Understanding the functions of SMC5/6 complex during the generative development in Arabidopsis

Fen YANG, M.Sc.

Doctoral thesis

Understanding functions of STRUCTURAL MAINTENANCE OF CHROMOSOMES 5/6 complex in seed developmet

Abstract:

The maintenance of genome stability is a vital issue for all organisms. Structural maintenance of chromosomes 5/6 (SMC5/6) complex is a crucial factor for preserving genome stability. Recently, studies in plants showed the SMC5/6 complex is important for plant fertility. However, the mechanism is little known so far. In this thesis, I aimed to investigate the functions of the SMC5/6 complex during generative development in diploid and autotetraploid plants of Arabidopsis (Arabidopsis thaliana). SMC5/6 complex is one of three SMC complexes, which are highly conserved to regulate chromosome architecture and genome organization in eukaryotes. The SMC5/6 complex is well known for its functions in DNA damage repair. We observed that loss-of-function mutations in NSE2 subunit of the SMC5/6 complex cause severe meiotic defects. The first defect is chromosome fragmentation observed in meiosis I, suggesting the SMC5/6 complex is partly required for the repair of SPO11-induced DNA double-strand breaks. The second independent defect is the absence of chromosome segregation in the first and/or the second meiotic division, leading to the formation of unreduced male gametes. The unreduced male gametes result in the production of triploid offspring in nse2 plants. And it may also cause seed abortion as the maternal and paternal genome dosage is disturbed in endosperm. The presence of aborted ovules showed that nse2 plants indicates any defects in female gametogenesis are maternally lethal. Polyploidization is a common phenomenon in the evolution of flowering plants. However, our knowledge about the maintenance of polyploid genome stability is still very limited. Our work uncovered the loss-of-function autotetraploid (4x) mutants of SMC5/6 complex enhance fertility defects, cause severe defects in meiosis, and produce hexaploid and aneuploid progeny, suggesting that the SMC5/6 complex is an important player in the maintenance of tetraploid genome stability. Additional, tetrads with micronuclei were formed in 4x mutant, which were not observed in the case of the diploid (2x) mutant pollen mother cells. Aneuploid offspring were equally caused maternally and paternally in 4x mutant. Rarely, hexaploid plants occurred by unreduced female gametes in 4x nse2 plants. The absence of aneuploidy offspring and the viable unreduced female gametes in 2x mutants sup-ports they are unique phenotypes of tetraploid plants, indicating the importance of certain molecular regulators may be changed when polyploidization occurs. In conclusion, our studies uncover a novel SMC5/6 complex function in the maintenance of gametophytic ploidy in both diploid and autotetraploid Arabidopsis. Our work in diploid and autotetraploid Arabidopsis supports that autotetraploid plants have a generally higher frequency of but also higher tolerance for aneuploidy. Moreover, our results emphasize the importance of studying the consequences of mutations in genes regulating the plant fertility in diploid versus polyploid conditions, which may provide the possibility to increase agri-culturally important traits as many crop species are polyploidy.

Abstract:

The maintenance of genome stability is a vital issue for all organisms. Structural maintenance of chromosomes 5/6 (SMC5/6) complex is a crucial factor for preserving genome stability. Recently, studies in plants showed the SMC5/6 complex is important for plant fertility. However, the mechanism is little known so far. In this thesis, I aimed to investigate the functions of the SMC5/6 complex during generative development in diploid and autotetraploid plants of Arabidopsis (Arabidopsis thaliana). SMC5/6 complex is one of three SMC complexes, which are highly conserved to regulate chromosome architecture and genome organization in eukaryotes. The SMC5/6 complex is well known for its functions in DNA damage repair. We observed that loss-of-function mutations in NSE2 subunit of the SMC5/6 complex cause severe meiotic defects. The first defect is chromosome fragmentation observed in meiosis I, suggesting the SMC5/6 complex is partly required for the repair of SPO11-induced DNA double-strand breaks. The second independent defect is the absence of chromosome segregation in the first and/or the second meiotic division, leading to the formation of unreduced male gametes. The unreduced male gametes result in the production of triploid offspring in nse2 plants. And it may also cause seed abortion as the maternal and paternal genome dosage is disturbed in endosperm. The presence of aborted ovules showed that nse2 plants indicates any defects in female gametogenesis are maternally lethal. Polyploidization is a common phenomenon in the evolution of flowering plants. However, our knowledge about the maintenance of polyploid genome stability is still very limited. Our work uncovered the loss-of-function autotetraploid (4x) mutants of SMC5/6 complex enhance fertility defects, cause severe defects in meiosis, and produce hexaploid and aneuploid progeny, suggesting that the SMC5/6 complex is an important player in the maintenance of tetraploid genome stability. Additional, tetrads with micronuclei were formed in 4x mutant, which were not observed in the case of the diploid (2x) mutant pollen mother cells. Aneuploid offspring were equally caused maternally and paternally in 4x mutant. Rarely, hexaploid plants occurred by unreduced female gametes in 4x nse2 plants. The absence of aneuploidy offspring and the viable unreduced female gametes in 2x mutants sup-ports they are unique phenotypes of tetraploid plants, indicating the importance of certain molecular regulators may be changed when polyploidization occurs. In conclusion, our studies uncover a novel SMC5/6 complex function in the maintenance of gametophytic ploidy in both diploid and autotetraploid Arabidopsis. Our work in diploid and autotetraploid Arabidopsis supports that autotetraploid plants have a generally higher frequency of but also higher tolerance for aneuploidy. Moreover, our results emphasize the importance of studying the consequences of mutations in genes regulating the plant fertility in diploid versus polyploid conditions, which may provide the possibility to increase agri-culturally important traits as many crop species are polyploidy.

Keywords

SMC5/6 complex, NSE2, meiosis, seed development, polyploidy, genome stability, Arabidopsis

Language used: English

Date on which the thesis was submitted / produced: 14. 12. 2021

Thesis defence

Palacký University Olomouc

Faculty of Science

Doctoral programme / field:
Biology / Molecular nad Cell Biology