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A transfection model for weak rhesus D expression
Author(s) -
Hemker M. B.,
Berger L.,
Van Der Schoot C. E.,
Van Rhenen D. J.,
Maaskantvan wijk P. A.
Publication year - 2000
Publication title -
transfusion medicine
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.471
H-Index - 59
eISSN - 1365-3148
pISSN - 0958-7578
DOI - 10.1046/j.1365-3148.2000.00261-30.x
Subject(s) - messenger rna , microbiology and biotechnology , transfection , biology , complementary dna , k562 cells , gene , genotype , phenotype , genetics
Weak D is recently reported to be encoded by 14 different weak D genotypes. The amount of RHD mRNA in donors expressing weak D was measured in previous studies. By semiquantitative RT‐PCR analysis Rouillac et al . ( Blood 1996) found lower RHD mRNA levels in donors expressing weak D compared to normal RhD donors, whereas Beckers et al . ( Transfusion 1997) described equal mRNA levels. The aim of the present study is to explore whether the pointmutations found in donors carrying the weak D phenotype can be responsible for weak D expression. Real‐time quantitative PCR analysis showed that levels of RHD mRNA from donors expressing different weak D genotypes (type 1, 2 and 3) did not differ from RHD mRNA levels from normal heterozygous RhD donors. This indicates that the mutations found in weak D genotypes do not affect RhD expression on transcription level. To investigate whether the described mutations influence translation of mRNA into RhD protein or the configuration of the RhD protein, weak D type 1 cDNA (T809G) and weak D type 3 cDNA (C8G) was transfected in K562 cells. Preliminary results indicate that the ratios between RhD expression and RHD mRNA level (RhD expression/ RHD mRNA level) do not differ between weak D type 1 and normal D K562 cells, but are reduced in weak D type 3 transfectants compared to normal D K562 cells. This suggests that in K562 neither the translation nor the configuration of RhD is influenced by the mutation T809G. There might be a missing or limited factor in the K562 model involved in the expression of RhD, hampering the interpretation of the weak D type 1. However, the lower expression of RhD with the mutation C8G might be caused by changed configuration or changed intracellular transport to the membrane.