The argininosuccinate lyase gene of Chlamydomonas reinhardtii: an important tool for nuclear transformation and for correlating the genetic and molecular maps of the ARG7 locus.

Debuchy, Robert; Purton, Saul; Rochaix, Jean‐David

doi:10.1002/j.1460-2075.1989.tb08426.x

Cited by 382 publications

(268 citation statements)

References 32 publications

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“…This pASL plasmid bears the Chlamydomonas ARG7 gene that codes for the argininosuccinate lyase (Debuchy et al, 1989) and is used as a selectable marker. Prototroph transformants were selected on TAP agar plates.…”

Section: Rna Interferencementioning

confidence: 99%

Inactivation of genes coding for mitochondrial Nd7 and Nd9 complex I subunits in Chlamydomonas reinhardtii. Impact of complex I loss on respiration and energetic metabolism

et al. 2014

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In Chlamydomonas, unlike in flowering plants, genes coding for Nd7 (NAD7/49 kDa) and Nd9 (NAD9/30 kDa) core subunits of mitochondrial respiratory-chain complex I are nucleus-encoded.Both genes possess all the features that facilitate their expression and proper import of the polypeptides in mitochondria. By inactivating their expression by RNA interference or insertional mutagenesis, we show that both subunits are required for complex I assembly and activity.Inactivation of complex I impairs the cell growth rate, reduces the respiratory rate, leads to lower intracellular ROS production and lower expression of ROS scavenging enzymes, and is associated to a diminished capacity to concentrate CO 2 without compromising photosynthetic capacity.

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Section: Rna Interferencementioning

confidence: 99%

Inactivation of genes coding for mitochondrial Nd7 and Nd9 complex I subunits in Chlamydomonas reinhardtii. Impact of complex I loss on respiration and energetic metabolism

et al. 2014

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“…Motility mutants were identified by growing positive transformants in liquid culture and screening by light microscopy. Cotransformations were performed using ODA5 genomic constructs and pARG7.8 plasmid (Debuchy et al, 1989); transformants were selected on TAP plates.…”

Section: Transformations and Insertional Mutagenesismentioning

confidence: 99%

“…The BAC clones were tested for their ability to rescue the Oda5Ϫ phenotype by cotransforming strain 112b with the BAC clones and plasmid pARG7.8 (Debuchy et al, 1989). Transformants were scored for Odaϩ/Ϫ phenotype by light microscopy.…”

Section: Cloning and Sequencing Of The Oda5 Gene And Cdnamentioning

confidence: 99%

Oda5p, a Novel Axonemal Protein Required for Assembly of the Outer Dynein Arm and an Associated Adenylate Kinase

Wirschell

Pazour

Yoda

et al. 2004

MBoC

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Of the uncloned ODA genes required for outer dynein arm assembly in Chlamydomonas, ODA5 and ODA10 are of particular interest because they do not encode known subunits of the outer arm or the outer dynein arm-docking complex (ODA-DC), and because genetic studies suggest their products interact. Beginning with a tagged oda5 allele, we isolated genomic and cDNA clones of the wild-type gene. ODA5 predicts a novel, 66-kDa coiled-coil protein. Immunoblotting indicates Oda5p is an axonemal component that assembles onto the axoneme independently of the outer arm and ODA-DC and is uniquely missing in oda5 and oda10 axonemes. Oda5p is released from the axoneme by extraction with 0.6 M KCl, but the soluble Oda5p does not cosediment with the outer dynein arm/ODA-DC in sucrose gradients. Quantitative mass spectrometry by using isotope coded affinity tagging revealed that a previously unidentified adenylate kinase is reduced 35-50% in oda5 flagella. Direct enzymatic assays demonstrated a comparable reduction in adenylate kinase activity in oda5 flagella, and also in oda10 flagella, but not in flagella of other oda mutants. We propose that Oda5p is part of a novel axonemal complex that is required for outer arm assembly and anchors adenylate kinase in proximity to the arm.

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“…To determine whether any of the recovered phage clones or PF6 gene constructs could rescue the motility defects in the pf6 mutants, pf6 arg7 strains were cotransformed with pARG7.8 (containing a wild-type copy of the argininosuccinate lyase gene; Debuchy et al, 1989) and the PF6 clone being tested. After growth for 7-10 d on selective media, the motility phenotypes of the argϩ transformants were scored using a dissecting microscope.…”

Section: Transformation and Screening For Rescue Of Motility Defectmentioning

confidence: 99%

TheChlamydomonas PF6Locus Encodes a Large Alanine/Proline-Rich Polypeptide That Is Required for Assembly of a Central Pair Projection and Regulates Flagellar Motility

Rupp

O’Toole

Porter

2001

MBoC

113

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Efficient motility of the eukaryotic flagellum requires precise temporal and spatial control of its constituent dynein motors. The central pair and its associated structures have been implicated as important members of a signal transduction cascade that ultimately regulates dynein arm activity. To identify central pair components involved in this process, we characterized a Chlamydomonas motility mutant (pf6-2) obtained by insertional mutagenesis. pf6-2 flagella twitch ineffectively and lack the 1a projection on the C1 microtubule of the central pair. Transformation with constructs containing a full-length, wild-type copy of the PF6 gene rescues the functional, structural, and biochemical defects associated with the pf6 mutation. Sequence analysis indicates that the PF6 gene encodes a large polypeptide that contains numerous alanine-rich, proline-rich, and basic domains and has limited homology to an expressed sequence tag derived from a human testis cDNA library. Biochemical analysis of an epitope-tagged PF6 construct demonstrates that the PF6 polypeptide is an axonemal component that cosediments at 12.6S with several other polypeptides. The PF6 protein appears to be an essential component required for assembly of some of these polypeptides into the C1-1a projection. INTRODUCTIONCilia and flagella are highly conserved structures found on diverse cell types, ranging from single-cell protozoa to multicellular tissues in humans, where they function to propel cells through a fluid environment or to transport fluid across a cell surface. Motile cilia are also found in the embryonic node (Sulik et al., 1994; Bellomo et al., 1996), where their motility appears to be critical for establishing the morphogenetic gradient that determines the left-right body axis in mammals (Nonaka et al., 1998). Defects in the assembly or activity of cilia and flagella result in a variety of abnormalities, including defects in left-right axis asymmetry, infertility, and respiratory disease (Afzelius et al., 1975; Afzelius, 1995;Supp et al., 1999). Most motile cilia and flagella contain an axoneme that consists of nine outer doublet microtubules surrounding two central singlet microtubules. The outer doublets contain binding sites for the inner and outer dynein arms, the molecular motors that power axoneme motility (reviewed by Porter, 1996;King, 2000). The dynein arms on one outer doublet interact transiently with the adjacent doublet to generate the force for interdoublet microtubule sliding. Other structures within the axoneme constrain and coordinate the activity of the multiple dynein motors and thereby convert microtubule sliding into flagellar bending.Both structural and genetic evidence indicate that the central pair microtubules and radial spokes play an important role in coordinating dynein activity. The two central pair microtubules are structurally asymmetric, and in several organisms, this apparatus has been shown to undergo clockwise rotation at a rate of approximately one turn per beat (reviewed by Omoto et al., 1999). Thes...

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The argininosuccinate lyase gene of Chlamydomonas reinhardtii: an important tool for nuclear transformation and for correlating the genetic and molecular maps of the ARG7 locus.

Cited by 382 publications

References 32 publications

Inactivation of genes coding for mitochondrial Nd7 and Nd9 complex I subunits in Chlamydomonas reinhardtii. Impact of complex I loss on respiration and energetic metabolism

Inactivation of genes coding for mitochondrial Nd7 and Nd9 complex I subunits in Chlamydomonas reinhardtii. Impact of complex I loss on respiration and energetic metabolism

Oda5p, a Novel Axonemal Protein Required for Assembly of the Outer Dynein Arm and an Associated Adenylate Kinase

TheChlamydomonas PF6Locus Encodes a Large Alanine/Proline-Rich Polypeptide That Is Required for Assembly of a Central Pair Projection and Regulates Flagellar Motility

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