TY - JOUR

T1 - Sterol mutants of Chlamydomonas reinhardtii

T2 - Characterisation of three strains deficient in C24(28) reductase

AU - Salimova, Ekaterina

AU - Boschetti, Arminio

AU - Eichenberger, Waldemar

AU - Lutova, Ludmila

N1 - Funding Information: The plasmid pARG7 was a kind gift from M. Goldschmidt-Clermont, University of Geneva. We are indebted to Dr T. Schneeberger for the GC-MS analyses. The work has been supported by the Swiss National Science Foundation (Grant 31-45901.95).

PY - 1999/4

Y1 - 1999/4

N2 - Three mutants of Chlamydomonas reinhardtii (strain arg7cw15) were obtained using the strategy of insertional mutagenesis by random plasmid integration with subsequent selection for resistance against the polyene antibiotic nystatin. Sterols were isolated by precipitation with digitonin, fractionated by both normal and argentation TLC, and then analysed by GLC and GC-MS. All the mutants accumulated ergosta-5,7,22,24(28)-tetraenol, ergosta-5,7,24(28)-trienol, ergosta-7,24(28)-dienol, stigmasta-5,7,22,24(28)-tetraenol, stigmasta-5,7,24(28)-trienol, stigmasta-8,24(28)-dienol and stigmasta-7,24(28)-dienol, while ergosterol and 7-dehydroporiferasterol which are the only major sterol components of the original strain were absent in the mutants. It is concluded that all these mutants are impaired in this C24(28) reductase which catalyses the reduction of the C24(28) tetraenol to the corresponding 24-alkyl sterol. There is strong evidence that the same enzyme acts on both the C28 and C29 sterol series. This view is also supported by Southern blot hybridisation analysis revealing that in all three mutants, plasmid insertion occurred at the same site indicating the disruption of the same gene. Due to the insertional nature of the mutations, the strains can be used for cloning the corresponding gene.

AB - Three mutants of Chlamydomonas reinhardtii (strain arg7cw15) were obtained using the strategy of insertional mutagenesis by random plasmid integration with subsequent selection for resistance against the polyene antibiotic nystatin. Sterols were isolated by precipitation with digitonin, fractionated by both normal and argentation TLC, and then analysed by GLC and GC-MS. All the mutants accumulated ergosta-5,7,22,24(28)-tetraenol, ergosta-5,7,24(28)-trienol, ergosta-7,24(28)-dienol, stigmasta-5,7,22,24(28)-tetraenol, stigmasta-5,7,24(28)-trienol, stigmasta-8,24(28)-dienol and stigmasta-7,24(28)-dienol, while ergosterol and 7-dehydroporiferasterol which are the only major sterol components of the original strain were absent in the mutants. It is concluded that all these mutants are impaired in this C24(28) reductase which catalyses the reduction of the C24(28) tetraenol to the corresponding 24-alkyl sterol. There is strong evidence that the same enzyme acts on both the C28 and C29 sterol series. This view is also supported by Southern blot hybridisation analysis revealing that in all three mutants, plasmid insertion occurred at the same site indicating the disruption of the same gene. Due to the insertional nature of the mutations, the strains can be used for cloning the corresponding gene.

KW - C24(28) reductase

KW - Chlamydomonas

KW - Mutants

KW - Nuclear transformation

KW - Sterols

UR - http://www.scopus.com/inward/record.url?scp=0033057044&partnerID=8YFLogxK

U2 - 10.1016/S0981-9428(99)80022-9

DO - 10.1016/S0981-9428(99)80022-9

M3 - Article

AN - SCOPUS:0033057044

VL - 37

SP - 241

EP - 249

JO - Plant Physiology and Biochemistry

JF - Plant Physiology and Biochemistry

SN - 0981-9428

IS - 4

ER -