title: snRNA and heterochromatin formation are involved in DNA excision during macronuclear development in stichotrichous ciliates.
abstract: Several models for specific excision of micronucleus-specific DNA sequences during macronuclear development in ciliates exist. While the template-guided recombination model suggests recombination events resulting in specific DNA excision and reordering of macronucleus-destined sequences (MDS) guided by a template, there is evidence that an RNA interference-related mechanism is involved in DNA elimination in holotrichous ciliates. We describe that in the stichotrichous ciliate Stylonychia, snRNAs homologous to micronucleus-specific sequences are synthesized during macronuclear differentiation. Western and in situ analyses demonstrate that histone H3 becomes methylated at K9 de novo during macronuclear differentiation, and chromatin immunoprecipitation revealed that micronucleus-specific sequences are associated with methylated H3. To link both observations, expression of a PIWI homolog, member of the RNA-induced silencing complex, was silenced. In these cells, the methylated micronucleus-specific histone H3 variant "X" is still present in macronuclear anlagen and no K9 methylation of histone H3 is observed. We suggest that snRNA recruits chromatin-modifying enzymes to sequences to be excised. Based on our and earlier observations, we believe that this mechanism is not sufficient for specific excision of sequences and reordering of MDS in the developing macronucleus and propose a model for internal eliminated sequence excision and MDS reordering in stichotrichous ciliates.
authors.affiliation: Institute of Cell Biology, University Witten/Herdecke, Stockumer Str. 10, D-58453 Witten, Germany.
authors.forename: Stefan A
authors.initials: SA
authors.lastname: Juranek
authors.forename: Sina
authors.initials: S
authors.lastname: Rupprecht
authors.forename: Jan
authors.initials: J
authors.lastname: Postberg
authors.forename: Hans J
authors.initials: HJ
authors.lastname: Lipps
mesh_terms: D000818:Animals; D060565:Argonaute Proteins; D042002:Chromatin Assembly and Disassembly; D004247:DNA / Q000378:metabolism*; D029721:Drosophila Proteins; D020868:Gene Silencing; D006570:Heterochromatin / Q000378:metabolism*; D006657:Histones / Q000378:metabolism; D006877:Hydroxamic Acids / Q000378:metabolism; D016810:Hypotrichida / Q000166:cytology / Q000235:genetics* / Q000502:physiology; D048648:Macronucleus / Q000502:physiology*; D011500:Protein Synthesis Inhibitors / Q000378:metabolism; D011506:Proteins; D034622:RNA Interference; D012342:RNA, Small Nuclear / Q000378:metabolism*; D034743:RNA-Induced Silencing Complex
chemical_list: D060565:Argonaute Proteins; D029721:Drosophila Proteins; D006570:Heterochromatin; D006657:Histones; D006877:Hydroxamic Acids; D011500:Protein Synthesis Inhibitors; D011506:Proteins; D012342:RNA, Small Nuclear; D034743:RNA-Induced Silencing Complex; C116498:piwi protein, Drosophila; C012589:trichostatin A; D004247:DNA