Frequencies of Circular Units of Nucleolar DNA in Oocytes of Two Insects, Acheta domesticus and Dytiscus marginalis, and Changes of Nucleolar Morphology during Oogenesis

Abstract

The organization of the extrachromosomal nucleolar material in oocytes of two insect species with different ovary types, the house cricket Acheta domesticus (panoistic ovary) and the water beetle Dytiscus marginalis (meroistic ovary), was studied with light and electron microscopic techniques. Stages early in oogenesis were compared with fully vitellogenic stages (mid-to-Iate diplotene). The arrangement of the nucleolar material undergoes a marked change from a densely aggregated to a dispersed state. The latter was characterized by high transcriptional activity. In spread and positively stained preparations of isolated nucleolar material, a high frequency of small circular units of transcribed rDNA was observed and rings with small numbers (1-5) of pre-rRNA genes were predominant. The observations suggest that the "extra DNA body" observed in early oogenic stages of both species represents a dense aggregate of numerous short circular units of nucleolar chromatin, with morphological subcomponents identifiable in ultrathin sections. These apparently remain in close association with the chromosomal nucleolar organizer(s). The observations further indicate that the individual small nucleolar subunit circles dissociate and are dispersed as actively transcribed rDNA units later in diplotene. The results are discussed in relation to principles of the ultrastructural organization of nucleoli in other cell types as well as in relation to possible mechanisms of gene amplification.

Lengths and patterns of transcriptional units in the amplified nucleoli of oocytes of Xenopus laevis

“…Such intra-axial heterogeneity is hardly explained by an amplification mechanism which involves starting units which contain only one pre-rRNA gene (for discussion see Wellauer et aL, 1976 b) , The existence of different rDNA units in the same amplified molecule seems to be compatible with the widely accepted concept that amplification takes place via a "rolling circle" mechanism (Hourcade et aL, 1973 ;Rochaix et aL , 1974 ;Bakken, 1975;Buongiorno-Nardelli et aL, 1976) only if one allows for multigenic rDNA circIes containing different units in the initial steps of the replicative process. Adjacent repeating units of different sizes have recently also been described in the amplified rDNA of the oocytes of two insect species, Dytiscus marginalis and Acheta domesticus (Trendelenburg et aL, 1976(Trendelenburg et aL, , 1977. Moreover, size differences of rDNA repeating units in the same strand (intraaxial heterogeneities) have been noted in spread prepanÜions of nucIeoli from primary nucIei of dasycladacean green algae (Spring et aL, , 1976Trendelenburg et aL, 1974) and from embryonic cells of Drosophila melanogaster (e.g.…”

“…Under preparative conditions that perfectly allow the deposition of single gene units of rDNA on the specimen grid (cf. Trendelenburg et al, 1976Trendelenburg et al, , 1977 we found only very rarely short linear or circular units (only three rings containing five and six repeating units, respectively, were encountered among a total of seve ral thousand repeating units evaluated, cf. Miller and Beatty, 1969 ; in general, rings of nucleolar chromatin were much rarer in oocytes of Xenopus laevis than in those of Triturus alpestris and T. cristatus).…”

“…The apparent spacer intercepts of nucleolar chromatin of Xenopus laevis show, at variable frequency , distinct groups of lateral fibrils of increasing lengths which are not continuous with the fibril gradient of the specific following prerRNA matri; unit. Such "prelude complexes " of variable lengths, some of which occupy most ofthe whole spacer intercept, have been observed in nucleolar chromatin from other organisms and cell types Trendelenburg et al , 1973Trendelenburg et al , , 1974Trendelenburg et al , , 1976Trendelenburg et al , , 1977Spring et al , 1974 ;Franke et al , 1976a). Their existence strongly suggests that parts of the apparent spacer region of rDNA are, at least occasionally, transcribed simultaneously with the pre-rRNA genes .…”

“…In contrast to earlier concepts emphasizing the homogeneity of rDNA repeating units in Xenopus (e.g., Wensink and Brown, 1971 ;Brown et al, 1972;Forsheit et al , 1974), recent results obtained from c1eavage patterns of rDNA with EcoRI restriction endonuclease have shown a striking size heterogeneity of repeating units which primarily reflects the heterogeneity of "spacer" lengths (e.g., Morrow et al , 1974 ;Wellauer et al, 1974Wellauer et al, , 1976aWellauer and Reeder, 1975). Heterogeneity of length of repeating units, including heterogeneity of apparent spacer units, has also been noted in the transcriptional units of spread nucleolar chromatin from diverse organisms such as certain green algae, insects, and amphibia (Miller and Beatty, 1969 ;Scheer et al , 1973;Spring et al , 1974Spring et al , , 1976Trendelenburg et al , 1974Trendelenburg et al , , 1976Trendelenburg et al , , 1977Berger and Schweiger, 1975;Foe et al , 1976). In order to answer the question whether the size heterogeneity of the repeating units observed in isolated Xenopus rDNA corresponds to the pattern observed in transcribed nucleolar chromatin we have undertaken a detailed quantitative study of the distribution and modes of arrangement of transcriptional units in the amplified rDNA of the Xenopus laevis oocytes.…”

Morphology of transcriptional units at different states of activity

“…Electron micrograph of a positively stained spread preparation of a ring of nucleolar chromatin isolated from diplotene oocytes of the water beetle, Dytiscus marginalis (for preparative details see Trendelenburg et al 1976Trendelenburg et al , 1977. This ring contains two transcribed genes coding for precursors to ribosomal RNAs and the interspersed apparent spacer intercepts.…”