Assignment of the human gene for liver-type 6-phosphofructokinase isozyme (PFKL) to chromosome 21 by using somatic cell hybrids and monoclonal anti-L antibody.

Abstract

Human 6-phosphofructokinase (PFK; ATP:D-fructose-6-phosphate 1-phosphotransferase, EC 2.7.1.11) is under the control of structural loci that code for muscle (M), liver (L) and platelet (P) subunits, which are variably expressed in different tissues; human diploid fibroblasts and leukocytes express all 3 genes. Random tetramerization of these subunits produces various isozymes, which can be distinguished from one another by ion exchange chromatography or by subunit-specific monoclonal antibodies. Seventeen somatic cell hybrids established between Chinese hamster cells and human diploid fibroblasts or leukocytes were examined for the expression of L-type subunits of human PFK. Electrophoresis does not distinguish between Chinese hamster PFK and human PFK, so an anti-human L-subunit-specific monoclonal antibody, which does not react with Chinese hamster PFK was used. The expression of human L subunits in the hybrids was detected by the enzyme-immunoprecipitation technique using staphylococci bearing protein A as an immunoadsorbent. Out of 17 hybrids 12 expressed human L subunits and retained chromosome 21, as determined by chromosome and isozyme marker analysis, whereas 5 did not express human PFKL and lacked chromosome 21. The mean erythrocyte PFK of 7 individuals with trisomy 21 was elevated (147% of normal). A specific increase in L subunits in trisomic erythrocytes was evident chromatographically by a striking increase in L4 species (50%; normal 10%) and immunologically by decreased precipitation with anti-M monoclonal antibody (50%; normal 80%). PFKL is located on chromosome 21; the previously noted elevation of erythrocyte PFK activity in individuals with trisomy 21 is due to a gene-dosage effect.