Male-sterile Cytoplasm Of Maize Research Articles

Translation in a wheat germ cell-free system of RNA from mitochondria of the normal and Texas male-sterile cytoplasms of maize (Zea mays L.).

RNA isolated from etiolated seedling shoot mitochondria of maize (Zea mays L.) with normal (N) or Texas male-sterile (T) cytoplasm stimulated the incorporation of [35S]-methionine into protein when added to a cell-free protein-synthesizing system from wheat germ. Discrete polypeptides with molecular masses of up to approximately 67 kDa were synthesized, and the pattern of bands was distinct from that obtained with total RNA. Products of translation of T-urf13 RNA were identified by immunoprecipitation, and of atpA, coxI, and coxII RNA by hybrid arrest of translation by the cloned gene. Several polypeptides were differentially synthesized from N and T mitochondrial RNA; these differences were more extensive than those found when isolated, intact, N and T mitochondria are allowed to synthesize proteins.

Chimeric mitochondrial genes expressed in the C male-sterile cytoplasm of maize.

Aberrant recombinations involving the mitochondrial atp9, atp6 and coxII genes have created unique chimeric sequences in the C male-sterile cytoplasm (cms-C) of maize. An apparent consequence of the rearrangements is the interchanging of transcriptional and/or translational regulatory signals for these genes, and alterations in the reading frames encoding the atp6 and coxII genes in the C cytoplasm. Particularly unusual is the organization of the atp6 gene in cms-C mitochondria, designated atp6-C. The atp6-C sequence is a triple gene fusion product comprised of DNAs derived from atp9, atp6 and an open reading frame of unknown origin. Although there is no direct evidence indicating that these chimeric genes are responsible for the cytoplasmic male sterility (cms) trait, their novel arrangements and the strong correlation between these genes and the C type of male sterility suggest such a role.

A mitochondrial gene is lost via homologous recombination during reversion of CMS T maize to fertility

The Texas (T) male sterile cytoplasm of maize is distinguished by a mitochondrially synthesized 13-kd polypeptide and a high susceptibility to the toxin produced by the fungal pathogen Helminthosporium maydis. Fertile, toxin-resistant revertants show an altered restriction profile for mitochondrial DNA and do not produce the 13-kd polypeptide. Characterization of cosmid clones from CMS T maize and a revertant shows that a heavily transcribed open reading frame named T-URF13, potentially coding a 13-kd product, is deleted in the revertant mitochondria. Six transcripts present in CMS T mitochondria, 4000, 3000, 2000, 1800, 1500 and 1200 nucleotides in length, are lacking in revertant mitochondria. T-URF25, an open reading frame coding for a 25-kd product, lies to the 3' end of T-URF13 but is retained in the revertants. T-URF13 and T-URF25 are co-transcribed in CMS T mitochondria; in the revertant T-URF25 is present on a 3100-nucleotide species. The recombination that caused these changes involved a 127-bp repeated sequence. Homologous recombination took place within the central 55 bp of this imperfect repeat. Hybridization analysis of DNA and RNA from other revertants demonstrates that a similar or identical event has taken place independently in these revertants.

Stoichiometric differences in DNA molecules containing the atpA gene suggest mechanisms for the generation of mitochondrial genome diversity in maize

Four genomic arrangements of the maize mitochondrial atpA gene (encoding the alpha subunit of the F(1) ATPase), have been characterized. Most N (fertile) and S (male-sterile) cytoplasms contain two atpA arrangements of equal abundance. Prolonged exposure of blots of maize mitochondrial DNA probed with atpA-specific sequences show that cytoplasms previously reported to lack one of the atpA arrangements do contain the second arrangement but at low levels. Similarly, restriction fragments containing the atpA gene previously thought unique to male-sterile S and T cytoplasms are present in low abundance in fertile cytoplasms. These observations suggest that fertile and male-sterile cytoplasms of maize may be more closely related than previously thought, and suggest possible mechanisms to explain the observed mitochondrial genome diversity.

The maize cytochrome c oxidase subunit I gene: sequence, expression and rearrangement in cytoplasmic male sterile plants

The single copy of the gene for cytochrome c oxidase subunit I (COX I) present in the mitochondrial genome of fertile maize (Zea mays L.) is encoded by a continuous open reading frame of 1584 nucleotides. The predicted polypeptide encoded by the gene has a mol. wt. of 58 219 daltons and shows >60% amino acid sequence homology with the corresponding fungal and animal polypeptides. Two major transcripts of 2400 and 2300 nucleotides can be detected and the 5' end of the larger transcript maps to a sequence from -161 to -153 (relative to the initiator codon) which shows high homology to the yeast mitochondrial promoter. In mitochondrial DNA from the S male-sterile cytoplasm of maize, which also characteristically contain two low mol. wt. linear DNAs (S1 and S2), rearrangements just 5' (at -175) to the COX I gene, generate additional DNA restriction fragments containing entire copies of the gene. These rearrangements involve a sequence identical to the terminal 186 bp of the 208-bp inverted repeat sequence found at either end of the S1 and S2 DNAs.

Nucleotide sequence of the S-1 mitochondrial DNA from the S cytoplasm of maize

Mitochondria from the S male-sterile cytoplasm of maize contain unique DNA-protein complexes, designated S-1 and S-2. These complexes consist of double-stranded linear DNAs with proteins covalently attached to the 5' termini. To learn more about these unusual DNAs we have determined the complete nucleotide sequence of the S-1 DNA molecule (6397 bp). The sequence of S-2 has been previously determined. S-1 and S-2 are structurally similar and contain 1.7kb of sequence homology. S-1 is terminated by exact 208-bp inverted repeats that are identical with the terminal inverted repeats of S-2. S-1 and S-2 also contain a 1462-bp region of nearly perfect homology, which includes one of the terminal inverted repeats. The homology between the two molecules may be maintained, in part, by homologous recombination. S-1 has three long unidentified open reading frames, URF2 (1017 bp), URF3 (2787 bp) and URF4 (768 bp). URF2 occurs in the 1462-bp region of homology and is identical in length and location in both S-1 and S-2. Based on their structural organization and their viral-like characteristics, we propose that S-1 and S-2 code for functions involved with their maintenance and replication.

LBN, a male-sterile cytoplasm of maize, contains two double-stranded RNAs

Two nucleic acids species discovered in a male-sterile cytoplasm of maize have been shown to be double-stranded RNAs (dsRNAs) of apparent mol. wt. 1.9 and 0.5 × 10 6. This cytoplasm has been designated ‘LBN’ for ‘L cytoplasm in inbred W182BN’. The dsRNAs are associated with mitochondria and are maternally transmitted. Nuclear genotype influences the amount of dsRNA recoverable.

Maize mitochondrial DNA contains a sequence homologous to the ribulose-1,5-bisphosphate carboxylase large subunit gene of chloroplast DNA

The mitochondrial genome of maize contains a DNA sequence homologous to the chloroplast gene coding for the large subunit of ribulose-1,5-bisphosphate carboxylase (LS gene). The presence in mitochondrial DNA of both coding and flanking sequences of this gene has been demonstrated first, by cross hybridization between the purified organelle DNAs and between cloned mitochondrial and chloroplast DNA sequences and second, by in vitro transcription-translation of cloned mitochondrial DNA in an E. coli cell free system where a 21,000 dalton polypeptide is synthesized that can be precipitated with antibodies to wheat ribulose-1,5-bisphosphate carboxylase. In contrast to the 12 kb chloroplast homologous sequence found in the mitochondrial genome ( Stern and Lonsdale, 1982), the sequence homologous to the LS gene is unaltered in mitochondrial DNA isolated from the male sterile cytoplasms of maize. The LS gene homologous sequence in the mitochondrial genome is located some 65 kb from the 18S mitochondrial rRNA gene and approximately 20 kb from the mitochondrial DNA sequence having homology to the chloroplast 16S rRNA gene.

S1 and S2, the linear mitochondrial DNAs present in a male sterile line of maize, possess terminally attached proteins.

S1 and S2 are short linear mitochondrial DNA molecules found in a particular male sterile cytoplasm of maize. We show here that these DNA molecules and two other related linear DNA species found in maize mitochondria, have proteins attached, probably covalently, to their 5' ends. This is the first demonstration of such a linear DNA-terminal protein association in higher eukaryotes. Such proteins may be involved in priming replication of these DNAs.

Organization of mitochondrial DNA in normal and texas male sterile cytoplasms of maize

AbstractRecombinant DNA and hybridization techniques have been used to compare the organization of mitochondrial DNA (mtDNA) from normal (N) and Texas male sterile (T) cytoplasms of maize. Bam H1 restriction fragments of normal mtDNA were cloned and used in molecular hybridizations against Southern blots of Bam H1 digested N and T mtDNA. Fifteen of the 35 fragments were conserved in both N and T as indicated by hybridization to comigrating bands in their restriction patterns. Only three fragments produced autoradiographs whose differences could reasonably be attributed to single changes in the cleavage site of the enzyme while approximately half (17/35) of the clones resulted in more complicated differences between N and T. The autoradiographs produced by these 17 clones indicated multiple cleavage site changes and/or sequence rearrangements of the mtDNA. Patterns of six of these 17 clones indicated partial duplication of the sequence and two showed variation in the intensity of hybridization between N and T, which may be related to the molecular heterogeneity phenomenon found in maize mitochondrial genomes. The large proportion of changes observed between N and T mtDNA indicates that rearrangements may have played an important role in the evolution of the maize mitochondrial genome.

Cytoplasmic Reversion of cms-S in Maize: Association with a Transpositional Event

Spontaneous reversion to fertility in S male-sterile cytoplasm of maize is correlated with the disappearance of the mitochondrial plasmid-like DNA's, S-1 and S-2, and changes in the mitochondrial chromosomal DNA. Hybridization data indicate that one of the plasmid-like DNA's, S-2, is prominently involved in the mitochondrial DNA rearrangements.