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. 1980 Jan;32(1):16–25.

Kinetic analysis of genetic complementation in heterokaryons of propionyl CoA carboxylase-deficient human fibroblasts.

B Wolf, H F Willard, L E Rosenberg
PMCID: PMC1685923  PMID: 7361761

Abstract

We studied genetic complementation of propionyl CoA carboxylase (PCC) deficiency in cultures of polyethylene glycol (PEG)-induced heterokaryons, using mutant fibroblast lines assigned to five mutant classes, designated bio, pcc A, pcc B, pcc C, and pcc BC. By measuring PCC activity directly in extracts of fused cells or indirectly in intact cells by [1-14C]propionate utilization, we confirmed the nonlinear nature of the PCC deficiency complementation map described by Gravel et al. [1]. When we studied the kinetics of complementation, we detected three distinct patterns using the [1-14C]propionate utilization assay. When either pcc A or pcc C lines were fused to bio cells, 14C-fixation increased to half of the maximally restored values within 4 hrs. In pcc A x pcc C crosses or in pcc A x pcc B crosses, however, complementation was much slower. In fusions between pcc B and pcc C cells, a third pattern was elicited; complementation was incomplete, maximum restoration of PCC activity begin less than 20% of that observed in other complementing crosses. From these data and previous biochemical evidence, we suggest (1) that the bio and pcc mutations affect different genes; (2) that complementation between pcc A and either pcc B, pcc C, or pcc BC lines is intergenic and involves subunit exchange and synthesis of new PCC molecules; and (3) that complementation between pcc B and pcc C mutants is interallelic.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Bartlett K., Gompertz D. Combined carboxylase defect: biotin-responsiveness in cultured fibroblasts. Lancet. 1976 Oct 9;2(7989):804–804. doi: 10.1016/s0140-6736(76)90640-1. [DOI] [PubMed] [Google Scholar]
  2. CRICK F. H., ORGEL L. E. THE THEORY OF INTER-ALLELIC COMPLEMENTATION. J Mol Biol. 1964 Jan;8:161–165. doi: 10.1016/s0022-2836(64)80156-x. [DOI] [PubMed] [Google Scholar]
  3. Case M. E., Giles N. H. COMPARATIVE COMPLEMENTATION AND GENETIC MAPS OF THE PAN-2 LOCUS IN NEUROSPORA CRASSA. Proc Natl Acad Sci U S A. 1960 May;46(5):659–676. doi: 10.1073/pnas.46.5.659. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Chasin L. A., Urlaub G. Mutant alleles for hypoxanthine phosphoriboxyltransferase: codominant expression, complementation, and segregation in hybrid Chinese hamster cells. Somatic Cell Genet. 1976 Sep;2(5):453–467. doi: 10.1007/BF01542725. [DOI] [PubMed] [Google Scholar]
  5. Fratantoni J. C., Hall C. W., Neufeld E. F. Hurler and Hunter syndromes: mutual correction of the defect in cultured fibroblasts. Science. 1968 Nov 1;162(3853):570–572. doi: 10.1126/science.162.3853.570. [DOI] [PubMed] [Google Scholar]
  6. Galjaard H., Hoogeveen A., de Wit-Verbeek H. A., Reuser A. J., Keijzer W., Westerveld A., Bootsma D. Tay-Sachs and Sandhoff's disease: intergenic complementation after somatic cell hybridization. Exp Cell Res. 1974 Aug;87(2):444–448. doi: 10.1016/0014-4827(74)90515-1. [DOI] [PubMed] [Google Scholar]
  7. Gravel R. A., Lam K. F., Scully K. J., Hsia Y. Genetic complementation of propionyl-CoA carboxylase deficiency in cultured human fibroblasts. Am J Hum Genet. 1977 Jul;29(4):378–388. [PMC free article] [PubMed] [Google Scholar]
  8. Gravel R. A., Mahoney M. J., Ruddle F. H., Rosenberg L. E. Genetic complementation in heterokaryons of human fibroblasts defective in cobalamin metabolism. Proc Natl Acad Sci U S A. 1975 Aug;72(8):3181–3185. doi: 10.1073/pnas.72.8.3181. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Hsia Y. E., Scully K. J., Rosenberg L. E. Human propionyl CoA carboxylase: some properties of the partially purified enzyme in fibroblasts from controls and patients with propionic acidemia. Pediatr Res. 1979 Jun;13(6):746–751. doi: 10.1203/00006450-197906000-00005. [DOI] [PubMed] [Google Scholar]
  10. LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
  11. Maul G. G., Steplewski Z., Weibel J., Koprowski H. Time sequence and morphological evaluations of cells fused by polyethylene glycol 6000(1). In Vitro. 1976 Nov;12(11):787–796. doi: 10.1007/BF02835453. [DOI] [PubMed] [Google Scholar]
  12. Nadler H. L., Chacko C. M., Rachmeler M. Interallelic complementation in hybrid cells derived from human diploid strains deficient in galactose-1-phosphate uridyl transferase activity. Proc Natl Acad Sci U S A. 1970 Oct;67(2):976–982. doi: 10.1073/pnas.67.2.976. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Sekiguchi T., Sekiguchi F., Tomii S. Genetic complementation in hybrid cells derived from mutagen-induced mouse clones deficient in HGPRT activity. Exp Cell Res. 1975 Jun;93(1):207–218. doi: 10.1016/0014-4827(75)90441-3. [DOI] [PubMed] [Google Scholar]
  14. Thomas G. H., Taylor H. A., Miller C. S., Axelman J., Migeon B. R. Genetic complementation after fusion of Tay-Sachs and Sandhoff cells. Nature. 1974 Aug 16;250(467):580–582. doi: 10.1038/250580a0. [DOI] [PubMed] [Google Scholar]
  15. Weyler W., Sweetman L., Maggio D. C., Nyhan W. L. Deficiency of propionyl-Co A carboxylase and methylcrotonyl-Co A carboxylase in a patient with methylcrotonylglycinuria. Clin Chim Acta. 1977 May 2;76(3):321–328. doi: 10.1016/0009-8981(77)90158-9. [DOI] [PubMed] [Google Scholar]
  16. Willard H. F., Ambani L. M., Hart A. C., Mahoney M. J., Rosenberg L. E. Rapid prenatal and postnatal detection of inborn errors of propionate, methylmalonate, and cobalamin metabolism: a sensitive assay using cultured cells. Hum Genet. 1976 Dec 15;34(3):277–283. doi: 10.1007/BF00295291. [DOI] [PubMed] [Google Scholar]
  17. Willard H. F., Mellman I. S., Rosenberg L. E. Genetic complementation among inherited deficiencies of methylmalonyl-CoA mutase activity: evidence for a new class of human cobalamin mutant. Am J Hum Genet. 1978 Jan;30(1):1–13. [PMC free article] [PubMed] [Google Scholar]
  18. Wolf B., Hsia Y. E., Rosenberg L. E. Biochemical differences between mutant propionyl-CoA carboxylases from two complementation groups. Am J Hum Genet. 1978 Sep;30(5):455–464. [PMC free article] [PubMed] [Google Scholar]

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