vavilovВавиловский журнал генетики и селекцииVavilov Journal of Genetics and Breeding2500-3259Institute of Cytology and Genetics of Siberian Branch of the RAS10.18699/VJ18.342vavilov-1375Research ArticleМОЛЕКУЛЯРНЫЕ МАРКЕРЫ В ГЕНЕТИКЕ И СЕЛЕКЦИИBIOINFORMATICS AND SYSTEM BIOLOGYЭКСПЕРИМЕНТАЛЬНОЕ ИЗУЧЕНИЕ ВЛИЯНИЯ РЕДКИХ ПОЛИМОРФИЗМОВ ТАТА-БОКСОВ ПРОМОТОРОВ ГЕНОВ HBB, HBD И F9 ЧЕЛОВЕКА НА КИНЕТИКУ ВЗАИМОДЕЙСТВИЯ С ТАТА-СВЯЗЫВАЮЩИМ БЕЛКОМAN EXPERIMENTAL STUDY OF THE EFFECT OF RARE POLYMORPHISMS OF HUMAN HBB, HBD AND F9 PROMOTER TATA BOXES ON THE KINETICS OF INTERACTION WITH THE TATA-BINDING PROTEINШарыповаЕ. Б.SharypovaE. B.

Новосибирск

Novosibirsk

ДрачковаИ. А.DrachkovaI. A.

Новосибирск

Novosibirsk

КашинаЕ. В.KashinaE. V.

Новосибирск

Novosibirsk

РассказовД. А.RasskazovD. A.

Новосибирск

Novosibirsk

ПономаренкоП. М.PonomarenkoP. M.ПономаренкоМ. П.PonomarenkoM. P.

Новосибирск

Novosibirsk

КолчановН. А.KolchanovN. А.СавинковаЛ. К.SavinkovaL. K.

Новосибирск

Novosibirsk

savinkl@mail.ruФедеральный исследовательский центр Институт цитологии и генетики Сибирского отделения Российской академии наукРоссияInstitute of Cytology and Genetics SB RASRussian FederationУниверситет Ла-ВернаСоединённые Штаты АмерикиUniversity of La VerneUnited StatesФедеральный исследовательский центр Институт цитологии и генетики Сибирского отделения Российской академии наук; Новосибирский национальный исследовательский государственный университетРоссияInstitute of Cytology and Genetics SB RAS; Novosibirsk State UniversityRussian Federation201822032018221145152Copyright © Шарыпова Е.Б., Драчкова И.А., Кашина Е.В., Рассказов Д.А., Пономаренко П.М., Пономаренко М.П., Колчанов Н.А., Савинкова Л.К., 20182018Шарыпова Е.Б., Драчкова И.А., Кашина Е.В., Рассказов Д.А., Пономаренко П.М., Пономаренко М.П., Колчанов Н.А., Савинкова Л.К.Sharypova E.B., Drachkova I.A., Kashina E.V., Rasskazov D.A., Ponomarenko P.M., Ponomarenko M.P., Kolchanov N.А., Savinkova L.K.This work is licensed under a Creative Commons Attribution 4.0 License.https://vavilov.elpub.ru/jour/article/view/1375

Гены человека HBB, HBD и F9 относятся к системе кроветворения. Недостаток или избыток продуктов этих генов является причиной наследственных талассемий разной тяжести и гемофилии В Лейдена. Ранее нами было показано, что ряд аннотированных однонуклеотидных полиморфизмов (SNPs) ТАТА-боксов этих генов, ассоциированных с возникновением ß-, δ-талассемий и гемофилии В Лейдена влияет на взаимодействие с ТАТА-связывающим белком (TBP), изменение взаимодействия пропорционально изменению количества продуктов этих генов. В настоящей работе исследуется влияние редких неаннотированных SNPs ТАТА-боксов этих генов на взаимодействие ТАТА-связывающего белка. Для изучения кинетических характеристик образования комплексов ТВР/ТАТА in vitro используются двуцепочечные олигодезоксинуклеотиды, идентичные ТАТА-содержащим участкам промоторов генов HBB, HBD и F9 (нормальным и минорным аллелям), и рекомбинантный ТВР человека. Показано, что замена –25A > C (rs281864525) и делеция –25АА (rs63750953) ТАТА-бокса гена ß-глобина одинаково влияют на аффинность ТВР/ТАТА, которая падает в обоих случаях в три раза, но значительно различается влияние этих SNPs на скорость образования комплексов ТВР/ТАТА: –25A > C приводит к снижению скорости в пять раз, а делеция (–25AA) – к снижению скорости более чем в семь раз. Различное влияние замен оказывается и на прочность комплексов ТВР/ТАТА. Если в случае –25A > C прочность комплексов увеличивается в 1.8 раза, то при делеции –25АА прочность комплексов возрастает в 2.4 раза, несмотря на уменьшение сродства ТАТА-связывающего белка к ТАТА-боксу. Сравнение изменения экспериментальных значений сродства (KD) ТВР/ТАТА нормальных и минорных аллелей c прогнозируемыми показало, что данные хорошо коррелируют друг с другом: коэффициент линейной корреляции r = 0.94 (α < 0.0001). Комплексный подход к изучению редких полиморфизмов может привести к определению наиболее чувствительных маркеров орфанных заболеваний, что внесет вклад в разработку надежных и быстрых методов их диагностирования и лечения.

Human genes HBB, HBD and F9 belong to the hematopoiesis system. The deficiency or excess of these genes’ products is the cause of hereditary thalassemias of various severity and haemophilia B Leyden. Previously, it was shown that a number of annotated single-nucleotide polymorphisms of TATA boxes of these genes associated with the occurrence of ß- and δ-thalassemia affect the interaction with the TATAbinding protein, the interaction changing proportionally with the change in the number of gene products. In the present work, we investigate the effect of rare not annotated single-nucleotide polymorphisms (SNPs) of TATA boxes of these genes with an unknown manifestation on the TATA-binding protein interaction. To study the kinetic characteristics of TBP/TATA complex formation in vitro, doublestranded oligodeoxynucleotides identical to the TATA-containing portions of the promoters of the HBB, HBD and F9 genes (“normal” and minor alleles) and recombinant human TBP were used. It was shown that the TATA-box SNP of –25A > C (rs281864525) and the deletion of the –25AA (rs63750953) TATA-box of the β-globin gene have the same effect on the TBP/TATA affinity, which decreases 3-folds in both cases. However, the effect of these substitutions on the rate of the TBP/TATA complex formation is significantly different: SNP –25A > C decreases the rate 5-fold, and the deletion decreases the rate more than 7-fold. The influence of substitutions on the strength of the TBP/TATA complexes has a different effect. If in the case of SNP –25A > C the strength of the complexes increases 1.8-fold, then in the case of the –25AA deletion, the strength of the complexes increases 2.4-fold, even though the affinity of the TATAbinding protein to the TATA box decreases. A comparison of experimental values of affinity (KD) of the TBP/T complexes of “normal” and minor alleles with the predicted has shown that data correlate well with each other. The coefficient of linear correlation r = 0.94 (α < 0.0001). A comprehensive approach to the study of rare polymorphisms may lead to the identification of the most sensitive markers of orphan diseases, which will contribute to the development of reliable and rapid methods for their diagnosis and treatment.

однонуклеотидный полиморфизмгены кроветворенияредкие полиморфизмыорфанные заболеванияпромоторТАТА-боксsingle nucleotide polymorphismhematopoies genesrare polymorphismsorphan diseasespromoterTATA-boxState Budgeted Project 03242018-0019; The development of the service pack of the Web service was supported by State Budgeted Project 03242018-0017; Computerized data analysis was supported by the Russian Federation Ministry of Education and Science, Program for improving the competitiveness of leading Russian universities among leading research and educational centers in the world (Project 5-100) and Integration Project 03242018-0021ReferencesBank A., O’Neill D., Lopez R., Pulte D., Ward M., Mantha S., Richardson C. Role of intergenic human γ-δ-globin sequences in human hemoglobin switching and reactivation of fetal hemoglobin in adult erythroid cells. Ann. N. Y. Acad. Sci. 2005;1054:48-54.Bank A., O’Neill D., Lopez R., Pulte D., Ward M., Mantha S., Richardson C. Role of intergenic human γ-δ-globin sequences in human hemoglobin switching and reactivation of fetal hemoglobin in adult erythroid cells. Ann. N. Y. Acad. Sci. 2005;1054:48-54.Bell C.J., Dinwiddie D.L., Miller N.A., Hateley S.L. Carrier testing for severe childhood recessive diseases by next-generation sequencing. Sci. Transl. Med. 2011;3:65ra4. DOI 10.1126/scitranslmed.3001756.Bell C.J., Dinwiddie D.L., Miller N.A., Hateley S.L. Carrier testing for severe childhood recessive diseases by next-generation sequencing. Sci. Transl. Med. 2011;3:65ra4. DOI 10.1126/scitranslmed.3001756.Cavalli M., Pan G., Nord H., Wallerman O., Arzt E.W., Berggren O., Elvers I., Eloranta M.-L., Rönnblom L., Toh K.L., Wadelius C. Allelespecific transcription factor binding to common and rare variants associated with disease and gene expression. Hum. Genet. 2016;135: 485-497.Cavalli M., Pan G., Nord H., Wallerman O., Arzt E.W., Berggren O., Elvers I., Eloranta M.-L., Rönnblom L., Toh K.L., Wadelius C. Allelespecific transcription factor binding to common and rare variants associated with disease and gene expression. Hum. Genet. 2016;135: 485-497.Dharssi S., Wong-Rieger D., Harold M., Terry S. Review of 11 national policies for rare diseases in the context of key patient needs. Orphanet J. Rare Dis. 2017;12:63.Dharssi S., Wong-Rieger D., Harold M., Terry S. Review of 11 national policies for rare diseases in the context of key patient needs. Orphanet J. Rare Dis. 2017;12:63.Dooms M. From promising molecules to orphan drugs: Early clinical drug development. Intractable Rare Dis. Res. 2017;(1):29-34. DOI 10.5582/irdr.2016.01094.Dooms M. From promising molecules to orphan drugs: Early clinical drug development. Intractable Rare Dis. Res. 2017;(1):29-34. DOI 10.5582/irdr.2016.01094.Drachkova I.A., Arshinova T.V., Ponomarenko P.M., Merkulova T.I., Kolchanov N.A., Savinkova L.K. Effect of Tata box polymorphisms in the human β-globin gene promoter associated with β-thalassemia on the interaction of the Tata-binding protein. Vestnik VOGiS = Herald Vavilov Society for Geneticists Breeding Scientists. 2010;14(4):698-705. (in Russian)Drachkova I.A., Arshinova T.V., Ponomarenko P.M., Merkulova T.I., Kolchanov N.A., Savinkova L.K. Effect of Tata box polymorphisms in the human β-globin gene promoter associated with β-thalassemia on the interaction of the Tata-binding protein. Vestnik VOGiS = Herald Vavilov Society for Geneticists Breeding Scientists. 2010;14(4):698-705. (in Russian)Drachkova I., Savinkova L., Arshinova T., Ponomarenko M., Peltek S., Kolchanov N. The mechanism by which TATA-box polymorphisms associated with human hereditary diseases influence interactions with the TATA-binding protein. Hum. Mutat. 2014;35(5):601-608. DOI 10.1002/humu.22535.Drachkova I., Savinkova L., Arshinova T., Ponomarenko M., Peltek S., Kolchanov N. The mechanism by which TATA-box polymorphisms associated with human hereditary diseases influence interactions with the TATA-binding protein. Hum. Mutat. 2014;35(5):601-608. DOI 10.1002/humu.22535.Frischknecht H., Dutly F. Two new delta-globin mutations: Hb A2-Ninive [!delta133(H11)Val-Ala] and a delta(+)-thalassemia mutation [-31 (A→G)] in the TATA box of the delta-globin gene. Hemoglobin. 2005;29(2):151-154.Frischknecht H., Dutly F. Two new delta-globin mutations: Hb A2-Ninive [!delta133(H11)Val-Ala] and a delta(+)-thalassemia mutation [-31 (A→G)] in the TATA box of the delta-globin gene. Hemoglobin. 2005;29(2):151-154.Galanello R., Origa R. Beta-thalassemia. Orphanet J. Rare Dis. 2010; 5:11.Galanello R., Origa R. Beta-thalassemia. Orphanet J. Rare Dis. 2010; 5:11.Haeussler M., Raney B.J., Hinrichs A.S., Clawson H., Zweig A.S., Karolchik D., Casper J., Speir M.L., Haussler D., Kent W.J. Navigating protected genomics data with UCSC genome browser in a box. Bioinformatics. 2015;31(5):764-766. DOI 10.1093/bioinformatics/ btu712.Haeussler M., Raney B.J., Hinrichs A.S., Clawson H., Zweig A.S., Karolchik D., Casper J., Speir M.L., Haussler D., Kent W.J. Navigating protected genomics data with UCSC genome browser in a box. Bioinformatics. 2015;31(5):764-766. DOI 10.1093/bioinformatics/ btu712.Keinan A., Clark A.G. Recent explosive human population growth has resulted in an excess of rare genetic variants. Science. 2012;336: 740-743.Keinan A., Clark A.G. Recent explosive human population growth has resulted in an excess of rare genetic variants. Science. 2012;336: 740-743.Kheradpour P., Ernst J., Melnikov A., Rogov P., Wang L., Zhang X., Alston J., Mikkelsen T.S., Kellis M. Systematic dissection of regulatory motifs in 2000 predicted human enhancers using a massively parallel reporter assay. Genome Res. 2013;23(5):800-811. DOI 10.1101/gr.144899.112.Kheradpour P., Ernst J., Melnikov A., Rogov P., Wang L., Zhang X., Alston J., Mikkelsen T.S., Kellis M. Systematic dissection of regulatory motifs in 2000 predicted human enhancers using a massively parallel reporter assay. Genome Res. 2013;23(5):800-811. DOI 10.1101/gr.144899.112.Kolchanov N.A., Ponomarenko P.M., Savinkova L.K., Drachkova I.A., Lysova M.V., Arshinova T.V., Ponomarenko M.P. A step-by-step model of TBP/TATA box binding allows predicting human hereditary diseases by single nucleotide polymorphism. Dokl. Biochem. Biophys. 2008;419:88-92.Kolchanov N.A., Ponomarenko P.M., Savinkova L.K., Drachkova I.A., Lysova M.V., Arshinova T.V., Ponomarenko M.P. A step-by-step model of TBP/TATA box binding allows predicting human hereditary diseases by single nucleotide polymorphism. Dokl. Biochem. Biophys. 2008;419:88-92.Kurachi K., Davie E.W. Isolation and characterization of a cDNA coding for human factor IX. Proc. Natl. Acad. Sci. USA. 1982;79(21):64616464.Kurachi K., Davie E.W. Isolation and characterization of a cDNA coding for human factor IX. Proc. Natl. Acad. Sci. USA. 1982;79(21):64616464.Levings P.P., Bungert J. The human beta-globin locus control region. Eur. J. Biochem. 2002;269:1589-1599.Levings P.P., Bungert J. The human beta-globin locus control region. Eur. J. Biochem. 2002;269:1589-1599.MacArthur D.G., Manolio T.A., Dimmock D.P., Rehm H.L., Shendure J., Abecasis G.R., Adams D.R., Altman R.B., Antonarakis S.E., Ashley E.A., Barrett J.C., Biesecker L.G., Conrad D.F., Cooper G.M., Cox N.J., Daly M.J., Gerstein M.B., Goldstein D.B., Hirschhorn J.N., LealS.M., PennacchioL.A., StamatoyannopoulosJ.A., SunyaevS.R., Valle D., Voight B.F., Winckler W., Gunter C. Guidelines for investigating causality of sequence variants in human disease. Nature. 2014;508(7497):469-476. DOI 10.1038/nature13127.MacArthur D.G., Manolio T.A., Dimmock D.P., Rehm H.L., Shendure J., Abecasis G.R., Adams D.R., Altman R.B., Antonarakis S.E., Ashley E.A., Barrett J.C., Biesecker L.G., Conrad D.F., Cooper G.M., Cox N.J., Daly M.J., Gerstein M.B., Goldstein D.B., Hirschhorn J.N., LealS.M., PennacchioL.A., StamatoyannopoulosJ.A., SunyaevS.R., Valle D., Voight B.F., Winckler W., Gunter C. Guidelines for investigating causality of sequence variants in human disease. Nature. 2014;508(7497):469-476. DOI 10.1038/nature13127.Moleirinho A., Seixas S., Lopes A.M., Bento C., Prata M.J., Amorim A. Evolutionary сonstraints in the β-globin cluster: the signature of purifying selection at the δ-globin (HBD) locus and its role in developmental gene regulation. Genome Biol. Evol. 2013;5(3):559-571. DOI 10.1093/gbe/evt029.Moleirinho A., Seixas S., Lopes A.M., Bento C., Prata M.J., Amorim A. Evolutionary сonstraints in the β-globin cluster: the signature of purifying selection at the δ-globin (HBD) locus and its role in developmental gene regulation. Genome Biol. Evol. 2013;5(3):559-571. DOI 10.1093/gbe/evt029.Persyn E., Karakachoff M., Scouarnec S. Le, Clézio C. Le, Campion D., French Exome Consortium Schott J.-J., Redon R., Bellanger L., Dina C. DoEstRare: A statistical test to identify local enrichments in rare genomic variants associated with disease. PLoS ONE. 12(7). DOI 10.1371/journal.pone.0179364.Persyn E., Karakachoff M., Scouarnec S. Le, Clézio C. Le, Campion D., French Exome Consortium Schott J.-J., Redon R., Bellanger L., Dina C. DoEstRare: A statistical test to identify local enrichments in rare genomic variants associated with disease. PLoS ONE. 12(7). DOI 10.1371/journal.pone.0179364.Picketts D.J., Mueller C.R., Lillicrap D. Transcriptional control of the factor IX gene: analysis of five cis-acting elements and the deleterious effects of naturally occurring hemophilia B Leyden mutations. Blood. 1994;84(9):2992-3000.Picketts D.J., Mueller C.R., Lillicrap D. Transcriptional control of the factor IX gene: analysis of five cis-acting elements and the deleterious effects of naturally occurring hemophilia B Leyden mutations. Blood. 1994;84(9):2992-3000.Ponomarenko M., Rasskazov D., Arkova O., Ponomarenko P., Suslov V., Savinkova L., Kolchanov N. How to use SNP_TATA_Comparator to find a significant change in gene expression caused by the regulatory SNP of this gene’s promoter via a change in affinity of the TATA-binding protein for this promoter. Biomed. Res. Int. 2015; 2015:359835. DOI 10.1155/2015/359835.Ponomarenko M., Rasskazov D., Arkova O., Ponomarenko P., Suslov V., Savinkova L., Kolchanov N. How to use SNP_TATA_Comparator to find a significant change in gene expression caused by the regulatory SNP of this gene’s promoter via a change in affinity of the TATA-binding protein for this promoter. Biomed. Res. Int. 2015; 2015:359835. DOI 10.1155/2015/359835.Savinkova L., Drachkova I., Arshinova T., Ponomarenko P., Ponomarenko M., Kolchanov N. An experimental verification of the predicted effects of promoter TATA-box polymorphisms associated with human diseases on interactions between the TATA boxes and TATA-binding protein. PLoS ONE. 2013;8(2):22-23.Savinkova L., Drachkova I., Arshinova T., Ponomarenko P., Ponomarenko M., Kolchanov N. An experimental verification of the predicted effects of promoter TATA-box polymorphisms associated with human diseases on interactions between the TATA boxes and TATA-binding protein. PLoS ONE. 2013;8(2):22-23.Schechter A.N. Hemoglobin research and the origins of molecular medicine. Blood. 2008;112(10):3927-3938. DOI 10.1182/blood-200804-078188.Schechter A.N. Hemoglobin research and the origins of molecular medicine. Blood. 2008;112(10):3927-3938. DOI 10.1182/blood-200804-078188.Sherry S.T., Ward M.H., Kholodov M., Baker J., Phan L., Smigielski E.M., Sirotkin K. dbSNP: the NCBI database of genetic variation. Nucleic Acids Res. 2001;29(1):308-311. DOI 10.1093/nar/29.1.308.Sherry S.T., Ward M.H., Kholodov M., Baker J., Phan L., Smigielski E.M., Sirotkin K. dbSNP: the NCBI database of genetic variation. Nucleic Acids Res. 2001;29(1):308-311. DOI 10.1093/nar/29.1.308.Steinberg M., Adams J.G. 3rd Hemoglobin A2: origin, evolution, and aftermath. Blood. 1991;78:2165-2177.Steinberg M., Adams J.G. 3rd Hemoglobin A2: origin, evolution, and aftermath. Blood. 1991;78:2165-2177.Wu J., Wu M., Li L., Liu Z., Zeng W., Jiang R. dbWGFP: a database and web server of human whole-genome single nucleotide variants and their functional predictions. Database (Oxford). 2016;pii: baw024. DOI 10.1093/database/baw024.Wu J., Wu M., Li L., Liu Z., Zeng W., Jiang R. dbWGFP: a database and web server of human whole-genome single nucleotide variants and their functional predictions. Database (Oxford). 2016;pii: baw024. DOI 10.1093/database/baw024.

The authors declare that there are no conflicts of interest present.