Identification of the mutation affecting the KEL gene responsible for a K0 phenotype

MATTALONI S; ARMONI C; TRUCCO BOGGIONE C; LUJÁN BRAJOVICH M; GARCÍA BORRÁS S; BIONDI C; CASTILHO L; COTORRUELO C

Londres

Congreso; 25th Regional Congress of the International Society of Blood Transfusion; 2015

International Society of Blood Transfusion

Backgound: the Kell system is one of the most clinically relevant blood group. It comprises at least 34 antigens being KEL1/KEL2 (K/k), KEL3/KEL4 (Kpa/Kpb) and KEL6/KEL7 (Jsa/Jsb) antithetical and the most important epitopes. Most Kell antibodies are involved in the pathogenesis of hemolytic disease of the fetus and newborn and severe hemolytic transfusion reactions. Kell antigens are encoded by the KEL gene which comprises 19 exons. Single nucleotide polymorphisms (SNPs) are the most common cause of the different Kell phenotypes. Molecular defects may lead to the rare K0 (Knull) phenotype characterized by the absence of Kell antigens expression. Limited clinical data have been published regarding the significance of anti-Ku, seen in K0 individuals.Aims: the aims of this study were to characterize the molecular basis of a K0 phenotype encountered in a pregnant woman and report the clinical outcome the pregnancy.Materials and Methods: a peripheral blood sample of a pregnant patient was referred to our reference laboratory because of the presence of a panreactive antierythrocyte antibody. Standard serological analyses based on hemagglutination were performed using tube and gel techniques. Genomic DNA was obtained with a modified salting-out method. Kel genotyping was performed by PCR-RFLP strategies. All coding and intron-exon splice regions were analysed by Sanger sequencing.Results: serological studies showed an IgG 37ºC-reactive alloantibody, titer 128. Extended red blood cell phenotype analysis failed to detect the expression of neither KEL1, KEL2, KEL3, KEL4, KEL6 nor KEL7 antigens suggesting a K0 phenotype with anti-Ku. KEL genotyping showed KEL*02/02, KEL*04/04 and KEL*07/07 genotypes. Sequence analysis of the 19 exons and intron-exon splice regions of KEL showed a guanine to an adenine substitution at the first nucleotide of intron 3 (IVS3+1g>a) originating the allele KEL*02N.06.Conclusions: the replacement of the conserved sequence gt at the donor splice site by at is responsible for an alternative assembly of mRNA with loss of part of exon 3 sequences. Alternatively, this mutation causes skipping of the entire exon 3. In both cases, a downstream premature stop codon is generated in exon 4 and no Kel protein synthesized. Molecular finding confirmed the suspected K0 phenotype. To our knowledge, this is first description of this unusual phenotype in an Argentinean patient. She gave birth to a preterm infant with hemolytic disease of the newborn, who was successfully treated with phototherapy. Identification of the mutation responsible for this exceptional phenotype allowed the development of a PCR-SSP strategy to detect this allele in family members and provide appropriate genetic counseling regarding transfusion therapy and pregnancy.