Plastid inheritance in Pisum sativum L.

Summary Cultivar variability for levels of plastid DNA (cpDNA) in the germ cell line of germinated pea pollen has suggested the possibility of biparental plastid transmission. In order to examine this possibility further, RFLP markers were used to follow the transmission of cpDNA from parents to their F₁ offspring. Results from these inheritance studies clearly indicate the presence of only maternal plastid markers in the F₁ progeny of each cross examined, irrespective of the pollen cpDNA levels of the paternal parent. The same result is obtained for F₁ progeny produced from crosses using pollen characterized by comparatively high cpDNA content, even when offspring are sampled at early developmental stages. Thus, there appears to be little correspondence between pollen cytological data indicating potential paternal plastid transmission and data from molecular marker studies confirming that P. sativum generally follows a uni-parental-maternal mode of plastid inheritance. Insufficient F₁ progeny were examined to exclude instances of trace biparentalism.     

Plastid inheritance in Oenothera: organelle genome modifies the extent of biparental plastid transmission

Summary The transmission abilities of four out of the five major plastome types of Oenothera (I–V) were analyzed in a constant nuclear background by assessing both the frequency of biparental inheritance and the extent of variegation in the progeny. Reciprocal crosses were performed between plants carrying one of four wild-type plastomes and plants carrying one of seven white plastid mutants. The frequency of biparental plastid transmission ranged from 0 to 56% depending on the plastid types involved in the crosses. The transmission abilities of the four representative wild-type plastids appear to be in the order of I > III > II > IV in the nuclear background of O. hookeri str. Johansen. In general, variegated seedlings from crosses that produced a higher frequency of biparental plastid transmission also had an increased abundance of tissue containing plastids of paternal origin. Although the transmission abilities of most Oenothera plastid mutants are comparable to the wild-type plastids, three mutant plastids derived from species having different type I plastids show three distinguishable transmission patterns. This study confirms the significant role of the plastome in the process of plastid transmission and possibly in plastid multiplication. However, the hypothesis of differential plastid multiplication rates suggested by earlier studies can explain the results only partially. The initiation of plastid multiplication within the newly formed zygote also seems to be plastome-dependent.     

Molecular analysis of mitochondrial DNA and its inheritance in Epilobium

Summary The mitochondrial genome of four Epilobium species has been characterized by restriction analysis and hybridizations with gene probes from Oenothera. Mitochondrial DNA of Epilobium has a complex restriction fragment pattern and an estimated size of about 320 kb. All species exhibit specific restriction patterns. Plasmid-like DNA molecules of 0.3 kb to 1.2 kb are found in preparations of undigested nucleic acids of mitochondria from E. montanum, E. watsonii, and E. lanceolatum. In contrast, the mitochondria of E. hirsutum contain double-stranded RNAs of 2.7 kb. The location of the genes for cytochrome c oxidase subunits I and III on the mitochondrial DNA seems to be conserved in those species analyzed. However, the genes for subunit II of this complex, and for the alpha subunit of ATPase, are located on different restriction fragments in the mitochondrial genomes of certain species. The location of the COX II gene on different BamHI fragments in E. watsonii and E. lanceolatum has been used for the analysis of mitochondrial inheritance in reciprocal hybrids. Like the plastids, mitochondria are inherited maternally in Epilobium.Abbreviations kb kilobase pairs - mtDNA mitochondrial DNA     

Plastid DNA is not detectable in the male gametes and pollen tubes of an angiosperm (Antirrhinum majus) that is maternal for plastid inheritance

Summary Prior cytological observations using DAPI/epifluorescence microscopy have suggested that the method could be used to rapidly screen plant species for their potential mode of plastid DNA transmission. Cytoplasmic DAPI-DNA aggregates were observed in generative cells of germinated pollen of Medicago sativa (alfalfa), a species known genetically to display biparental transmission, but not in Antirrhinum majus (snapdragon), a species known to be maternal for plastid transmission. If, as suggested, these aggregates are plastid DNA nucleoids, then M. sativa pollen should contain plastid DNA detectable by molecular biology methods and A. majus pollen should not. Total DNA was isolated from germinated pollen and analyzed by Southern blot hybridization. A clone containing part of the rbcL gene from the garden pea plastome was used as a probe for plastid DNA. This probe hybridized with a restriction fragment from M. sativa pollen DNA, but not detectably with A. majus pollen DNA, thereby corroborating the identification of the cytoplasmic DAPI-DNA aggregates in M. sativa pollen as plastid DNA, and confirming the cytologically determined absence of plastic DNA in A. majus pollen.     

Maternal inheritance of chloroplast genome and paternal inheritance of mitochondrial genome in bananas (Musa acuminata)

Restriction fragment length polymorphisms (RFLPs) were used as markers to determine the transmission of cytoplasmic DNA in diploid banana crosses. Progenies from two controlled crosses were studied with heterologous cytoplastmic probes. This analysis provided evidence for a strong bias towards maternal transmission of chloroplast DNA and paternal transmission of mitochondrial DNA in Musa acuminata. These results suggest the existence of two separate mechanisms of organelle transmission and selection, but no model to explain this can be proposed at the present time. Knowledge of the organelle mode of inheritance constitutes an important point for phylogeny analyses in bananas and may offer a powerful tool to confirm hybrid origins.     

Biased organelle transmission in somatic hybrids ofLycopersicon esculentum andSolanum lycopersicoides

Summary Restriction fragment length polymorphisms (RFLPs) were used to determine the transmission of organelle genomes in somatic hybrid plants of tomato and its wild relativeSolanum lycopersicoides. Biased frequencies of organelle combinations were observed in a population of 70 somatic hybrid plants each derived from a separate callus. The plastids in 68 of 70 hybrids examined were fromL. esculentum. One of the remaining hybrids, plant 240, hadS. lycopersicoides plastids and the other, plant 63, had a mixture of parental plastids. Forty-six of the same 70 plants were analyzed for mtDNA and all had that ofS. lycopersicoides including plant 240. One of these hybrids had novel mtDNA fragments which mayhave resulted from recombination or rearrangement. The biased transmission may have resulted from an initial unequal input of organelles, differential replication of organelles, or nucleo-organelle incompatibility.Michigan Agricultural Experiment Station Journal Article No. 12538. Supported by Grant No. I-751-84R from BARD, The United States-Israel Binational Agricultural Research and Development Fund     

Plastid inheritance in Oenothera: paternal input may influence transmission patterns

Summary Prior genetic analysis of Oenothera to assess the mechanism(s) controlling differential (biparental) plastid transmission patterns have indicated that the plastome plays an integral role. However, the influence of putative variation in paternal plastid input remains unclear. Pollen collected from Oenothera hookeri plants containing one of four different plastome types (I–IV) in a constant nuclear background (A₁A₁) was examined cytologically by DAPI/ epifluorescence microscopy. The number of plastid DNA aggregates per pollen generative cell was found to differ significantly. Plants containing plastome types I or II displayed an average of about ten plastid DNA aggregates per generative cell whereas plants containing types III or IV displayed, on average, 15 plastid DNA aggregates. The potential paternal plastid contribution to the egg cell at syngamy (III=IV>I=II) differs from the previously determined survival frequencies of the same four plastid types (I>III>II>IV) progeny.Scientific article no. A-5036, contribution no. 8084 of the Maryland Agricultural Experimental Station     

Restriction map and clone bank of tomato plastid DNA

Summary Tomato plastid DNA has been isolated from leaf chloroplasts andPst1 fragments cloned in the plasmid vector pUC8. Recombinant plasmids containing all but the largestPst1 fragment were identified by colony hybridisation. A restriction map of the plastid DNA for four restriction endonucleases was generated by digestion of the clonedPst1 fragments and by hybridisation to Southern blots. The plastid DNA is a circular molecule of 156.6–159.4 kbp with a large inverted repeat, and shows high conservation of restriction sites with plastid DNA from tobacco andPetunia.     

Non-random plastid segregation in somatic hybrids of Datura innoxia with several other Solanaceous species

Summary A non-random plastid segregation was found in somatic hybrids of Datura innoxia with seven different Solanaceous species. 14 out of 17 examined somatic hybrids showed the plastid features of Datura innoxia. Within the limits of sensitivity of the applied methods, one line could be shown to contain mixed plastids. Since sexual offspring of this line contains only one set of plastids, it is assumed that this is probably a periclinal chimaera due to the plastome, i.e., the plastid mixture is present on a plant rather than a cell level.     

Plastid fusion as an agent to arrest sorting out

Summary Plastid fusions were noted in an ultrastrucutal study of a mutant of Hosta showing slow sorting out of plastid genes. These data suggest that fusions between wild type and mutant plastids might increase the mixing of plastid DNA and hence slow the process of sorting out. It is likely that peripheral reticula are involved in the process of plastid fusion between the mutant and wild-type plastids.     

Transfer of paternal mitochondrial DNA during fertilization of honeybee (Apis mellifera L.) eggs

Strict maternal inheritance of mitochondrial (mt) DNA is believed to be the rule in most eukaryotic organisms because of exclusion of paternal mitochondria from the egg cytoplasm during fertilization. In honeybees, polyspermic fertilization occurs, and many spermatozoa, including their mitochondria-rich flagellum, can completely penetrate the egg, thus allowing for a possibly high paternal leakage. In order to identify paternal mtDNA in honeybee eggs, restriction fragment length polymorphisms (RFLP) of different subspecies were used. Total DNA extracts of different developmental stages of an Apis mellifera carnica x Apis mellifera capensis hybrid brood were tested with a radioactively-labelled diagnostic mtDNA probe. Densitograms of autoradiographs indicated that the male contribution represents up to 27% of the total mitochondrial DNA in the fertilized eggs 12 h after oviposition. In subsequent developmental stages the portion of paternal mtDNA slowly decreased until hatching of the larvae when only traces were found. Although rapid disintegration of paternal mtDNA does not occur, the initially high paternal mitochondrial contribution is not maintained in the adult animal.     

Maternal transmission of mitochondrial DNA in interspecific hybrids of Populus

Summary Restriction fragment analysis was conducted to investigate the mode of inheritance of mitochondrial (mt) DNA in F₁ progeny of two P. deltoides x P. deltoides, three P. deltoides x P. nigra, and two P. deltoides x P. maximowiczii controlled crosses, and in Populus x canadensis by using 16 restriction endonucleases and two heterologous probes of cloned mtDNA fragments of maize. Five restriction fragment length polymorphisms (RFLPs) of mtDNA differentiated P. deltoides from P. nigra, whereas three RFLPs of mtDNA separated P. deltoides from P. maximowiczii. In all cases, F₁ progeny of P. deltoides x P. nigra, and P. deltoides x P. maximowiczii, crosses had mtDNA restriction fragments of only their maternal P. deltoides parents. P. x canadensis had mtDNA restriction fragments of only P. deltoides. F₁ progeny of intraspecific P. deltoides crosses also had the same mtDNA fragments as their maternal parent. The results clearly demonstrate uniparental-maternal inheritance of the mitochondrial genome in F₁ interspecific hybrids of P. deltoides with P. nigra and P. maximowiczii.     

Cultivar variability for the presence of plastid DNA in pollen of Pisum sativum L.: implications for plastid transmission

Summary The mode of plastid transmission in the garden pea (Pisum sativum L.) was analyzed cytologically using the DNA-fluorochrome 4,6-diamidino-2-phenylindole (DAPI) in conjunction with epifluorescence microscopy. The reproductive cells of mature pollen obtained from 12 inbred lines and cv Early Alaska were examined for the presence or absence of DAPI-stained plastid DNA aggregates. Plastid DNA was detected in all 13 pea lines examined, although there was variability with regard to the percentage of pollen graines showing plastid DNA aggregates of generative cells (ranging from 3% in accession 82-12r to 65% in accession 82-14n). These cytological results may indicate genetic variability for plastic DNA inheritance in the garden pea. This paper is dedicated to the memory of Gerald A. Marx     

Inheritance of mitochondrial DNA in sexual crosses and protoplast cell fusions in Lentinula edodes

By using mitochondrial DNA (mtDNA) restriction fragment length polymorphisms (RFLPs) as genetic markers, the modes of mitochondrial inheritance in sexual crosses and protoplast cell fusions of the higher basidiomycete Lentinula edodes were examined. All newly established dikaryons from reciprocal crosses between compatible monokaryons carrying different mtDNA RFLP phenotypes retained mtDNA genotypes from one of the monokaryons, suggesting that mitochondrial inheritance is principally uniparental. In contrast, it was shown that recombinant mtDNA genomes arose in some dikaryons obtained after protoplast cell fusion. Based on these results, a possible mechanism for mitochondrial inheritance in L. edodes is discussed.     

Angelman and Prader-Willi syndromes share a common chromosome 15 deletion but differ in parental origin of the deletion

Many Prader-Willi syndrome (PWS) and Angelman syndrome (AS) patients have a cytogenetic deletion of 15q11q13. While AS and PWS share a similar cytogenetic anomaly, they have very different clinical phenotypes. DNAs from 4 AS patients were examined using 5 chromosome 15q11q13-specific cloned DNA segments. With the present level of resolution, the molecular deletions between AS and those previously reported for PWS did not appear to differ. However, in contrast to the paternal inheritance of the deleted chromosome 15 observed in the majority of PWS patients, maternal inheritance of the deleted chromosome 15 was demonstrated in the AS patients by restriction fragment length polymorphisms (RFLPs).     

Plastid DNA from Pyrenomonas salina (Cryptophyceae): physical map,genes, and evolutionary implications

Summary Cryptomonads are thought to have arisen from a symbiotic association between a eukaryotic flagellated host and a eukaryotic algal symbiont, presumably related to red algae. As organellar DNAs have proven to be useful tools in elucidating phylogenetic relationships, the plastid (pt) DNA of the cryptomonad alga Pyrenomonas salina has been characterized in some detail. A restriction map of the circular 127 kb ptDNA from Pyrenomonas salina was established. An inverted repeat (IR) region of about 5 kb separates two single-copy regions of 15 and 102 kb, respectively. It contains the genes for the small and large subunit of rRNA. Ten protein genes, coding for the large subunit of ribulose-1,5-bisphosphate carboxylase, the 47 kDa, 43 kDa and 32 kDa proteins of photosystem II, the ribosomal proteins L2, S7 and S11, the elongation factor Tu, as well as the - and -subunits of ATP synthase, have been localized on the restriction map either by hybridization of heterologous gene probes or by sequence homologies. The gene for the plastidal small subunit (SSUr) RNA has been sequenced and compared to homologous SSU regions from the cyanobacterium Anacystis nidulans and plastids from rhodophytes, chromophytes, euglenoids, chlorophytes, and land plants. A phylogenetic tree constructed with the neighborliness method and indicating a relationship of cryptomonad plastids with those of red algae is presented.     

Light and benzylaminopurine induce changes in ultrastructure and gene expression in plastids of Petunia hybrida cell cultures

Summary The regulatory effect of light and the cytokinin 6-benzylaminopurine (BA) on the plastid ultrastructure and plastid DNA gene expression is studied in white and mutant green cell suspension cultures of Petunia hybrida. By electron microscopy we show that both light and 6-benzylaminopurine induce the formation of thylakoid membranes and grana structures in plastids of the green cultures. For membrane formation in plastids of white cultures, light in combination with BA is required. Light and benzylaminopurine also influence the plastid DNA gene expression. By in-organello protein synthesis with isolated plastids we show that light as well as benzylaminopurine affects the synthesis of plastid DNA encoded proteins. A characteristic effect of benzylaminopurine on plastids from white and green cultures is the reduction in the synthesis of the CFI subunits of 55,000 and 57,000 D, and the reduction in the synthesis of large polypeptides with a molecular weight higher than 67,000 D. In contrast to benzylaminopurine, light only affects the DNA gene expression of plastids from white cell cultures, that are in a very early stage of plastid development. Light stimulates the synthesis of polypeptides with a molecular weight of 84,000, 70,000 and 46,000 D which are encoded by cpDNA in these white culture plastids. In green cell cultures both plastids with a etioplast-like phenotype and with a chloroplast like morphology synthesize similar polypeptides, resulting in the same polypeptide pattern. Our results indicate that qualitative differences in plastid DNA gene expression as an effect of light do occur but only in plastids at very early stages of chloroplast development. We observe a gradual reduction in the number of high molecular weight polypeptides at later stages of chloroplast development. This suggests that these large polypeptides are characteristic for plastids at an early developmental stage.Abbreviations LSU of RuBPCase large subunit of Ribulose-1, 5-bisphosphate carboxylase - CF₁ coupling factor of the ATPase complex - LCH chlorophyll a/b protein - BA 6-benzylaminopurine - cpDNA chloroplast DNA     

Parental origin of chromosome 5 deletions in the cri-du-chat syndrome

The parental origin of de novo deletions leading to the cri-du-chat syndrome has been investigated. Since the cri-du-chat syndrome is correlated with deletions involving the short arm of chromosome 5 (5p), DNA fragments known to detect restriction fragment length polymorphisms (RFLPs) along 5p were used to establish whether the paternal or the maternal chromosome had suffered the deletion. In cases where only one parent was available, somatic cell hybrids were used in conjunction with RFLP analysis to determine the origin of the deleted chromosome. The deleted chromosome 5 was of paternal origin in 20/25 cases.     

Evidence for tissue-specific cytosine-methylation of plastid DNA from Zea mays

Summary Total DNA isolated from leaves, etiolated seedlings, roots, endosperm or embryos of Zea mays was digested separately with each of the restriction enzymes HpaII, MspI and HhaI, and the resulting fragment patterns, which were specific for the plastid rRNA operon, were analyzed by Southern hybridization. While most of the fragment patterns were consistent with previously established physical maps, the partial resistance shown by one HpaII site and one HhaI site, both of which reside in the 16S/23S rDNA spacer region, was observed in DNA isolated from embryo, root tissue and endosperm. The partially resistant HpaII site was susceptible to cleavage with restriction enzyme MspI. From this and from the known inhibition of restriction enzyme Hhal at methylated HhaI sites, we conclude that the partial resistance of the two sites is caused by C-specific methylation of plastid DNA in the respective tissues. The tissue specificity of this DNA methylation is likely to reflect a differential expression of plastome encoded genes.