Related Experiment Videos
Point mutations causing the McLeod phenotype
David C W Russo1, Soohee Lee, Marion E Reid
1Lindsley F. Kimball Research Institute, The New York Blood Center, New York, New York 10021, USA. drusso@nybc.org
Background:
The McLeod phenotype is defined by absence of Kx, weakening of Kell system antigens, and acanthocytosis. Individuals with the McLeod phenotype usually develop late-onset neuromuscular abnormalities. Gene deletions, insertions, and point mutations that affect RNA splicing or that lead to premature stop codons have been reported to cause the McLeod phenotype. The McLeod phenotype may also be caused by mutations at a different splice site and by a novel mutation encoding an amino acid substitution that prevents transport to the cell surface.
Study Design And Methods:
The coding and flanking intron regions of XK from four male, unrelated individuals with the McLeod phenotype and non-chronic granulomatous disease were sequenced and compared with the wild type sequence. Genomic DNA was amplified by PCR, and the products were sequenced. In one case, the mutant cDNA was expressed in a heterologous cell, and cell surface expression was determined.
Results:
Three individuals with the McLeod phenotype had mutations that disrupted conserved GT sequences present at RNA splice sites. Two of them had G>C mutations at the 5' splice site of intron 1, and one had a G>A mutation at the 5' splice site of intron 2. One person with the McLeod phenotype had a 746C>G mutation in exon 3 encoding an R222G substitution. In a transfected cell, the expressed protein from the latter mutant did not travel to the cell surface.
Conclusion:
The McLeod phenotype may be caused by several different mutations.
Insights
Multiple mutations in the XK gene can cause the McLeod phenotype, affecting Kx antigen expression and leading to neuromuscular issues. These genetic alterations disrupt RNA splicing or protein transport to the cell surface.
Area of Science:
- Genetics
- Molecular Biology
- Hematology
Background:
- The McLeod phenotype is characterized by absent Kx, weakened Kell antigens, and acanthocytosis, often leading to late-onset neuromuscular problems.
- Previously identified causes of the McLeod phenotype include gene deletions, insertions, and point mutations affecting RNA splicing or causing premature stop codons.
Purpose of the Study:
- To investigate the genetic basis of the McLeod phenotype in four unrelated males.
- To identify novel mutations in the XK gene responsible for the McLeod phenotype.
Main Methods:
- Sequencing of the coding and flanking intron regions of the XK gene in four individuals with the McLeod phenotype.
- PCR amplification of genomic DNA followed by sequencing.
- Expression of mutant cDNA in a heterologous cell system to determine cell surface protein expression.
Main Results:
- Three individuals exhibited mutations disrupting conserved GT sequences at RNA splice sites (two with G>C at intron 1 5' splice site, one with G>A at intron 2 5' splice site).
- One individual had a 746C>G mutation in exon 3, resulting in an R222G amino acid substitution.
- The R222G mutant protein failed to be transported to the cell surface in transfected cells.
Conclusions:
- The McLeod phenotype can result from various mutations within the XK gene.
- Mutations can affect RNA splicing or protein trafficking, leading to the McLeod phenotype.