Identification of a new WT1 mutation in a sporadic Wilms' tumour.

A new mutation in WT1 is described in a sporadic unilateral Wilms' tumour consisting of a 17 bp duplication in exon 7 generating a stop codon. The second allele is either partially deleted or presents the same alteration. LOH analysis at 11p15.5 and at the 16q13-16q24.3 regions indicated retention of heterozygosity in the tumour DNA for the markers analysed. The results are consistent with Knudson's hypothesis and confirm that loss of function of WT1 contributes to the development of at least some Wilms' tumours.

Dominantly transmitted beta-thalassemia arising from the production of several aberrant mRNA species and one abnormal peptide.

We describe a dominantly inherited beta-thalassemia intermedia phenotype observed in a five-generation Portuguese family. Carriers are characterized by moderate anemia, hypochromia, microcytosis, elevated hemoglobin (Hb)A2 and HbF levels, splenomegaly, hepatomegaly, and inclusion bodies in peripheral red blood cells after splenectomy. The molecular basis of this condition is a small deletion within the 5' consensus splicing sequence of the second intron of the beta-globin gene, IVS-II-4,5 (-AG). Reticulocyte RNA studies performed by reverse transcription-polymerase chain reaction (RT-PCR) and primer extension analysis showed three abnormally processed transcripts, which, upon sequencing, were shown to correspond to (1) skipping of exon 2, and (2) activation of two cryptic splice sites (between codons 59/60, and at IVS-II-47). In vitro translation studies of these patients' reticulocyte RNA have shown that at least one of these aberrant mRNA species is translated into an abnormally elongated peptide whose cytotoxic properties could, in part, be causing the atypical dominant mode of inheritance observed in this family. We suggest that this elongated beta chain is unable to combine with an alpha-globin chain to form a functional Hb molecule. Its degradation would, then, exhaust the proteolytic defense mechanism of the erythroid precursors, leading to inefficient proteolysis of the free alpha chains in excess.

A novel mosaic Bantu/Benin/Bantu beta s haplotype found in several African populations.

In a survey of the chromosomal backgrounds associated with the sickle cell gene in Portuguese-speaking populations from Europe and Africa, a discordance between the classical haplotype and the predicted allele at the RsaI polymorphism 5' to the beta globin gene was observed in four patients. Extensive typing of the corresponding beta s chromosomes at simple polymorphic repeat motifs revealed a novel "extended" haplotype that appeared to be a mosaic of (1) a Bantu-type DNase I hypersensitive site 2 within the beta globin gene cluster locus control region, (2) a Benin 5' subhaplotype, and (3) a Bantu 3' subhaplotype. We propose two alternative schedules for the generation of yet another chromosomal background of the sickle cell gene.

Insertional inactivation of the WT1 gene in tumour cells from a patient with WAGR syndrome.

The WT1 gene was analysed using DNA from a Wilms' tumour derived from a patient with the WAGR syndrome using single strand conformation polymorphism analysis and polymerase chain reaction sequencing. A 14-bp insertion was found in the intron part of the splice donor site of exon 7 and was a tandem duplication of an upstream exon sequence. This mutation would be expected to disrupt the correct processing of the WT1 mRNA and is predicted to result in a non-functional protein. This observation further supports the role of WT1 in Wilms' tumorigenesis in patients with constitutional 11p13 deletions.

Importation route of the sickle cell trait into Portugal: contribution of molecular epidemiology.

To elucidate the origin and spread of the sickle cell trait into the Portuguese population, we examined nine polymorphic DNA markers within the beta globin gene cluster defining the haplotype. The population sample included 64 sickle-cell-gene-bearing individuals from defined Portuguese-speaking white, black, and Asian Indian populations. The nature and geographic distribution of the different beta S haplotypes in Portugal suggest that the sickle cell trait has been imported twice: between the eighth and the thirteenth centuries from the Mediterranean basin (in association with the Benin haplotype) and after the fifteenth century from black Africa over an Atlantic route (Senegal and Bantu haplotypes).

Novel promoter and splice junction defects add to the genetic, clinical or geographic heterogeneity of beta-thalassaemia in the Portuguese population.

In order to delineate the spectrum and the relative abundance of beta-globin gene defects causing thalassaemia in the Portuguese population, a representative sample was analysed including 51 beta-thalassaemia carriers along with 26 patients representing different clinical phenotypes. Seven mutations were identified, four of which [codon 39 (C----T), 39%; intervening sequence (IVS) 1 nucleotide (nt) 1 (G----A), 26%; IVS 1 nt 110 (G----A), 17%; IVS 1 nt 6 (T----C), 15%] account for 97% of 93 beta-thalassaemia chromosomes. Two previously undescribed mutations, namely a C----T substitution at position--90 in the proximal CACCC box, and the deletion of nucleotides 4 and 5 (AG) in IVS2 were identified. The uncommon, though ubiquitous, G----T transversion at codon 121 was found once upon haplotype V. Direct prenatal diagnosis can be offered to 95% of couples at risk of bearing a thalassaemic child.

Alpha-thalassemia resulting from deletion of regulatory sequences far upstream of the alpha-globin structural genes.

We describe an alpha-thalassemia determinant in which alpha-globin expression is silenced by a deletion located 27 kb 5' to the transcription start site of the alpha 2-globin gene. This alpha-thalassemic determinant, (alpha alpha)MM, is a member of a newly described group of thalassemic mutations resulting from deletion of locus-controlling sequences critical to globin gene expression.