Skip to main content
Fig. 3 | Epigenetics & Chromatin

Fig. 3

From: 27nt-RNAs guide histone variant deposition via ‘RNA-induced DNA replication interference’ and thus transmit parental genome partitioning in Stylonychia

Fig. 3

Differential excision of micronuclear sequence classes and immunolocalization of PIWI1 and H3K27me3 in polytene developing macronuclei. a, b Agarose gel electrophoresis of amplicons from PCR analyses used for the examination of sequence excision. x-axis/line labelling: time PC (hours). In each box the different columns represent replicate experiments. a MDP2 gene—PIWI1-minus samples (left) and non-RNAi control samples (right); b MaA81 repetitive bulk DNA—PIWI1-minus samples (left) and non-RNAi control samples (right). For each experiment several replicates are shown in columns. c–e Immunolocalization of PIWI1 (c, d; red) or H3K27me3 (e; yellow) during development (c1-4) and in polytene chromosomes at higher magnification (d, e). To-Pro-3 was used for DNA counterstaining (blue). Briefly as previously described [14], PIWI1 accumulates in parental macronuclei (p) during conjugation (c1) and persists immediately after cell separation (c2). Subsequently, PIWI1 vanishes from parental macronuclei and translocates to the prospective macronucleus (A) (c3-7). Thereafter, premature macronuclei, whose nanochromosomes undergo a second series of DNA replication until they reach their final copy numbers, become devoid of PIWI1. Remarkably, we observed an extranuclear PIWI1 spot and speculate that it could be a degradation body (c8). p: parental (old) macronucleus fragments; m(*): micronucleus (*during mitosis); A: anlage, developing macronucleus; M: macronucleus. Lines in c8 point to replication bands, the arrowhead points to an unknown PIWI1-positive extranuclear structure. Arrowheads in D point to areas, where the shape of the polytene chromosomes as well as the banding pattern can be well recognized

Back to article page