De novo deletion within the telomeric region flanking the human a globin locus as a cause of a thalassaemia Vip Viprakasit, 1,2 Alexa M. J. Kidd, 3 * Helena Ayyub, 1 Sharon Horsley, 1  Jim Hughes 1 and Douglas R. Higgs 1 1 MRC Molecular Haematology Unit, Weatherall Institute of Molecular Medicine, John Radcliffe Hospital, University of Oxford, UK, 2 Department of Paediatrics, Faculty of Medicine, Siriraj Hospital, Mahidol University, Bangkok, Thailand, and 3 Department of Medical Genetics, Grampian University Hospital, University of Aberdeen, Foresterhill, Aberdeen, UK Received 24 August 2002; accepted for publication 7 November 2002 Summary. We have identified and characterized a Scot- tish individual with a thalassaemia, resulting from a de novo 48 kilobase (kb) deletion from the telomeric flanking region of the a globin cluster which occurred as a result of recombination between two misaligned repet- itive elements that normally lie 83 kb and 131 kb from the 16p telomere. The deletion removes two previously described putative regulatory elements (HS-40 and HS-33) but leaves two other elements (HS-10 and HS-8) intact. Analysis of this deletion, together with eight other pub- lished deletions of the telomeric region, showed that they all severely downregulated a globin expression. Together they defined a 20Æ4-kb region of the human a cluster, which contains all of the positive cis-acting elements required to regulate a globin expression. Comparative analysis of this region with the corresponding segment of the mouse a globin cluster demonstrated conserved non- coding sequences corresponding to the putative regulatory elements HS-40 and HS-33. Although the role of HS-40 as an enhancer of a globin expression is fully established, these observations suggest that the role of HS-33 and other sequences in this region should be more fully investigated in the context of the natural human and mouse a globin loci. Keywords: a thalassaemia, HS-40, HS-33a major regulatory element, Alu-associated homologous recombination. The human a and b globin gene clusters have been among the most intensively studied of all mammalian multigene families, not least because mutations which downregulate globin gene expression cause a and b thalassaemia, which are the most common single gene disorders throughout all tropical and subtropical regions of the world (Steinberg et al, 2001). The human a globin gene cluster includes an embryonic (f) and two fetal/adult genes (a2 and a1) lying close [165 kilobase (kb)] to the subtelomeric region of the short arm of chromosome 16 arranged in the order, telomere-f-a2-a1-centromere. The b globin cluster includes an embryonic gene (e), two fetal (c) genes and one major adult gene (b) within the short arm of chromosome 11, arranged telomere-e-c-c-b-centromere. Thalassaemia most frequently results from deletions or point mutations affect- ing the structural a and b globin genes. However, three unique, sporadic mutations which cause b thalassaemia remove sequences located upstream of the b cluster but leave the structural b gene intact (Wood, 2001). All three deletions remove the five regulatory elements, associated with DNAse I hypersensitive sites (called HS1-5), which are collectively referred to as the b globin locus control region (bLCR) (Tuan & London, 1984; Forrester et al, 1987; Grosveld et al, 1987). These deletions were crucially important in first identifying the bLCR. We have previously identified eight sporadic deletions, each affecting single families, that similarly remove sequences lying upstream of the a globin cluster while leaving the structural a genes intact (Hatton et al, 1990; Liebhaber et al, 1990; Wilkie et al, 1990a; Romao et al, 1991, 1992; Flint et al, 1994, 1996). The situation in the a cluster is somewhat different from that in the b cluster. First, although there are at least four erythroid-specific Correspondence: D. R. Higgs, MRC Molecular Haematology Unit, Weatherall Institute of Molecular Medicine, John Radcliffe Hospital, Headington, University of Oxford, Oxford, OX3 9DS, UK. E-mail: drhiggs@molbiol.ox.ac.uk *Present address: Department of Medical Genetics, Wellington Hospital, New Zealand.  Present address: Leukaemia Research Fund Centre, Institute of Cancer Research, Chester Beatty Laboratories, London, UK. British Journal of Haematology, 2003, 120, 867–875 Ó 2003 Blackwell Publishing Ltd 867