Abstract
BackgroundProgrammed DNA elimination and reorganization frequently occur during cellular differentiation. Development of the somatic macronucleus in some ciliates presents an extreme case, involving excision of internal eliminated sequences (IESs) that interrupt coding DNA segments (macronuclear destined sequences, MDSs), as well as removal of transposon-like elements and extensive genome fragmentation, leading to 98% genome reduction in Stylonychia lemnae. Approximately 20–30% of the genes are estimated to be scrambled in the germline micronucleus, with coding segment order permuted and present in either orientation on micronuclear chromosomes. Massive genome rearrangements are therefore critical for development.Methodology/Principal FindingsTo understand the process of DNA deletion and reorganization during macronuclear development, we examined the population of DNA molecules during assembly of different scrambled genes in two related organisms in a developmental time-course by PCR. The data suggest that removal of conventional IESs usually occurs first, accompanied by a surprising level of error at this step. The complex events of inversion and translocation seem to occur after repair and excision of all conventional IESs and via multiple pathways.Conclusions/Significance This study reveals a temporal order of DNA rearrangements during the processing of a scrambled gene, with simpler events usually preceding more complex ones. The surprising observation of a hidden layer of errors, absent from the mature macronucleus but present during development, also underscores the need for repair or screening of incorrectly-assembled DNA molecules.
Highlights
DNA elimination and reorganization occur in a variety of differentiating eukaryotic cells
MDS numbers m are consecutive in the macronucleus, and internal eliminated sequences (IESs) numbers i are consecutive in the micronucleus, as in chromosome formation to the beginning of DNA degradation (Figure 2)
We surveyed the population of partially processed DNA molecules from two scrambled genes at different time intervals during macronuclear development in two ciliates, Stylonychia lemnae and Oxytricha trifallax
Summary
DNA elimination and reorganization occur in a variety of differentiating eukaryotic cells. Non-coding DNA segments (IESs) [19] interrupt most of the ,30,000 genes in the germline genome of stichotrichous ciliates. Macronuclear development in these species removes intergenic DNA as well as deleting more than 100,000 IESs to allow assembly of the gene segments (MDSs) into functional genes. Development of the somatic macronucleus in some ciliates presents an extreme case, involving excision of internal eliminated sequences (IESs) that interrupt coding DNA segments (macronuclear destined sequences, MDSs), as well as removal of transposon-like elements and extensive genome fragmentation, leading to 98% genome reduction in Stylonychia lemnae. 20–30% of the genes are estimated to be scrambled in the germline micronucleus, with coding segment order permuted and present in either orientation on micronuclear chromosomes.
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