F8 and F9 mutations fail to co-segregate in a family with co-incident haemophilia A and B.

Haemophilia A and B in one individual may arise from co-incident inheritance of independent mutations in the F8 and F9 genes. However, this association is rare and has been studied poorly at a genetic level. We report a male patient with abnormal bleeding and reduced factor VIII:C (26 IU dL(-1)) and factor IX:C (35 IU dL(-1)). This index case harboured a F8 c.979C>G transversion (predictive of p.Leu327Val) and a F9 c.845A>G transition (predictive of p.His282Arg) which have been previously associated with mild haemophilia A and B, respectively. Identical F8 and F9 mutations were identified in the mother and maternal grandmother. However, an affected maternal uncle showed only the F8 c.979C>G mutation, indicating haemophilia A alone. The sister of the index case was heterozygous only for F9 c.845A>G, indicating carriership of haemophilia B alone. The non-Mendelian inheritance of F8 c.979C>G and F9 c.845A>G in this kindred is consistent with recombination between F8 and F9 and illustrates the large recombination distance between these loci. Recognition of this phenomenon was essential for accurate genetic counselling in this kindred.

Effects of colchicine, cytochalasin-B and papaverine on wound healing in Xenopus early embryos.

The effects of colchicine, cytochalasin-B and papaverine on wound healing in Xenopus early embryos have been studied. Colchicine does not prevent wound healing, whereas cytochalasin-B does. Papaverine, under conditions which prevent the completion of neurulation, does not prevent wound healing. A model is given which might explain these observations.

Haemopoietic stem cells: possibility of toxic effects of 5-fluorouracil on spleen colony formation.

Spleen colony formation (Till and McCulloch, 1961) by cells collected from mice treated with the cytotoxic drug 5-fluorouracil (5-FUBM) differs from spleen colony formation by normal bone marrow cells (NBM). The number of spleen colonies formed after the graft of 5-FUBM rises with the passage of time since the graft (extra spleen colonies). It does not after the graft of NBM. Hodgson and Bradley (1979) proposed that extra spleen colonies might form in this way: a progenitor of CFUs ('the pre-CFUs') seeded preferentially to the marrow of recipient mice. It formed CFUs there which migrated to the spleen and formed extra colonies. This hypothesis, if it were true, would support another hypothesis, the generation-age hypothesis (Rosendaal et al., 1976, 1979), which proposes that the number of divisions there have been in the history of a stem cell is one determinant of the order in which they start cycling to form other stem cells and differentiated cells. We investigated four possible sources of extra colonies. They might arise by reseeding of CFUs formed in the marrow or the spleen; they might arise from CFUs which settle in spleen but require a variable time until they can start to cycle, or they might arise after a variable time as CFUs repair the damage done them by 5-FU. We have established that the CFUs which eventually form spleen colonies are in the spleens of recipients 24 hours after the graft. Extra colonies too, do not arise by reseeding of CFUs formed in the spleens of recipients. It is possible that extra colonies arise as CFUs repair the damage done by 5-FU.

Human minisatellites, repeat DNA instability and meiotic recombination.

Minisatellites include some of the most variable loci in the human genome and are superb for dissecting processes of tandem repeat DNA instability. Single DNA molecule analysis has revealed different mutation processes operating in the soma and germline. Low-level somatic instability results in simple intra-allelic rearrangements. In contrast, high frequency germline instability involves complex gene conversions and is therefore recombinational in nature, almost certainly occurring at meiosis. To determine whether true meiotic crossovers occur at human minisatellites, we have used polymorphisms near the repeat array to recover recombinant DNA molecules directly from sperm DNA. Analysis of minisatellite MS32 has revealed an intense and highly localised meiotic crossover hotspot centred upstream of the array, the first example of a human hotspot defined at the molecular level. This hotspot extends into the beginning of the repeat array, resulting in unequal and equal crossovers. Array crossovers occur much less frequently than array conversions but appear to arise by a common process, most likely by alternative processing of a recombination initiation complex. The location of MS32 at the boundary of a recombination hotspot suggests that this locus has evolved as a by-product of localised meiotic recombination activity, and that minisatellites might in general mark recombinationally proficient hotspots or hot domains in the genome. Finally, sperm crossover analysis makes it possible to explore the molecular rules that govern human meiotic recombination, and to detect phenomena such as meiotic drive that could provide a possible connection between recombination and DNA sequence diversity itself.