Rh(D) antigen expression and isolation of a new Rh(D) cDNA isoform in human erythroleukemic K562 cells

Kimita Suyama, Ruth Lunn, Stephanie Haller, Jack Goldstein

Research output: Contribution to journalArticle

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Abstract

Human erythroleukemic K562 cells are known to have several erythroid properties. K562 cells possess Rh mRNAs, but expression of Rh proteins has not previously been reported. We immunoprecipitated Rh protein from K562 cell lysate using rabbit anti-Rh and detected Rh(D) antigens on K562 cells using fluorescence-activated cell sorting (FACS). These results suggest that K562 cells will be useful as an expression model for most Rh antigens. We also cloned a new Rh(D) cDNA isoform (RhK562-II), from a K562 cDNA library using polymerase chain reaction (PCR) with 5' and 3' end oligonucleotides of the published Rh(e/E) antigen encoding cDNA sequence as primers. Sequence analysis showed that RhK562-II is composed of 951 nucleotides (316 amino acids), identical to the first 939 nucleotides (exons 1 to 6) of one of the Rh(D) cDNAs (RhXIII), except for nucleotide 654 (C → G exchange). However, this exchange is the same as that of another published Rh(D) cDNA (RhPII cDNA). RhK562-II is deprived of exons 7 and 8 (nucleotides 940 to 1,153), followed by an identical sequence up to the 3' end of the open-reading frame of the RhXIII cDNA, which causes a frameshift mutation and produces a premature stop codon. In vitro expression of RhK562-II using the transcription and translation rabbit reticulocyte lysate system produced two major Rh-related proteins (30 kD and 25 kD), which were immunoprecipitated by rabbit polyclonal anti-Rh and separated on sodium dodecyl sulfate- polyacrylamide gel electrophoresis (SDS-PAGE).

Original languageEnglish (US)
Pages (from-to)1975-1981
Number of pages7
JournalBlood
Volume84
Issue number6
StatePublished - Sep 15 1994
Externally publishedYes

Fingerprint

K562 Cells
Protein Isoforms
Complementary DNA
Antigens
Nucleotides
Rabbits
Exons
Frameshift Mutation
Proteins
Hepatitis B e Antigens
Nonsense Codon
Reticulocytes
Polymerase chain reaction
Transcription
Electrophoresis
Gene Library
Sorting
Oligonucleotides
Sodium Dodecyl Sulfate
Open Reading Frames

ASJC Scopus subject areas

  • Hematology

Cite this

Rh(D) antigen expression and isolation of a new Rh(D) cDNA isoform in human erythroleukemic K562 cells. / Suyama, Kimita; Lunn, Ruth; Haller, Stephanie; Goldstein, Jack.

In: Blood, Vol. 84, No. 6, 15.09.1994, p. 1975-1981.

Research output: Contribution to journalArticle

Suyama, K, Lunn, R, Haller, S & Goldstein, J 1994, 'Rh(D) antigen expression and isolation of a new Rh(D) cDNA isoform in human erythroleukemic K562 cells', Blood, vol. 84, no. 6, pp. 1975-1981.
Suyama, Kimita ; Lunn, Ruth ; Haller, Stephanie ; Goldstein, Jack. / Rh(D) antigen expression and isolation of a new Rh(D) cDNA isoform in human erythroleukemic K562 cells. In: Blood. 1994 ; Vol. 84, No. 6. pp. 1975-1981.
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