Chironomus thummi thummi contains a repetitive AT-rich 118 bp sequence mainly in the centromere regions and elsewhere in the genome (1). A large cluster of repeats is regularly present in the non-transcribed spacer of rDNA. Dimers and multimers of the repeat migrate slower in small pore gels than would be expected from their size. The results indicate a solenoidal structure with a coil girth of appr. 350 bp. This structure is most probably due to a highly periodic positioning of di-nucleotides of the type purinepurine or pyrimidine-pyrimidine with distances of appr. 10 bases. In a cluster~f 118 bp repeats, regions of dyad-symmetry are positioned such that a 142 -2 bp palindrome-frame is generated. Evidence is presented favouring the assumption that the repeat functions primarily in sister chromatid exchange.
The ribosomal DNAs from Ch. thummi piger and Ch. th. thummi were cloned and analysed by a variety of restriction endonucleases. Comparison of rDNA clones from the two subspecies revealed a considerable length difference: the length of the analysed rDNA cistrons is approximately 9.0 kb for Ch. th. piger and approximately 14.5 kb for Ch. th. thummi. The nearly 5 kb additional DNA in Ch. th. thummi is clearly located within the non-transcribed spacer region, and consists of AT-rich, repetitive DNA elements. These elements with a basic repeat length of approximately 120 bp, are arranged tandemly in stretches of up to about 50 identical copies, which are characterized by a cleavage site for ClaI restriction endonuclease. They are found only in the Ch. th. thummi rDNA clones and not in the Ch. th. piger clones. Southern hybridizations between cloned ribosomal DNA and "centromeric" highly repetitive DNA have shown that the ribosomal repetitive Cla-elements are closely related to a highly repetitive DNA sequence family, which is present in various chromosomal sites particularly the centromeres. Sequence analysis has revealed more than 90% homology between the ribosomal Cla-elements and the "centromeric" Cla-elements.--Since it is clear from cytological investigations that Ch. th. piger with the small rDNA repeating unit is the phylogenetically older subspecies, we postulate a transposition of Cla-elements into the nucleolar DNA during the evolution of Ch. th. thummi.
The rDNA of Ch. tepperi is homogeneous in structure with a repeat size of 8.4 kb. This size seems to be typical for the basic repeat unit in Chironomus species. Ch. th. piger rDNA cistrons are slightly increased in length (9.0 kb). In the non-transcribed spacer (NTS) an appr. 0.18 kb segment is additionally present in about 50 % of the repeats. Ch. th. thummi DNA contains largely heterogeneous rDNA repeats, mainly between 10 and 16 kb. The heterogeneity is due to varying numbers of 120 bp elements present in the NTS. The different spacer size classes are not randomly distributed. The short repetitive 120 bp elements (Cla I elements) hybridize in situ with the nucleolus and with centromere regions. The Cla I elements are regularly present in the thummi NTS, but are absent in the piger NTS. Only very few piger rDNA cistrons may contain Cla I elements.
In order to induce chromosomal rearrangements, males were exposed to x-rays and then mated to non-irradiated females. The number of each type of structural alteration was determined by examination of the polytene chromosomes of the F1 progeny. -- A comparison of the results with similar studies made on Drosophila revealed a significantly greater sensitivity in Phryne. Parallel to that an extremely high frequency of small inversions was ascertained in Phryne, and the observed ratio of inversions to translocations was the inverse of that which would be expected from purely mathematical considerations based on the lengths of the different chromosomes. These facts allow the conclusion that the paternal pronuclear chromosomes in Phryne are highly spiralized. Besides, the kinetochore-to-translocation-breakpoint distance was measured in both of the chromosomes involved in each reciprocal translocation and the differences (kinetochore-break distance differences) were registered and from them the arrangement of the chromosomes in the pronucleus of Phryne deduced. The data obtained support the assumption of an ordered, polar-field type of orientation. In Drosophila, in contrast, the comparable data showed that the pronuclear chromosomes are not spiralized and are randomly arranged (Bauer, 1939). -- These results seem to indicate that a close correlation exists between the different radiation sensitivities of Drosophila and Phryne and the different states of spiralisation and arrangements of their chromosomes in the pronucleus stage. It is hypothesized that the influence of the maternal genome on the degree of spiralization of the paternal chromosomes could account for differences in the pronuclear chromosome structure of both species.
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