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doi: 10.1242/10.1242/dev.00550

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The meiotic protein SWI1 is required for axial element formation and recombination initiation in Arabidopsis

¹ School of Biosciences, The University of Birmingham, Birmingham, B15 2TT, UK
² Institut National de la Recherche Agronomique, Station de Génétique et d'Amélioration des Plantes, Route de Saint-Cyr, 78026 Versailles, cedex, France
³ Department of Chemistry and Biochemistry, Miami University, Oxford, OH 45056, USA

View larger version (112K): [in a new window]   Fig. 1. Silver staining of spread nuclei at meiotic prophase in wild type and swi1-2 mutant. (A-D) Wild-type meiotic prophase. (A) Leptotene, axial elements appear. (B) Zygotene, synapsis is progressing. (C) Pachytene, synapsis is completed. (D) Diakinesis; five bivalents are condensing. (E-H) swi1-2 meiotic prophase. Ten univalents progressively condense without any sign of an axial element or synapsis. Scale bar: 10 µm.

View larger version (62K): [in a new window]   Fig. 2. ASY1 immunolocalization during meiosis in wild type and swi1-2 mutant. In each panel, the upper picture shows a DAPI-stained nucleus (blue) and the lower one shows the ASY1 immunolocalization (green) on the same nucleus. (A-C) Wild-type meiotic nuclei. (A) Pre-leptotene nucleus, ASY1 is associated with chromatin. (B) Leptotene, ASY1 localization stains the axial elements. (C) Pachytene, ASY1 localizes along synapsed chromosomes. (D-I) swi1-2 meiotic cells. (D,E) ASY1 localizes on chromatin as it does in wild type. (F,G) ASY1 is present on the condensing ten univalents. (H) The sister chromatid cohesion is released, resulting in 20 free chromatids on which ASY1 is still present. (I) A polyad composed of nine micronuclei on which ASY1 localizes. Scale bars: 10 µm.

View larger version (89K): [in a new window]   Fig. 3. SYN1/DIF1 immunolocalization during meiosis in wild type and swi1-2 mutant. In each panel, the upper picture shows a DAPI-stained nucleus (blue) and the lower one shows the SYN1/DIF1 immunolocalization (red) on the same nucleus. (A-D) Wild-type nuclei. (A) Pre-leptotene nucleus, SYN1/DIF1 is associated with chromatin. (B) Leptotene, SYN1/DIF1 localizes along chromosome axes. (C) Pachytene, SYN1/DIF1 immunolocalization stains synapsed chromosomes. (D) Metaphase I, a weak SYN1/DIF1 signal is detected on condensed bivalents. (E-H) swi1-2 meiotic cells. (E) SYN/DIF1 localizes on chromatin as it does in wild type. (F) SYN1/DIF1 surrounds the 10 condensing univalents. (G) While sister cohesion is released, SYN1/DIF1 remains on the chromatids. (H) A polyad, SYN1/DIF1 localizes in the four nuclei. Scale bars: 10 µm.

View larger version (74K): [in a new window]   Fig. 4. RAD51 immunolocalization during meiosis in wild type and swi1-2 mutant. RAD51 appears in green and the DAPI-stained DNA in blue. (A) Wild-type leptotene, RAD51 immunolocalization reveals numerous spots on chromosomes. (B) Wild-type pachytene, the number of RAD51 spots has decreased. (C-F) swi1-2 meiotic cells. (C) Early meiotic cells, no RAD51 signal. (D) 10 condensing univalents, no RAD51 signal. (E) 20 free chromatids, no RAD51 signal. (F) A polyad, RAD51 appears in the two nuclei. Scale bars: 10 µm.

View larger version (76K): [in a new window]   Fig. 5. swi1-2 mutation suppresses the dif1-1 chromosome fragmentation phenotype. (A) Metaphase I in a wild-type male meiocyte. Five bivalents are observed. (B) dif1-1 meiocyte exhibiting chromosome fragmentation. (C) swi1-2 meiocyte at the end of prophase. Twenty chromatids are observed. (D,E) swi1-2/dif1-1 double mutant meiocytes. (D) Sister chromatid cohesion is lost at the end of prophase I. (E) The resulting 20 free chromatids at metaphase I. (F) Some cells contained 10 condensed univalents. Scale bar: 10 µm.

View larger version (39K): [in a new window]   Fig. 6. SWI1 immunolocalization. (A) Three examples of meiotic cells expressing SWI1. (B) SWI1 is not detected at leptotene stage. (C) Diagrams outlining the three kinds of BrdU incorporation experiment. (D-I) Double localization of SWI1 and incorporated BrdU. In each panel, the upper picture shows a DAPI-stained nucleus (blue), the middle one the BrdU immunolocalization (green), and the lower one shows the SWI1 immunolocalization (red) on the same nucleus. (D) Experiment 1. Tissues were harvested directly at the end of the pulse. The cell shown is at S phase, according to the BrdU labeling, and expresses SWI1. (E-H) Experiment 2. Samples were harvested 2 hours, 6 hours or 10 hours after the end of the 2-hour pulse. After a 2-hour delay, half of the BrdU-positive cells express SWI1 (E) while the other half do not (F). After a 6-hour or a 10-hour delay, BrdU-positive cells do not express SWI1 (G-H). (I) Experiment 3. Samples were harvested at the end of an 8-hour BrdU treatment. Small meiotic cells express SWI1, and are BrdU negative, indicating they are in G1. Scale bar: 10 µm.