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The organisation of genes for transfer RNA and ribosomal RNA in amoebae and plasmodia of Physarum polycephalum.

1. Using hybridisation techniques nuclei from both amoebae and plasmodia of Physarum polycephalum were found to contain 275 genes each coding for 5.8-S, 19-S and 26-S rRNA, 685 genes for 5-S rRNA and 1050 genes for tRNA. 2. Hybridisation of these RNA species to both amoebal and plasmodial DNA fractionated on CsCl gradients reveal that the 5.8-S, 19-S and 26-S rRNA genes are located at a satellite position (formula: see text) with respect to the main band of DNA, whereas 4-S RNA genes are located exclusively in the main band of DNA (formula: see text). 3. This result was confirmed by demonstrating that only the 5.8-S, 19-S and 26-S rRNA species hybridise to purified plasmodial ribosomal DNA. 4. The 19-S and 26-S rRNA genes of amoebae are located on extrachromosomal DNA molecules of a discrete size (Mr = 38 X 10(6)) with identical properties to plasmodial ribosomal DNA.

Cell Nucleus↗

Metabolic stability of the extrachromosomal ribosomal RNA genes in the slime mould Physarum polycephalum.

The rRNA genes of the slime mould Physarum polycephalum are located on free, linear DNA molecules of a discrete size, Mr=38X10(6). Using an isotope dilution technique we have examined the metabolic stability of these extrachromosomal genes during active, balanced growth. Microplasmodia, prelabelled with [3H]thymidine, were used to prepare synchronous surface plasmodial cultures which were subsequently grown on unlabelled medium. The gross synthesis of ribosomal DNA was then determined over three consecutive mitotic divisions from the ratio of 3H to 14C in a hybrid formed between the extracted ribosomal [3H]DNA and a [14C]rRNA probe. It was found that ribosomal DNA, like chromosomal DNA, is completely stable during active growth.

Drug Stability↗

Mobile genetic elements in animal cells and their biological significance.

Mobile genetic elements were discovered by McClintock while analysing unstable mutations in maize. The structural and functional studies of such elements became possible after their cloning, first from the genome of Drosophila melanogaster. In particular, Ilyin et al. demonstrated the varying location of the described elements in D. melanogaster chromosomes, thus providing the first evidence of their mobility. Mobile elements comprise a significant part of the genetic material in D. melanogaster (not less than 10%). Several classes of mobile elements do exist. Mobile dispersed genetic elements (mdg elements) are among the best characterized ones. Mdg elements are represented in the genome by dozens of families, each consisting of 10-150 copies. They are very similar structurally to proviruses of endogenous retroviruses. In particular, the both contain long terminal repeats (LTRs). The nucleotide sequences of LTRs and their flanking sequences of several mdg elements were determined. Their analysis suggested that RNA reverse transcription should be involved in the mdg amplification. It has been found that putative transposition intermediates, i.e. extrachromosomal DNA copies of mdg elements, are synthesized by reverse transcriptase in D. melanogaster culture cells. Another type of mobile genes is represented by P factor and similar elements. P factor seems to encode 'transposase' participating in direct excision and insertion of P elements themselves as well as of other mobile genes (mdg and fold-back elements). Besides these 'active transposons' which encode the enzyme machinery for transposition, a number of other sequences which may be transposed are present in the genome. RNAs synthesized on such elements can serve as a template for reverse transcriptase, and the DNA formed can then be inserted at new sites of the genome. Among such sequences are the so-called short ubiquitous repeats: B1 and B2 in mouse genome and Alu in human genome. We found that, at least in several cases, B-type sequences were located at the 3' end of mRNA. Short repetitive sequences were also detected at the 3' end of certain mRNAs of D. melanogaster. Usually the transpositions of mobile genes occur very rarely. However, under certain conditions, for example, in hybrid dysgenesis, they become more frequent. The strain with a mutation in cut locus was obtained in hybrid dysgenesis. This mutation depends on an insertion of mdg4 at the cut locus. Genetic instability in this strain is maintained for a long time. 'Transposition bursts' were found to occur in some germ cells.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗

Formation and fate of extranuclear chromatin bodies in Tetrahymena.

The fate of extranuclear chromatin bodies (ECBs) formed by exclusion of macronuclear material at the time of karyokinesis was followed quantitatively in Tetrahymena pyriformis strain GL-I. In a logarithmic growth phase culture, 51% of the dividing cells produced one (43%) or more (8%) ECBs. Most of these gradually disappear before the next cell division, but approximately 13% are retained and carried into subsequent cell cycles. The random distribution of ECBs into anterior or posterior daughter cells, their staining and morphological characteristics, and their rapid loss in cells in starvation medium, all indicate that ECBs play no more of a role in cellular activity than that of an internally produced food vacuole.

Animals↗

A double-stranded RNA mycovirus in Botrytis cinerea.

In wild-type Botrytis cinerea CVg25 strain we have detected the presence of extrachromosomal genetic elements corresponding to double-stranded RNA molecules. These genetic elements have been designated L, M1 and M2 with molecular sizes of 8.3, 2.0 and 1.4 kb, respectively. The visualization by electron microscopy of mycelium ultrathin sections from B. cinerea CVg25 showed the presence of isometric virus-like particles of about 40 nm in diameter. Linear sucrose gradient centrifugation of mycelium-free extracts was done to determine if the double-stranded RNAs were associated with virus-like particles. The gradient profile obtained at 260 and 280 nm revealed a major peak that was analyzed by both agarose-gel electrophoresis and electron microscopy. It was observed that only the L-double-stranded RNA molecule copurified with isometric virus-like particles. These virus-like particles had a similar morphology and size as those detected by electron microscopy in the mycelium sections. These results suggest that only the L-double-stranded RNA would be encapsidated.

Extrachromosomal Inheritance↗

Macrorestriction analysis of Desulfurella acetivorans and Desulfurella multipotens.

The genomes of the phylogenetically and physiologically unique bacteria Desulfurella acetivorans DSM 5264T and D. multipotens DSM 8415T were characterized and compared by pulsed field gel electrophoresis (PFGE). Macrorestriction patterns made of large PFGE separated DNA fragments were generated by digesting the genomic DNAs of both strains with the rare cutting restriction endonucleases ApaI, AscI, EagI, RsrII, SacII, SalI as well as with the intron encoded endonuclease I-CeuI. The sum of calculated fragment sizes from digests of the first six enzymes yielded estimates for the chromosome sizes of D. acetivorans with a mean of 1939.0 +/- 26.0 kb and for D. multipotens with a mean of 1864.0 +/- 23.0 kb. Within the patterns obtained from EagI and RsrII cleavages the apparent differences could be attributed to DNA insertion or deletion and to point mutation. The single, circular chromosomes of the two strains contain two copies of 23S rRNA genes each. Different extrachromosomal elements were detected in both strains.

DNA, Bacterial↗