes and
Entamoeba EF1 alpha may arise from a lateral gene transfer. If this were true then
the 2 amino acid deletion would only have happened in the fungi and some of the
alveolates. It is complicated by the observation that some but not all ciliophorans
have the deletion, while all apicomplexans seem to have it. Note: one of the Euplotes sequences
is probably an animal contamination. It contains the animal specific sequence STEPPYS.
One scenario for these results is that the ancestor to the alveolates suffered the 2 amino acid
deletion, and the line of ciliophorans including Paramecium, Tetrahymena, Colpoda, Stentor and
Stolonychia regained the 2 amino acida as an insert.

1. NKMDSTEPPYSQKRYEEIVKEVSTYIKKIGY XM_004398.1 ANIMALS
2. NKMDSTEPPYSQARFEEITKEVSAYIKKIGY L23807 ANIMALS
3. NKMDSTEPPYSEARYEEIKKEVSSYIKKIGY AE003779 ANIMALS
4. NKMDSTEPPFSEARFTEITNEVSGFIKKIGY U51994.1 ANIMALS
5. NKMD..TTGWSQARFEEIVKETSNFIKKVGF D82571.1 FUNGI
6. NKMD..TTQWSQTRFEEIIKETKNFIKKVGY D45837.1 FUNGI
7. NKMD..SVKWDESRFQEIVKETSNFIKKVGY M15666.1 FUNGI
8. NKMD..SVKWDESRFQEIVKETSNFIKKVGY M15667.1 FUNGI
9. NKMD..STKYSEARYNEIVKEVSTFIKKIGF AF157303 FUNGI
10. NKMD..STKYSEARYNEIVKEVSTFIKKIGY AF157274 FUNGI
11. NKMD..STKYSEARYNEIVKEVSTFIKKIGY AF157283 FUNGI
12. NKMD..STKYSEARYNEIVKEVSTFIKKIGY AF157250 FUNGI
13. NKMDEKSVNWSQARYDEIVKETSSFVKKIGY AF016243 AMOEBOZOA
14. NKMDEKSTNYSQARYDEIVKEVSSFIKKIGY X55972.1 AMOEBOZOA
15. NKMD..AIQYKQERYEEIKKEISAFLKKTGY M92073.1 AMOEBOZOA ?
16. NKMDATTPKYSKARYDEIIKEVSSYLKKVGY AC026875.4 PLANTS
17. NKMDATTPKYSKARYDEIVKEVSSYLKKVGY X14449.1 PLANTS
18. NKMDATTPKYSKARYDEIVKEVSSYLKKVGY U76259.1 PLANTS
19. NKMDATTPKYSKARYDEIVKEVSSYLKKVGY D63582.1 PLANTS
20. NKMDDKNVNWSKERYEEVSKEMDLYLKKVGY U08844.1 RHODOPHYTA
21. NKMDEKSVNYGQPRFEEIKKEVSAYLKKIGY AF092951.1 GLACOCYSTOPHYCEAE
22. NKMDDKSVNYSEARYKEIKAEMTSFLTKVGY AF091355.2 HETEROKONTA
23. NKMDDSSVMYGQARYEEIKSEVTTYLKKVGY AJ249839.1 HETEROKONTA
24. NKMDDSSVMYGESRYTEIKEEVAIYLKKVGY AW400940 HETEROKONTA
25. NKMD..TVKYSEDRYEEIKKEVKDYLKKVGY AJ224150.1 ALVEOLATA Apicomplexa
26. NKMD..TVKYSEDRYEEIKKEVKDYLKKVGY AJ224151.1 ALVEOLATA Apicomplexa
27. NKMD..TVKYSEDRYEEIKKEVKDYLKKVGY AJ224153.1 ALVEOLATA Apicomplexa
28. NKMD..TVKYSEDRYEEIKKEVKDYLKKVGY X60488 ALVEOLATA Apicomplexa
29. NKMD..TCEYKQSRFDEIFNEVDGYLKKVGY U71180.1 ALVEOLATA Apicomplexa
30. NKMD..SCNYSEDRFNEIQKEVAMYLKKVGY N81319 ALVEOLATA Apicomplexa
31. NKMD..AAEYDEERYNEIKKEVSEYLAKVGY AF056100.1 ALVEOLATA Ciliophora
32. NKMD..AAEYDETRYKEIKKEVSEYLDKVGY AF056099.1 ALVEOLATA Ciliophora
33. NKMD..TIDYDEERFNEIVENVSDHLGKIGY U26267.1 ALVEOLATA Ciliophora
34. NKMD..AVQYNEERFTDIKKEVIDYLKKMGS AF056104.1 ALVEOLATA Ciliophora
35. NKMD..AISYNEERFTDIKKEVIDYLKKLGF AF056105.1 ALVEOLATA Ciliophora
36. NKMDEKTVNYAQGRYDEIVKEMRDYLKKVGY AF172083.2 ALVEOLATA Ciliophora
37. NKMDEKTVNFSEERYQEIKKELSDYLKKVGY D11083.1 ALVEOLATA Ciliophora
38. NKMDDKSVNWDQGRFLEIKKELSDYLKKIGY AF056108 ALVEOLATA Ciliophora
39. NKMDEKTVAWSQSRFEEIQKEVQEYLKKVGY AF056098 ALVEOLATA Ciliophora
40. NKMDDKSCNWSQERYEEIKKEVSQYLKKVGY AF056106 ALVEOLATA Ciliophora
41. NKMDSTEPPYSEDRYEEIKKEVSTFLAKVGY U26260.1 ALVEOLATA Probable ANIMAL contaminant
42. NKMDDKTVNYGQERYDEIVKEVSAYIKKVGY AJ234094.1 EUGLENOZOA
43. NKMDDKSVNFAQERYDEIVKEVSAYLKKVGY AI069750 EUGLENOZOA
44. NKMDDKTVQYSQARYEEISKEVGTYLKRVGY U72244.1 EUGLENOZOA
45. NKFDDKTVKYSQARYEEIKKEVSGYLKKVGY X16890.1 EUGLENOZOA
46. NKFDDTSVSYKEDRYNEIKSEVGRYLKGLGF AF058283.1 HETEROLOBOSEA
47. NKMDDKSVQYKEDRYKEIQKEVADYLKKVGY AF190771.1 HETEROLOBOSEA
48. NKMDDPNVNYSKDRYNEIKTEMTKTLVAIGY U94406.1 DIPLOMONADIDA
49. NKMDDPQVNYSEARYTEIKTEMQKTFKQIGF U37081.1 DIPLOMONADIDA
50. NKMDDPQVNYSEARYKEIKEEMQKNLKQIGY U29442.1 DIPLOMONADIDA
51. NKMDDGQVKYSKERYDEIKGEMMKQLKNIGW D14342.1 DIPLOMONADIDA
52. NKMDDNTVNYAESRYKEITEEMKNVLKQVGW AF230353.1 PARABASALA
53. NKMDDKTVNYNKARFDEIQSEMTRILTGIGY D78479.1 PARABASALA
54. NKMDDKTVNYNKARFDEITAEMTRILTGIGY AF058282.1 PARABASALA
55. NKMDDKSVNWAESRYNEIKTEMRTYLKKIGY AF230349.1 PARABASALA Oxymonadida
56. NKMDDKSVNWAESRYNEVKTEMGTYLKKIGY AB007029.1 PARABASALA Oxymonadida

1. Homo sapiens EF 1 alpha mRNA
2. Danio rerio EF 1 alpha mRNA
3. Drosophila melanogaster
4. Caenorhabditis elegans
5. S. pombe mRNA for EF 1 alpha-A
6. Neurospora crassa EF 1-alpha gene
7. S.cerevisiae EF1 alpha-A gene
8. S.cerevisiae EF1 alpha-B gene
9. Zychaea mexicana gene
10. Phascolomyces articulosus gene
11. Rhizomucor pusillus gene
12. Fennellomyces linderi gene
13. Physarum polycephalum (tef1) gene
14. D. Discoideum gene
15. Entamoeba histolytica mRNA
16. Arabidopsis thaliana Genomic There are 4 EF1-alpha genes in Arabidopsis A1-A4
17. Tomato LeEF-1 mRNAcds
18. Zea mays mRNA
19. Oryza sativa mRNA
20. Porphyra purpurea
21. Cyanophora paradoxa Glacocystophyceae
22. Blastocystis hominis gene
23. Phytophthora infestans mRNA
24. Laminaria digitata cDNA
25. Plasmodium berghei EF-1alpha A-gene
26. Plasmodium berghei EF-1alpha B-gene
27. Plasmodium knowlesi EF-1alpha A-gene
28. Plasmodium falciparum MEF-1 gene
29. Crytposporidium parvum MRNA
30. Toxoplasma gondii cDNA
31. Euplotes aediculatus TEF2 GENE
32. Euplotes aediculatus TEF1 GENE
33. Euplotes crassus EFA2 gene
34. Spathidium sp. (TEF1) gene CILIOPHORA
35. Spathidium sp. (TEF2) gene CILIOPHORA
36. Paramecium tetraurelia gene
37. Tetrahymena pyriformis mRNA
38. Stylonychia mytilus TEF1 gene
39. Colpoda inflata TEF1 gene
40. Stentor coeruleus TEF1 gene
41. Euplotes crassus GENE 80% identical to Drosophila, 70% to seq 29 Euplotes
contains STEPPY seq found in animals. Probable animal DNA contamination
42. Trypanosoma brucei cDNA
43. T. cruzi cDNA
44. Leishmania braziliensis gene
45. Euglena gracilis tef mRNA
46. Naegleria andersoni gene
47. Acrasis rosea (tef1) gene Heterolobosea
48. Spironucleus vortens gene Diplomonad
49. Hexamita inflata gene Diplomonad
50. Diplomonad ATCC50330 gene Diplomonad
51. Gardia lamblia mRNA Diplomonad
52. Trichonympha agilis mRNA Parabasala
53. Trichomonas tenax gene Parabasala
54. Trichomonas vaginalis mRNA Parabasala
55. Dinenympha exilis mRNA Oxymonadida
56. Unidentified Oxymonadida A-14 mRNA

Ribosomal protein L24 has a four amino acid insert in Euglenozoa that needs more work to clarify
the extent of this insert in other groups.

MRIEKCYFCSSPIYPGHGIQFV....RNDSTVFKFCRSRCNKLF C. elegans Animals
MKVELCSFSGYKIYPGHGRRYA....RTDGKVFQFLNAKCESAF Human Animals
MKVEIDSFSGAKIYPGRGTLFV....RGDSKIFRFQSSKSASLF K. lactis Fungi
MRVHTCYFCSGPVYPGHGIMFV....RNDSKVFRFCRSKCHKNF S. pombe Fungi
MKVEVDSFSGAKIYPGRGTLFV....RGDSKIFRFQNSKSASLF yeast B Fungi
LKTELCRFSGQKIYPGRGIRFI....RSDSQVFLFLNSKCKRYF Arabidopsis Plants
LKTELCRFSGAKIYPGRGIRFI....RGDSQVFLFVNSKCKRYF Cicer arietinum (chickpea) Plants
LKTELCRFSGQKIYPGKGIRFI....RSDSQVFLFANSKCKRYF Hordeum vulgare (barley) Plants
VKTEVCQYSGFRIYPGHGSRFI....RVDGKSYVFANSKCEASF Porphyra yezoensis AV436717 Rhodophyta
RTSLCNYSEFKIYPARGMKFV....RGDSKVFHFINTKVESLF Dicty Mycetozoa
KTQVCAFSGFKIPVAKGRKYV....RLDLKSFTFINKKSLMQF Entamoeba histolytica AJ012925 Entamoebidae
IKTETCSFSESRIYPGHGSRFI....RRDGSAYVFINSKSKSLF Phytophthora sojae BE583183 Heterokonta
IKTDMCSFSEYRIYPGRGQRFV....AKDGKVHTFIHRKEASLF Toxoplasma gondii cDNA AA531998 Alveolata
IKTELCSFSEYRIYPGRGQRFV....AKDGKVHTFLFRKEASLF Sarcocystis neurona BE635379 Alveolata
IKTELCSFSEYRIYPGRGLKFV....SRDGKVHTYIHSKEARLG Eimeria tenella BE028025 Alveolata
MRTIDCEFSHFAVHPGHGRRYVPFAFLSTKPVLTFARPKCFAMY Trypanosoma brucei Euglenozoa
MRTIECEFSHFAVHPGHGRRYVPFAFLSTKPVLTFSRPKCFALY Leishmania major embl CAC02879 Euglenozoa
MEKRVCSFCGYDIEPGTGKMYV....RRDGRVFYFCSGKCEKNM Archaeoglobus fulgidus Archaea
PEWRTCSFCGYEIEPGKGKMVV....EKDGTVLYFCSSKCEKSY Methanococcus jannaschii Archaea

Ribosomal large subunit 34 Plant specific insertion (look for other taxa with insert, especially
Rhodophyta and Glaucophyta)

CGVCPGKLRGVRPVRPKVLMR--LSKTKKHVSRAYGG human Animals
CGVCPGRLRGVRAVRPKVLMR--LSKTKKHVQPGLWW rat Animals
CGVLPGRLRGVVAVRPKVLMR--LSKTKKHVQQGLWW pig Animals
CGQCKEKLSGIKPSRPSERPR--MCRRLKTVTRTFGG aedes albopictus Animals
CGDCGSALQGISTLRPRQYAT--VSKTHKTVSRAYGG yeast A gene Fungi
CGDTGVPLQGIPALRPREFAR--LSHNKKTVQRAYGG S. pombe Fungi
CGECGVNLAGIPALRPYQYKN--LPKSRRTVSRAYGG Dictyostelium AU038253 Mycetozoa
CGDCGCVLAGIRHIRPHQYGW--LGKSKRTVTRAYGG Entamoeba histolytica AZ687907 Entamoebidae
CPVTGKRIQGIPHLRPTEYKRSRLSRNRRTVNRAYGG tobacco Plants
CPVTGKRIQGIPHLRPTEYKRSRLSRNRRTVNRAYGG pea Plants
CPVTGKRIQGIPHLRPSEYKRSRLSRNRRTVNRAYGG Arabidopsis Plants
CGDCGISLPGIKHLRPKQYKN--LKKREKTVSRAYGG Laminaria digitata AW400906 Heterokonta
CGDCKQTLKGIKHLQSKEYKN--VSKTQRTVSRAYGG Phytophthora infestans BE775634 Heterokonta
CGDCKKPLAGIPACAPYEMKH--LKKRERTVARAYGG Cryptosporidium parvum AA555437 Alveolata
CGGCGRLLPGIPARRPPQFRL--LKKRERTVNRAYGG Eimeria tenella AI757126 Alveolata
CANCHRALPGIPAVAPHKMKL--LKKKDRTVHRAYGG Sarcocystis neurona BF324030 Alveolata
CGNCDRALPGIPAVAPHRLRL--LKKRERTVHRAYGG Toxoplasma gondii AA520444 Alveolata
CAICGAELHGVPRGRPVEIRK--LPKSQRRPERPYGG Methanococcus jannaschii Archaea

Return to index

References

1. Baldauf SL, Palmer JD. Animals and fungi are each other's closest relatives: congruent
evidence from multiple proteins. Proc Natl Acad Sci U S A. 1993 Dec 15;90(24):11558-62.

2. Baldauf SL, Roger AJ, Wenk-Siefert I, Doolittle WF.
A kingdom-level phylogeny of eukaryotes based on combined protein data.
Science. 2000 Nov 3;290(5493):972-7.

3. Joseph W. Chihade, James R. Brown, Paul R. Schimmel, and Llu�s Ribas de
Pouplana. Origin of mitochondria in relation to evolutionary history of eukaryotic
alanyl-tRNA synthetase PNAS 97, 12153-12157 2000

4. Terao M, Kurosaki M, Demontis S, Zanotta S, Garattini E.
Isolation and characterization of the human aldehyde oxidase gene: conservation of
intron/exon boundaries with the xanthine oxidoreductase gene indicates a common origin.
Biochem J. 1998 Jun 1;332 ( Pt 2):383-93.

5. Biochim Biophys Acta 1998 Jul 9;1398(3):397-402
Biochemical and genetic characterization of three molybdenum cofactor
hydroxylases in Arabidopsis thaliana.
Hoff T, Frandsen GI, Rocher A, Mundy J
Aldehyde oxidases and xanthine dehydrogenases/oxidases belong to the molybdenum
cofactor dependent hydroxylase class of enzymes. Zymograms show that Arabidopsis
thaliana has at least three different aldehyde oxidases and one xanthine oxidase.

6. Baldauf SL, Doolittle WF.
Origin and evolution of the slime molds.
Proc Natl Acad Sci U S A. 1997 Oct 28;94(22):12007-12.

Ribosomal protein L24 has a four amino acid insert in Euglenozoa that needs more work to clarify the extent of this insert in other groups.

Ribosomal large subunit 34 Plant specific insertion (look for other taxa with insert, especially Rhodophyta and Glaucophyta)