We describe the molecular etiology of β(+)-thalassemia that is caused by the insertion of the full-length transposable element LINE-1 (L1) into the intron-2 of the β-globin gene (HBB). The transcript level of the affected β-globin gene was severely reduced. The remaining transcripts consisted of full-length, correctly processed β-globin mRNA and a minute amount of three aberrantly spliced transcripts with a decreased half-life due to activation of the nonsense-mediated decay pathway. The lower steady-state amount of mRNA produced by the β-globin(L1) allele also resulted from a reduced rate of transcription and decreased production of full-length β-globin primary transcripts. The promoter and enhancer sequences of the β-globin(L1) allele were hypermethylated; however, treatment with a demethylating agent did not restore the impaired transcription. A histone deacetylase inhibitor partially reactivated the β-globin(L1) transcription despite permanent β-globin(L1) promoter CpG methylation. This result indicates that the decreased rate of transcription from the β-globin(L1) allele is associated with an altered chromatin structure. Therefore, the molecular defect caused by intronic L1 insertion in the β-globin gene represents a novel etiology of β-thalassemia.

Department of Biology Faculty of Medicine and Dentistry Palacky University Olomouc Czech Republic; Departments of Medicine Pathology and Genetics University of Utah and Medical Service George E Wahlen Department of Veterans Affairs Medical Center Salt Lake City Utah

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Antonarakis SE, Kazazian HH, Jr, Orkin SH. DNA polymorphism and molecular pathology of the human globin gene clusters. Hum Genet. 1985;69:1–14. PubMed

Boehringer RR, Hammer RE, Brinster RL, Palmiter RD, Townes TM. Two 3′ sequences direct adult erythroid-specific expression of human β-globin genes in transgenic mice. Proc Natl Acad Sci USA. 1987;84:7056–7060. PubMed PMC

Divoky V, Indrak K, Mrug M, Brabec V, Huisman THJ, Prchal JT. A novel mechanism of β-thalassemia. The insertion of L1 retrotransposable element into β globin IVSII. Blood. 1996;88:148a. (abstract)

Garcia-Perez JL, Morell M, Scheys JO, Kulpa DA, Carter CC, Hammer GD, Collins KL, O’Shea KS, Menendez P, Moran JV. Epigenetic silencing of engineered L1 retrotransposition events in human embryonic carcinoma cells. Nature. 2010;4669:769–773. PubMed PMC

Han JS, Szak ST, Boeke JD. Transcriptional disruption by the L1 retrotransposon and implications for mammalian transcriptomes. Nature. 2004;429:268–274. PubMed

Hancks DC, Kazazian HH., Jr Active human retrotransposons: variation and disease. Curr Opin Genet Dev. 2012;22:191–203. PubMed PMC

Indrak K, Brabec V, Indrakova J, Chrobak L, Sakalova A, Jarosova M, Cermak J, Fei YJ, Kutlar F, Gu YC, Baysal E, Huisman THJ, et al. Molecular characterization of β-thalassemia in Czechoslovakia. Hum Genet. 1992;88:399–404. PubMed

Kimberland ML, Divoky V, Prchal JT, Schwahn U, Berger W, Kazazian HH., Jr Full-length human L1 insertions retain the capacity for high frequency retrotransposition in cultured cells. Hum Mol Genet. 1999;8:1557–1560. PubMed

Mabaera R, Greene MR, Richardson CA, Conine SJ, Kozul CD, Lowrey CH. Neither DNA hypomethylation nor changes in the kinetics of erythroid differentiation explain 5-azacytidine’s ability to induce human fetal hemoglobin. Blood. 2008;111:411–420. PubMed PMC

Mabaera R, Richardson CA, Johnson K, Hsu M, Fiering S, Lowrey CH. Developmental- and differentiation-specific patterns of human gamma- and betaglobin promoter DNA methylation. Blood. 2007;110:1343–1352. PubMed PMC

Montoya-Durango DE, Liu Y, Teneng I, Kalbfleisch T, Lacy ME, Steffen MC, Ramos KS. Epigenetic control of mammalian LINE-1 retrotransposon by retinoblastoma proteins. Mutat Res. 2009;665:20–28. PubMed PMC

Orkin SH, Kazazian HH, Jr, Antonarakis SE, Goff SC, Boehm CD, Sexton JP, Waber PG, Giardina PJ. Linkage of β-thalassaemia mutations and β-globin gene polymorphisms with DNA polymorphisms in human β-globin gene cluster. Nature. 1982;296:627–631. PubMed

Papayannopoulou T, Brice M, Stamatoyannopoulos G. Analysis of human hemoglobin switching in MEL × human fetal erythroid cell hybrids. Cell. 1986;46:469–476. PubMed

Pfaffl MW. A new mathematical model for relative quantification in real-time RT-PCR. Nucleic Acids Res. 2001;29:e45. PubMed PMC

Qin T, Jelinek J, Si J, Shu J, Issa JP. Mechanisms of resistance to 5-aza-2′-deoxycytidi-ne in human cancer cell lines. Blood. 2009;113:659–667. PubMed PMC

Swank RA, Skarpidi E, Papayannopoulou T, Stamatoyannopoulos G. The histone deacetylase inhibitor, trichostatin A, reactivates the developmentally silenced gamma globin expression in somatic cell hybrids and induces gamma gene expression in adult BFUe cultures. Blood Cells Mol Dis. 2003;30:254–257. PubMed

Wheelan SJ, Aizawa Y, Han JS, Boeke JD. Gene-breaking: a new paradigm for human retrotransposon-mediated gene evolution. Genome Res. 2005;15:1073–1078. PubMed PMC

Whitelaw E, Martin DI. Retrotransposons as epigenetic mediators of phenotypic variation in mammals. Nat Genet. 2001;27:361–365. PubMed

Wu J, Grindlay GJ, Bushel P, Mendelsohn L, Allan M. Negative regulation of the human epsilon-globin gene by transcriptional interference: role of an Alu repetitive element. Mol Cell Biol. 1990;10:1209–1216. PubMed PMC

Yang C, Feng J, Song W, Wang J, Tsai B, Zhang Y, Scaringe WA, Hill KA, Margaritis P, High KA, Sommer SS. A mouse model for nonsense mutation bypass therapy shows a dramatic multiday response to geneticin. Proc Natl Acad Sci USA. 2007;104:15394–15399. PubMed PMC