A new PKLR gene mutation in the R-type promoter region affects the gene transcription causing pyruvate kinase deficiency

Mutations in the PKLR gene responsible for pyruvate kinase (PK)-deficient a naemia are mainly located in the coding regions: 11 are in the splicing sit es and, recently, three mutations have been described in the promoter regio n. We now report a novel point mutation A --> G on nucleotide 72, upstream from the initiation codon of the PKLR gene, in four Portuguese PK-deficient patients. This new regulatory mutation occurs within the most proximal of the four GATA motifs (GATA-A element) in the R-type promoter region. In two patients who were homozygous for this mutation, a semiquantitative reverse transcription polymerase chain reaction (PCR) procedure was used to evalua te the amount of R-PK mRNA transcript in the reticulocytes. The mRNA level was about five times lower than in normal controls, demonstrating that the PKLR gene transcription is severely affected, most probably because the -72 A --> G point mutation disables the binding of the erythroid transcription factor GATA-1 to the GATA-A element. Supporting these data, the two patient s homozygous for the -72A --> G mutation had severe haemolytic anaemia and were transfusion dependent until splenectomy. Two other patients who were c ompound heterozygous for this mutation and the previously described missens e mutation 1456C --> T had a mild condition.