Summary
In vivo complementation between different wild and mutant strains defective for nitrate assimilation has been performed by isolating diploid strains from the appropriate crosses. Twenty-two diploids homozygous or heterozygous with respect to nitrate reduction and able to grow on nitrate medium were obtained and their diploid character demonstrated from analyses of mating type, cell volume, nuclear size and progeny of crosses with haploid wild-type. All diploids were assayed for overall- and terminal-nitrate reductase (NR) activity and for the occurrence of the NR-diaphorase subunit. Data on NR activities in heterozygotes carrying mutation(s) in structural gene(s) (nit-1 or nit-1a, nit-1b) agree with the heteromultimeric nature of the enzyme complex previously described (Franco et al. (1984) EMBO J 3: 1403–1407), and indicate that subunits are exchangeable to form hybrid enzymes. In addition, in vitro complementation tests with mutant nit-1 of C. reinhardtii indicate that this mutant has defective NR-diaphorase subunits but intact terminal subunits. Super-repression caused by the mutant allele nit-2 is suppressed by the wild allele in heterozygotes, which suggests a positive control by the nit-2 product on structural gene(s) transcription. Mutant alleles of genes for the biosynthesis of molybdenum-containing cofactor, either nit-4 or nit-5 and nit-6, were recessive in diploids carrying them. The mutant allele of nit-3, from strain 307, was codominant in all heterozygotes suggesting that nit-3 codes for a protein whose activity is limiting for the molybdenum-cofactor biosynthetic pathway.
Similar content being viewed by others
Abbreviations
- NR:
-
nitrate reductase
- MNNG:
-
N-methyl-N+-Nitro-N-nitrosoguanidine
- MoCo:
-
molybdenum-containing cofactor
- WT:
-
wild-type
- DAPI:
-
4′,6-diamidino-2-phenylindol
References
Arnon DI (1949) Plant Physiol 24:1–15
Barea JL, Cárdenas J (1975) Arch Microbiol 105:21–25
Bradford MM (1976) Anal Biochem 72:248–256
Cove DJ (1979) Biol Rev 54:291–327
Crick FHC, Orgel LE (1964) J Mol Biol 8:161–165
Ebersold WT (1967) Science 157:447–449
Fernández E, Cárdenas J (1981a) Biochim Biophys Acta 657:1–12
Fernández E, Cárdenas J (1981b) Planta 153:254–257
Fernández E, Cárdenas J (1982a) Biochim Biophys Acta 681:530–537
Fernández E, Cárdenas J (1982b) Mol Gen Genet 186:164–169
Fernández E, Cárdenas J (1983a) Biochim Biophys Acta 745:12–19
Fernández E, Cárdenas J (1983b) Z Naturforsch 38:439–445
Fernández E, Matagne RF (1984) Curr Genet 8:635–640
Franco AR, Cárdenas J, Fernández E (1984a) EMBO J 3:1403–1407
Franco AR, Cárdenas J, Fernández E (1984b) FEBS Lett 176:453–456
Gilham NW (1965) Genetics 52:529–537
Gorman DS, Levine RP (1965) Proc Natl Acad Sci USA 54:1665–1669
Guerrero MG, Vega JM, Losada M (1981) Ann Rev Plant Physiol 32:169–204
Hewitt EJ, Notton BA (1980) Nitrate reductase systems in eukaryotic and prokaryotic organisms. In: Coughlan MP (ed) Molybdenum and molybdenum-containing enzymes. Pergamon Press, Oxford, pp 273–325
Levine RP, Ebersold WT (1960) Annu Rev Microbiol 14:197–216
Loppes R (1966) Z Vererbungsl 98:193–202
Loppes R (1970) Experientia 26:660–661
Loppes R, Matagne RF (1972) Genetica 43:422–430
Loppes R, Matagne RF (1975) Principles of genetic regulation in lower and higher plants. In: Ledoux L (ed) Genetic manipulations with plant material. Plenum Press, New York, pp 89–105 (Nato Advanced Study Institutes Ser)
Loppes R, Matagne RF, Strijkert PJ (1972) Heredity 28:239–251
Matagne RF (1976) Mol Gen Genet 146:209–214
Matagne RF, Hermesse MP (1981) Genetics 99:371–381
Matagne RF, Loppes R (1975) Interallelic complementation in the study of gene action. In: Ledoux L (ed) Genetic manipulations with plant material. Plenum Press, New York, pp 77–88 (Nato Advanced Study Institutes Ser)
Marzluf GA (1981) Microbiol Rev 45:437–461
Mendel RR, Müller AJ (1979) Mol Gen Genet 177:145–153
Nichols GL, Syrett PJ (1978) J Gen Microbiol 108:71–77
Nichols GL, Shehata SAM, Syrett PJ (1978) J Gen Microbiol 108:79–88
Pineda M, Fernández E, Cárdenas J (1984) Physiol Plant 62:453–457
Sosa FM, Ortega T, Barea JL (1978) Plant Sci Lett 11:51–58
Thacker A, Syrett PJ (1972) New Phytol 71:435–441
Zabin I, Villarejo MR (1975) Annu Rev Biochem 44:295–313
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Fernández, E., Matagne, R.F. In vivo complementation analysis of nitrate reductase-deficient mutants in Chlamydomonas reinhardtii . Curr Genet 10, 397–403 (1986). https://doi.org/10.1007/BF00418413
Received:
Issue Date:
DOI: https://doi.org/10.1007/BF00418413