2023 Undergraduate Research Symposium: Katelynn Haygood The Pax6 gene is well known for its importance in developing the visual system. The gene is able to, in the early stages of the embryo, place eye location, and control eye function. Many species of arthropods and arachnids normally possess two copies of the Pax6 gene called the eyeless and the twin of the eyeless (Yang 2009). Both genes are essential for the development of the ocular head segment (Luan et al. 2014). The horseshoe crab Limulus polyphemus has more Pax6 genes than most arthropods (Friedrich, 2022), i.e., two homologs of eyeless and three homologs of twin of eyeless. Horseshoe crabs represent an ancient chelicerate lineage and are known as some of the world's living fossils that are over 445 million years old. In this project, we were studying the horseshoe crab eyeless and twin of eyeless protein sequences to identify sequence regions that distinguish the two closely related paralogs.

1. Mmus_Pax6_NP_00 --------------------------------------------------------------------------SHSGVNQLGGVFVNGRPLPDSTRQKIVELAHSGARPCDISRILQVSNGCVSKILGRYYETGSIRPRAIGGSKPRVATPEVVSKIAQYKRECPSIFAWEIRDRLLSEGVCTNDNIPSVSSINRVLRNLASEKQQM-------------GADGMYDKLRMLNGQTGSWGTR--PGWYPGTSV-------------------
Tcas_ey_ABW2313 MA----------------------------------------------------------------------HKGHSGVNQLGGVFVGGRPLPDSTRQKIVELAHSGARPCDISRILQVSNGCVSKILGRYYETGSIRPRAIGGSKPRVATAEVVSKISSYKRECPSIFAWEIRDRLLQEGVCTNDNIPSVSSINRVLRNLAAQKEQTSQNP-----PVSSGNDSVYDKLRLLNGNQGSW-RP--TPWYSPGNTS--F--P------------
Dmag_ey_KZS1363 MIKQVDDK---------------------------------------EVKNLHIKK-----KEDIGLDIGAMMDSHSGVNQLGGVFVSGRPLPDQTRQKIVELAHSGARPCDISRILQVSNGCVSKILGRYYETGSIRPRAIGGSKPRVATNDVVGKISQFKRECPSIFAWEIRDRLLQEAICNNDNIPSVSSINRVLRNLAAQKEHQQQHQHPHQHVAIPSPDSVYDKLRMLNGQSQ-WPRTNTNTWYPNGNGA--GL-G------------
Dpul_ey_EFX7578 MLME-----------------------------------------------------------DVGLDSGAMMDSHSGVNQLGGVFVSGRPLPDQTRQKIVELAHSGARPCDISRILQVSNGCVSKILGRYYETGSIRPRAIGGSKPRVATNDVVGKISQFKRECPSIFAWEIRDRLLQEAICNNDNIPSVSSINRVLRNLAAQKEHQQQHQHPHQHATIPSPDSVYDKLRMLNGQSQ-WPRTNTNTWYPNGNGA--GL-G------------
Ptep_ey_XP_0159 MQIGLRSVDATLA-VSAASSLFRDLSAPGQSSSLTIPGATVYQGCAGATDDVMPHKDSRRSVYSPIPLIAVQDPGHSGINQLGGVYVNGRPLPDSTRQKIVELAHSGARPCDISRILQVSNGCVSKILGRYYETGSIRPRAIGGSKPRVATPDVVAKIAHFKRECPSIFAWEIRDRLLSEGACTNDSVPSVSSINRVLRNLAAQKEQAQV----------QAQDAVYDKLRMLNGQG--WPRP--NPWYPGGTTF--G--G------------
Lpol_ey_XP_0137 ML--------------------------------------------------------------------LFPPGHSGINQLGGIYVNGRPLPDSTRQKIVELAHSGARPCDISRILQVSNGCVSKILGRYYETGSIRPRAIGGSKPRVATPDVVSNIAQYKRECPSIFAWEIRDRLLSDGICTNDNVPSVSSINRVLRNLAAQKEQHVH-----------AQDAVYDKLRMLNGQG--WPRP--NPWYPASATF--G--G------------
Lpol_ey_XP_0222 MGIH-------------------------------------------TIEDFDPC------RPGPLLQSSVSDPGHSGINQLGGIYVNGRPLPDSTRQKIVELAHSGARPCDISRILQVSNGCVSKILGRYYETGSIRPRAIGGSKPRVATPPVVNKIAQYKRECPSIFAWEIRDRLLSDGICNNDNVPSVSSINRVLRNLAAQKEQQVH---------AQAQDAVYDKLRILNGQG--WPRP--NPWYPGSATF--G--G------------
Lpol_ey_XP_0222 MGIH-------------------------------------------ATEDFDPC------RPGPLRQSSITDPGHSGINQLGGIYVNGRPLPDSTRQKIVELAHSGARPCDISRILQVSNGCVSKILGRYYETGSIRPRAIGGSKPRVATPSVVAKIAHYKRECPSIFAWEIRDRLLSDGVCNNDNVPSVSSINRVLRNLAAKKEQQ-------------AQDTVYDKLRLLNGQA--WPRP--NPWYPGSATF--G--G------------
Tcas_toy_KYB279 MPHKVGLFPEEDLMHHAS--------------------------------------------MGQNTIFGCSTAGHSGINQLGGVYVNGRPLPDSTRQKIVELAHSGARPCDISRILQVSNGCVSKILGRYYETGSIKPRAIGGSKPRVATQPVVNKIAEFKRECPSIFAWEIRDRLLSEGVCNNDNIPSVSSINRVLRNLASQKEQQAS----------AQNESVYDKLRMFNGQTP----G--WAWYPGTPTA--PHLGLPPAPAALTT--
Dmag_toy_KZS136 MP----------------------------------------------------------------------HKGHSGVNQLGGVYVNGRPLPDSTRQKIVELAHSGARPCDISRILQVSNGCVSKILGRYYETGSIKPRAIGGSKPRVATSCVVTKIADYKRECPSIFAWEIRDRLLSEGVCNNDTIPSVSSINRVLRNLAAQKEQQSA---------AAASESVYDKLRMFNGQAG----G--WAWYPGAPPPTHPALT------------
Dpul_toy_EFX757 MPQCNDG-------------------------------------C------------------SALADFCFHHSGHSGVNQLGGVYVNGRPLPDSTRQKIVELAHSGARPCDISRILQVSNGCVSKILGRYYETGSIKPRAIGGSKPRVATSCVVTKIADYKRECPSIFAWEIRDRLLSEGVCNNDTIPSVSSINRVLRNLAAQKEQQSA---------AAASESVYDKLRMFNGQAG----G--WAWYPGAPPPTHPALA------------
Ptep_toy_XP_042 MP----------------------------------------------------------------------HKGHSGINQLGGVYVNGRPLPDSTRQKIVELAHSGARPCDISRILQVSNGCVSKILGRYYETGSIKPRAIGGSKPRVATPHVVAKIADYKRECPSIFAWEIRDRLLSEGVCNNDNIPSVSSINRVLRNLAAQKEQQQQHHQ-H-QLTVPGAELVYDKLRMLNGGQ-----P--WPWYHTGASH-LTAAGVPHGTSPAGLSA
Lpol_toy_XP_013 MP----------------------------------------------------------------------HKGHSGVNQLGGVYVNGRPLPDSTRQKIVELAHSGARPCDISRILQVSNGCVSKILGRYYETGSIKPRTIGGSKPRVATSAVVNKIADYKRECPSVFAWEIRDRLLADGVCNSENIPSVSSINRVLRNLTSQKDQQEPSPH---HVPMTGPESVYDKLRLLNGSQ-----P--WPWYPTGPTH--HFHGIPPTSSPVALGQ
Lpol_toy_XP_022 MNIQVNQVVEFVILPPCAYKV--GIDAPGKGL---IIGVNYVTMCAILESET----------MNWKTAKVLRLPSHSGVNQLGGMYVNGRPLPDSTRQKIVELAHSGARPCDISRILQVSNGCVSKILGRYYETGSIKPRTIGGSKPRVATAAVVNRIADYKRECPSIFAWEMRDRLLADGVCNSENIPSVSSINRVLRNLSAQKDHHQSSPQ----VPMTGPESVYDKLRMLNGAQ-----P--WPWYPTGTSH--HFHGVPPPSSPVTLGP
Mmus_Pax6_NP_00 ----------------------PG------------QPTQDGCQ--QQEGG---------GE--NT-N-SIS--SNG-EDSDEAQMRLQLKRKLQRNRTSFTQEQIEALEKEFERTHYPDVFARERLAAKIDLPEARI-----QVWFSNRRAKWRREEKLRNQRRQASN-----------------------------------------------TPSHIP-ISSSF-STSVYQPIPQPTTPVSSFTSGSMLGRTDTALTNTYSA--LP-PM
Tcas_ey_ABW2313 -------LQ----PLSPPPTILAD-----------DI--------NKKVT----DLDGINSDETNS-GDNSNA-GSS-IGPDDDQARLRLKRKLQRNRTSFTNDQIDSLEKEFERTHYPDVFARERLAAKIGLPEARI-----QVWFSNRRAKWRREEKLRNQRRGAPAPAEHP--------------------------------------GPAASPPRL----QGFNNTSMYSPLPPPM-----------------SIADTYSS--MS-SM
Dmag_ey_KZS1363 -------LTTGVGAGSPPYTPLPGATSPDQTPLCHQISSNDGSYNHKKGGDDVMSDDGGVSSGENSRSAPPTP-TTT-TPSDDDQARLRLKRKLQRNRTSFTNEQIESLEKEFERTHYPDVFARERLAAKIGLPEARI-----QVWFSNRRAKWRREEKLRNQRRGPEQQQSSSQQP--DQVNPQQQQQQQQQQ----QQQQQQQQLIPVVEPAPSSPRSVS-HSANY-PSGIYSSLATTM------------------MTDTYSS--AFNGM
Dpul_ey_EFX7578 -------LTTGVGAGSPPYTPLPGATSPDQTPLCHQISSNDGSYNHKKGGDDVMSEDGGVSSGENSRSAPPTP-TTT-TPSDDDQARLRLKRKLQRNRTSFTNEQIESLEKEFERTHYPDVFARERLAAKIGLPEARI-----QVWFSNRRAKWRREEKLRNQRRGPDQQQSSSQQP--DQVNPQQQQQQQQQQPQQQQQQQQQQQQINVVEPPPSSPRSVS-HSANY-PGGIYSSLATSM------------------MTDTYSS--AFNGM
Ptep_ey_XP_0159 -------IAP---SYIA-PVTTPA------------TPLENGLNP-KKEGS---------VT--EA-S-TPSD-QSG-SGEEDSAARLRLKRKLQRNRTSFTPEQIEALEKEFERTHYPDVFARERLAAKIDLPEARI-----QVWFSNRRAKWRREEKLRNQRRAAEQAT--------------------------------------------TTPGRLP-LNTGF-PNSLYPSIGQPMA----------------SMAETYRCGTIP-TM
Lpol_ey_XP_0137 -------ITP---ACVPPPVTVPG------------LSTENGPIQ-KKEGD--------SGT--ES-S-SRSD-PGGSGDDEENAARLRLKRKLQRNRTSFTPEQIEALEKEFERTHYPDVFARERLASKIDLPEARI-----QVWFSNRRAKWRREEKLRNQRRAAEQAAAAAA------------------------------------A-VNTTPGRLP-LNSGF-PNSLYPGIGQPMA----------------SMGDSYSS--IP-PM
Lpol_ey_XP_0222 -------ITS---ACVPPPVTAPS------------LPTENGQIQ-KKEGD--------SGT--ES-S-SRSD-PGG-SGDEDNAARLRLKRKLQRNRTSFTPEQIEALEKEFERTHYPDVFARERLAAKIDLPEARIQAVSVQVWFSNRRAKWRREEKLRNQRRAAEQAAAA--------------------------------------A-ANTAPARLP-LNAGF-PNALYPSIGQPMA----------------SMTDTYSS--IP-PM
Lpol_ey_XP_0222 -------ITP---ACIPPPITASG------------HHSDNGQVQ-KKEGD--------SGN--DS-S-SRSD-PGG-SGDEDNEARLRLKRKLQRNRTSFTPEQIEALEKEFERTHYPDVFARERLAAKIDLPEARIQALSVQVWFSNRRAKWRREEKLRNQRRVAEQAAVAAA------------------------------------A-ANSTPARLS-LNSGF-PNALYPGIGQPMA----------------SMTDSYSS--IP-PM
Tcas_toy_KYB279 -----------------------------------QIARDDLQ-K-RADAM--------HEN--TS-D-GNSE-HNS-SGDEDSQLRLRLKRKLQRNRTSFTNEQIDSLEKEFERTHYPDVFARERLAEKIGLPEARI-----QVWFSNRRAKWRREEKLRNQRRAVDQVGAVS---------------------------------------PPSSAGRLP-INSGF--NSMYSSIPQPIA----------------TMADTYSS--MS---
Dmag_toy_KZS136 -------LP----HGHVPTTPIPSG----------GHHLIRDDPH-KREIP--------SYR--LA-C-GLSL--YL-HI-YCCLLIMQKGSKLARQSTATMTDRLGKKKKNI----APPILKKKWT--------HIG-----RVWFSNRRAKWRREEKLRNQRRGPESTSGSGSSA--GGVGS--S----------------SSAAAAAAA-AAAATGRLNSINSG--LNSMYPSLSQPLA----------------SMAEPYSS--MA--T
Dpul_toy_EFX757 -------LP----HGHVPATPLPSGG--------GGHHLIRDDPH-KRDCN--------DGN--IS-D-GNSDLHNS-SGDEDPQMRLRLKRKLQRNRTSFTNEQIESLEKEFERTHYPDVFARERLAEKIGLPEARI-----QVWFSNRRAKWRREEKLRNQRRGPESTSGGSGGSGGSGVGSSSS----------------SSAAAAAAA-AAAATGRLNSINSG--LNSMYPSLTQPLA----------------SMAEPYSS--MA--T
Ptep_toy_XP_042 AAAAAAAATG---GLVPPVVSSAGLG------VCNGQHNGSDSSA-KKSVN---------EA--TS-D-GNSE-NNS-SADEDSQMRMRLKRKLQRNRTSFANEQIEALEKEFERTHYPDVFARERLAEKISLPEARI-----QVWFSNRRAKWRREEKLRNQRRTVDQVSASS-------------------------------------V-ATPTAGRLP-LNGTF-ANTMYPSLGQPMG----------------AMPDSYSK--DM--S
Lpol_toy_XP_013 NAGLVGP--------VHGGNGHPSLA------ACESHRQDHPS-V-KKDSI---------DA--TS-D-GNSE-NNS-SADEDSQLRMRLKRKLQRNRTSFTNEQIEALEKEFERTHYPDVFARERLAEKITLPEARI-----QVWFSNRRAKWRREEKLRNQRRVVEQPTGTT-------------------------------------V-LAPPNGRLP-INGTF-YNSMYPSLGQPMG----------------GMGDSYSM--PP--S
Lpol_toy_XP_022 NTGLGGS--------VPIVNGHSSLG------SCDSHRHDHMSVT-KKDSI---------DA--TS-D-GNSE-NNS-SADEDSQLRMRLKRKLQRNRTSFTNEQIEALEKEFERTHYPDVFARERLAEKISLPEARI-----QVWFSNRRAKWRREEKLRNQRRVIEQPACTS-------------------------------------M-LSPPNGRLP-INGSF-YNSMYPSLAQPLG----------------GMGDSYSV--PP--A
Mmus_Pax6_NP_00 P----------------------------------SFTMA--NNLP--MQPPVPSQT---------------------------------SSYS------CMLPTSP------------------S----------VN-GR-SYD-T-YTP--PH-------MQTHMNSQ---P-----MGTSGTTSTGLISPGVSVPVQVPGSEPDM-----SQYWPRLQ
Tcas_ey_ABW2313 SSFNHG-------------------------------GLT--SSMGPSPGACLQQRESP--------------------------------SMGGAMSGYCMGR-PP--SYDL------------G----------IG-YG-SRP-SPCSPTQPY-------Q---MNGHQ--------YNTNGTSSTGLISPGVSVPIAVPGQA-----DMTSQYWPRLQ
Dmag_ey_KZS1363 NSMNGFGAMGHHPGGHGAVGPVAAAAMTPVTPAMSHIGAV--PPIGTG-GCSLQQQQQQQQHQ-QLQQQS-SHQAQQPRDHHSAHSAHSSGGYS------CMSRMSG--GYDISAYNRHHPHLSHS----------MG-QV-SGP-I-GPTATSVSNSLAHNYHQSVNSQY-GM-----HHNSHNSATGLISPGVSVPVAVPGQP-TA--EMHHQYWTRLQ
Dpul_ey_EFX7578 NSMTGFSAMGHHPGGHGAVGPVAAAAMTPVTPSMSHIAAV--PPIGTG-PCSLQQQQQQP-HQQQQQQQQPPHQTQQPRDHHSAHSVHSSGGYS------CMSRMSG--GYDISSYNRHHPHLSHA----------MG-QV-SGP-I-GPTATSVSNSLAHNYHQSVNSQY-GM-----HHNSHNSTTGLISPGVSVPVAVPGQP-TA--EMHHQYWTRLQ
Ptep_ey_XP_0159 P----------------------------------TYS----SNMT--PSSCLQQQT---------------------------------TSYS------CMLPPGGTRSYD---------P--LS----------LG-NY-TRP-T-CPTAQPH-------VMQPVNSQF-PS-----VNNGTAASTGLISPGVSVPVQVPGQSNDL--NMASNYWTRLQ
Lpol_ey_XP_0137 S----------------------------------TYSMP--NNMA--PNSCLQQQA---------------------------------SSYS------CMLPPGSARSYD---------H--LS----------LS-SY-SRP-S-CPTAQPY-------VMQPVNGQF-ST-----VNNGGAGSTGLISPGLSVPVQVPGQPGDL--NMSSAYWPRIQ
Lpol_ey_XP_0222 S----------------------------------TYSMS--NNMG--PSSCLQQQA---------------------------------PSYS------CMLPPGSARSYD---------P--LS----------LG-SY-SRP-S-CPTAQPY-------VMQPVNGQF-HT-----VNNASAGSTGLISPGLSVPVQVPAQTGEL--NMSSAYWPRIQ
Lpol_ey_XP_0222 S----------------------------------TYSMS--NNMA--PSSCLQQQT---------------------------------PSYS------CMLPSGSARNYD---------P--LS----------LG-SY-GRP-S-CPTAQPY-------VMQPINGQF-AT-----VNNGSAGSTGLISPGLSVPVQVPSQPGEL--NMSTAYWPRIQ
Tcas_toy_KYB279 ------------------------------------------GG-L--SSSCLQQRDA--------------------------------TGYP------YMF-------HD---------P--LHS---------LSSSYNSRATA-CNPAVAH-------TQPT--THPSYASATTPTSVPASTGTGVISAGVSVPVQIPSQTPDLSTNLASNYWPRIQ
Dmag_toy_KZS136 -----------------------------------SLSTM--GGMG--GSCGLQQRD---------------------------------PATAAAYPS-YMFAHH---GGD---------P--LSSLQSSYNQHQRA-AA-AAA-A-CSAAQAS-------SAAS--AHS-YA-----AASSAHAQTGVISAGVSVPLQIPSHQG-S--DIAANYWHRLQ
Dpul_toy_EFX757 -----------------------------------SLSSM--GGMG--GSCGLQQRD---------------------------------PATASAYPS-YMFAHH---GGD---------P--LSSLQSSYNQHQRA-AA-AAA-A-CSAAQAS-------SVAS--AHS-YA-----T--SAH-AQTVVEEIV--------------------------
Ptep_toy_XP_042 S----------------------------------FYSMAPVSGMT--PSSCLQQRDP--------------------------------GAYS------YML-------HD---------T--LS----------LG-AY-SRA-S-CTPAQPM-------N-----THPSYA-----VSGASPHSAGVISPGVTVPVQVPGQTPDI--SVTSNYWPRIQ
Lpol_toy_XP_013 ------------------------------------------SSMT--SNHCLQQRDP--------------------------------TSYT------YMF-------HD---------P--LS----------LG-SY-SRS-S-CTPSQSI-------N-----GHPSYS-----VNGGNAHGAGVISPGVTVPLQVPSQGPDL--SMATNYWHRIQ
Lpol_toy_XP_022 ------------------------------------------SSVA--SNHCLQQRDP--------------------------------SSYT------YMF-------HD---------Q--LT----------LG-SY-SRP-S-CTPSQPI-------N-----GHASYS-----VSASSPHGAGVISPGVTVPLQVPSQGPDL--SMATNYWHRIQ
Using the unusually gene duplication rich horseshoe crab genome
to define protein sequence domains that distinguish the Pax6 paralogs
eyeless and twin of eyeless across the arthropods
Abstract
The Pax6 gene is well known for its importance in developing the
visual system. The gene is able to, in the early stages of the
embryo, place eye location, and control eye function. Many
species of arthropods and arachnids normally possess two copies
of the Pax6 gene called the eyeless and the twin of the eyeless
(Yang 2009). Both genes are essential for the development of the
ocular head segment (Luan et al. 2014). The horseshoe crab
Limulus polyphemus has more Pax6 genes than most arthropods
(Friedrich, 2022), i.e., two homologs of eyeless and three
homologs of twin of eyeless. Horseshoe crabs represent an
ancient chelicerate lineage and known as some of the world's
living fossils are over 445 million years old. In this project we were
studying the horseshoe crab eyeless and twin of eyeless protein
sequences to identify sequence regions that distinguish the two
closely related paralogs.
We compared the protein sequences of eyeless and twin of eyeless
homologs from different species, using different programs obtain a protein
alignment. All sequences were acquired from the National Center of
Biotechnology Information (https://www.ncbi.nlm.nih.gov/).
Our results showed that there is a distinct differences between the Eyeless
and Twin of eyeless gene. By comparing the proteins between several
species, we found several proteins arrangements that were only presents to
that particular gene.
Approach
Findings
Katelynne Haygood and Markus Friedrich, Wayne State University, Detroit, MI 48202
● Mus musculus (mouse)
● Lpol = Limulus polyphemus = Horseshoe
crab
● Tcas = Tribolium castaneum (beetle)
● Dpul = Daphnia pulex (water flea)
● Dmag = Daphnia magna (water flea)
● Ptep = Parasteatoda tepidariorum (spider)
Future Directions
We plan to add more Pax gene sequences for comparison and explore the
conservation of our currently identified eyeless and twin of eyeless specific
sequence sites. Interestingly, there are three additional species of
horseshoe crabs besides Limulus polyphemus which may have evolved
differently from each with respect to Pax6 gene numbers.
Acknowledgments
We would like to thank the MARC program for support and our group
members Mohamad Kassem, Tanay Dakarapu, and Zainab Jamshed for
input and discussion.
Fan, Zheng, et al. “A Chromosome-Level Genome of the Spider Trichonephila Antipodiana Reveals the Genetic Basis of Its Polyphagy and Evidence of an Ancient Whole-Genome Duplication Event.” GigaScience, vol. 10, no. 3,
2021, https://doi.org/10.1093/gigascience/giab016.
Friedrich, Markus. “Ancient Genetic Redundancy of Eyeless and Twin of Eyeless in the Arthropod Ocular Segment.” Science Direct, Academic Press, 6 Oct. 2017,
https://www.sciencedirect.com/science/article/pii/S0012160617306279.
Yang, Xiaoyun, et al. “Probing the Drosophila Retinal Determination Gene Network in Tribolium (II): The PAX6 Genes Eyeless and Twin of Eyeless.” Developmental Biology, vol. 333, no. 1, 2009, pp. 215–227.,
https://doi.org/10.1016/j.ydbio.2009.06.013.
References