L1 sequences are a human-specific family of long, interspersed, repetitive elements, present as approximately 10(5) copies dispersed throughout the genome. The full-length L1 sequence is 6.1 kilobases, but the majority of L1 elements are truncated at the 5' end, resulting in a fivefold higher copy number of 3' sequences. The nucleotide sequence of L1 elements includes an A-rich 3' end and two long open reading frames (orf-1 and orf-2), the second of which encodes a potential polypeptide having sequence homology with the reverse transcriptases. This structure suggests that L1 elements represent a class of non-viral retrotransposons. A number of L1 complementary DNAs, including a nearly full-length element, have been isolated from an undifferentiated teratocarcinoma cell line. We now report insertions of L1 elements into exon 14 of the factor VIII gene in two of 240 unrelated patients with haemophilia A. Both of these insertions (3.8 and 2.3 kilobases respectively) contain 3' portions of the L1 sequence, including the poly (A) tract, and create target site duplications of at least 12 and 13 nucleotides of the factor VIII gene. In addition, their 3'-trailer sequences following orf-2 are nearly identical to the consensus sequence of L1 cDNAs (ref. 6). These results indicate that certain L1 sequences in man can be dispersed, presumably by an RNA intermediate, and cause disease by insertional mutation.

Haemophilia A is a common disorder of blood coagulation caused by a deficiency of factor VIII. It is inherited as an X-linked recessive trait, and one-third of all cases are thought to result from de novo mutations. The clinical severity of haemophilia A varies markedly among different families and a subset of the patients with severe disease develop antibodies against factor VIII, called inhibitors. Because of this heterogeneity, it is likely that many different molecular lesions result in haemophilia A. Indeed, of the nine mutations described to date, all appear to be unique changes. However in this study of 83 patients with haemophilia A we have identified two different point mutations, one in exon 18 and one in exon 22, that have recurred independently in unrelated families. Each mutation produces a nonsense codon by a change of CG to TG, and each occurred de novo on the X-chromosome donated by the maternal grandfather. These observations strongly support the view that CpG dinucleotides are mutation hotspots.

Hemophilia A is an X-linked disorder of coagulation caused by a deficiency of factor VIII. By using cloned DNA probes, we have characterized the following five different partial deletions of the factor VIII gene from a panel of 83 patients with hemophilia A: (i) a 7-kilobase (kb) deletion that eliminates exon 6; (it) a 2.5-kb deletion that eliminates 5' sequences of exon 14; (ii) a deletion of at least 7 kb that eliminates exons 24 and 25; (iv) a deletion of at least 16 kb that eliiminates exons 23-25; and (v) a 5.5-kb deletion that eliminates exon 22. The first four deletions are associated with severe hemophilia A. By contrast, the last deletion is associated with moderate disease, possibly because of in-frame splicing from adjacent exons. None of those patients with partial gene deletions had circulating inhibitors to factor VIII. One deletion occurred de novo in a germ cell of the maternal grandmother, while a second deletion occurred in a germ cell of the maternal grandfather. These observations demonstrate that de novo deletions of X-linked genes can occur in either male or female gametes.