Isozymes of Human Phosphofructokinase in Blood Cells and Cultured Cell Lines: Molecular and Genetic Evidence for a Trigenic System

• Published in: Blood 1981-04, Vol.57 (4), p.724-732
• Main Author: Vora, Shobhana
• Format: Article
• Language: English
• Subjects: Alleles; Antibodies; Blood Cells - enzymology; Blood Platelets - enzymology; Cell Line; Cells, Cultured; Chemical Precipitation; Erythrocytes - enzymology; Humans; Isoenzymes - blood; Isoenzymes - genetics; Lymphocytes - enzymology; Neutrophils - enzymology; Phosphofructokinase-1 - blood; Phosphofructokinase-1 - genetics
• ISSN: 0006-4971; 1528-0020
• DOI: 10.1182/blood.V57.4.724.724

We have previously demonstrated that human red cell (RBC) phosphofructokinase (PFK) is under dual genetic control, since it is a mixture of five tetrameric isozymes composed of two distinct subunits, M (muscle-type) and L (liver-type), i.e., M4, M3L, M2L2, ML3, and L4. The residual RBC PFK in a patient with Tarui's disease (glycogenosis type VII) was shown to consist exclusively of L4 type isozyme, indicating homozygosity for the deficiency of M-type subunits. In this study, we have investigated various normal and M-deficient blood cells and cultured cell lines for their quantitative and qualitative PFK profiles, using monoclonal antibodies in addition to protein hybridization technique. Our results indicate that erythrocytes and platelets exhibit the least PFK activities as compared to various nucleated blood cells. However, the highest activities are exhibited by cultured fibroblasts and lymphoblastoid cell lines. The lymphoblastoid cell line from our M-deficient patient exhibits a relative PFK deficiency as compared to the normal cell lines. The platelet PFK consists of three species with subunit structures of P4, P3L, and P2L2, since dissociation and reassociation of platelet PFK yields a five-membered set. The “P” subunit is under separate genetic control, since it exhibits a unique chromatographic mobility, complete resistance to anti-M and anti-L monoclonal antibodies, and normal expression in Tarui's disease. Lymphocytes and polymorphonuclear leukocytes exhibit predominantly L4 isozyme with some hybrids consisting of all 3 subunits. Fibroblast PFK is resolved chromatographically into 10–12 species, 5 being identical to those of RBC and 3 to those of platelets, suggesting simultaneous expression of all 3 genes. The hybridization of RBC PFK (M + L) and platelet PFK (P + L) yields the same 10–12 fibroblastic species, proving coexpression of all 3 genes. Normal lymphoblastoid cell lines exhibit the same 10–12 species seen in fibroblasts, as do the RBC of a few otherwise healthy individuals. In accord with the above data, the lymphoblastoid cell line from the M-deficient patient exhibits only 5 species composed of P and L subunits. These studies demonstrate the multimolecular forms of PFK in various blood cells and cultured cell lines and provide evidence for the existence of 3 structural loci coding for human PFK, i.e., M, P, and L.