Changes of the dna content of differentiating and adult macronuclei of the ciliate Loxodes magnus (karyorelictida)

Abstract

The DNA content of isolated micronuclei, differentiating macronuclei (macronuclear Anlagen), and adult macronuclei of Loxodes magnus was measured cytofluorimetrically in preparations stained with a Schiff-type reagent, auramine-SO2, following hydrochloric acid hydrolysis. The DNA content of the youngest macronuclear Anlagen proved to be the same as that of telophasic micronuclei (2 c). The Anlagen thus differentiate from micronuclei which are still in G1. The quantity of DNA in the macronuclear Anlagen thereafter rises to the 4-c level, simultaneously with DNA replication in the micronuclei which immediately follows mitosis. In non-dividing animals most micronuclei are already in G2. Adult macronuclei here contain on average 1.5 times more DNA than the micronuclei; their DNA content is about 5–6 c (in some individual nuclei, up to 10 c). These data are consistent with autoradiographic evidence indicating a weak DNA synthesis in the macronuclei of Loxodes and make likely the existence of partial DNA replication (e.g. gene amplification) in the macronuclei. The DNA content of adult macronuclei isolated from dividing animals proved to be significantly smaller than that of macronuclei isolated from non-dividing specimens of the same clone. In 3 clones studied, the former value amounted on average to 71–79, 78 and 95% of the latter, respectively. This drop of DNA content cannot be explained by ‘dilution’ of the old macronuclei with newly formed ones. The quantity of DNA in adult macronuclei thus seems to undergo cyclical changes correlated with cytokinesis, despite the fact that, in Loxodes magnus, the macronuclei themselves never divide and are simply segregated at every cell division. The macronuclei of Loxodes can be termed paradiploid or hyperdiploid.

Nuclear dualism without extensive DNA elimination in the ciliateLoxodes magnus

“…Nonetheless, other karyorelicts may have some degree of editing: developing MACs of Trachelonema sulcata have distinctly less DNA than MICs or mature MACs, suggesting some DNA elimination in early MAC development. 57 Old MACs in Loxodes have higher and more variable DNA content than recently matured MACs, 38 but this may be nonspecific amplification in senescent nuclei, as we did not observe distinct subclusters of MACs by DNA content (Figure 1D) nor evidence for differential amplification. Chromosome breakage and unscrambling were not directly addressed here, but unscrambling has only been found in conjunction with IES elimination, 5,58 and is likely also absent.…”

“…Unique k-mers did not show discernible frequency peaks (Figure 2A), nor was there an obvious cluster of k-mers with different coverage between the two libraries (Figure 2B), contrary to what would be expected if much of the genome was MIC-limited like in other ciliates (Supplementary Results 1), or if there was differential amplification of specific loci, as previously proposed. 38,43 There was no evidence for amplification of the rRNA locus in particular (Supplementary Results 2).…”

“…Schematic of nuclei in Loxodes striatus sexual and asexual cycles (above), adapted from 37,71,72 ; diagram of DNA content in Loxodes nuclei at different developmental stages (below), adapted from 38 . (B) Diagrammatic tree of ciliate classes (after 66 , branch lengths arbitrary) with genera used as laboratory models.…”

“…in the karyorelict Loxodes magnus, the DNA content in MACs is only about twice that of MICs. 38 Karyorelicts were formerly considered a "basal" group with a "primitive" form of nuclear development. 37,39 However, molecular phylogenies now show that karyorelicts are not the earliest-diverging branch but in fact sister to the Heterotrichea, [40][41][42] which have dividing, ampliploid MACs and at least one genus with extensive genome editing like other ciliates, 9,12 so non-dividing paradiploid MACs must be a derived character of karyorelicts.…”