High lipid order of Arabidopsis cell‐plate membranes mediated by sterol and DYNAMIN‐RELATED PROTEIN1A function

Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/109568/1/tpj12674.pdfhttp://deepblue.lib.umich.edu/bitstream/2027.42/109568/2/tpj12674-sup-0002-FigS2.pdfhttp://deepblue.lib.umich.edu/bitstream/2027.42/109568/3/tpj12674-sup-0001-FigS1.pdfhttp://deepblue.lib.umich.edu/bitstream/2027.42/109568/4/tpj12674-sup-0003-FigS3.pdfhttp://deepblue.lib.umich.edu/bitstream/2027.42/109568/5/tpj12674-sup-0004-FigS4.pd

Endocytosis restricts Arabidopsis KNOLLE syntaxin to the cell division plane during late cytokinesis

Cytokinesis represents the final stage of eukaryotic cell division during which the cytoplasm becomes partitioned between daughter cells. The process differs to some extent between animal and plant cells, but proteins of the syntaxin family mediate membrane fusion in the plane of cell division in diverse organisms. How syntaxin localization is kept in check remains elusive. Here, we report that localization of the Arabidopsis KNOLLE syntaxin in the plane of cell division is maintained by sterol-dependent endocytosis involving a clathrin- and DYNAMIN-RELATED PROTEIN1A-dependent mechanism. On genetic or pharmacological interference with endocytosis, KNOLLE mis-localizes to lateral plasma membranes after cell-plate fusion. Fluorescence-loss-in-photo-bleaching and fluorescence-recovery-after-photo-bleaching experiments reveal lateral diffusion of GFP-KNOLLE from the plane of division to lateral membranes. In an endocytosis-defective sterol biosynthesis mutant displaying lateral KNOLLE diffusion, KNOLLE secretory trafficking remains unaffected. Thus, restriction of lateral diffusion by endocytosis may serve to maintain specificity of syntaxin localization during late cytokinesis

Chromosome and DNA methylation dynamics during meiosis in autotetraploid Arabidopsis arenosa

Variation in chromosome number due to polyploidy can seriously compromise meiotic stability. In autopolyploids, the presence of more than two homologous chromosomes may result in complex pairing patterns and subsequent anomalous chromosome segregation. In this context, chromocenter, centromeric, telomeric and ribosomal DNA locus topology and DNA methylation patterns were investigated in the natural autotetraploid, Arabidopsis arenosa. The data show that homologous chromosome recognition and association initiates at telomeric domains in premeiotic interphase, followed by quadrivalent pairing of ribosomal 45S RNA gene loci (known as NORs) at leptotene. On the other hand, centromeric regions at early leptotene show pairwise associations rather than associations in fours. These pairwise associations are maintained throughout prophase I, and therefore likely to be related to the diploid-like behavior of A. arenosa chromosomes at metaphase I, where only bivalents are observed. In anthers, both cells at somatic interphase as well as at premeiotic interphase show 5-methylcytosine (5-mC) dispersed throughout the nucleus, contrasting with a preferential co-localization with chromocenters observed in vegetative nuclei. These results show for the first time that nuclear distribution patterns of 5-mC are simultaneously reshuffled in meiocytes and anther somatic cells. During prophase I, 5-mC is detected in extended chromatin fibers and chromocenters but interestingly is excluded from the NORs what correlates with the pairing patter

Dissecting sterol function during clathrin-dependent endocytosis and cytokinesis in Arabidopsis thaliana

Sterols are lipid components of eukaryotic membranes. Alterations of membrane sterol composition perturb the execution of cell division, which in diverse eukaryotes can have severe consequences for development of the organism. Partitioning of the cytoplasm during cell division occurs at the final stage of cell division named cytokinesis. In somatic plant cells, cytokinesis is initiated by fusion of membrane vesicles in the plane of cell division resulting in a transient compartment termed the cell plate. Cell plate maturation relies on temporal and spatial orchestration of membrane fusion and endocytosis. Impaired vesicle fusion or defects in endocytosis result in cytokinetic defects. In Arabidopsis thaliana, the KNOLLE and DYNAMIN-RELATED PROTEIN 1A (DRP1A) contribute to cytokinesis. KNOLLE mediates fusion of vesicles at the plane of cell division while DRP1A appears to be involved in cell plate maturation through its role in clathrin-mediated endocytosis. This thesis shows that KNOLLE is specifically restricted to the cell division plane through sterol-dependent endocytosis that involves a clathrin- and DRP1A-mediated mechanism. Sterols affect internalization of KNOLLE through their role in lateral membrane organization by keeping diffusion of KNOLLE to lateral membranes in check via its endocytic removal. It is shown that the cell plate represents a high-lipid-order membrane domain that depends on the correct composition and the right concentration of sterols. Accumulation of DRP1A at the cell plate requires correct sterol concentration and composition similar to high-lipid order. Conversely, high-lipid-order at the cell plate relies on DRP1A activity suggesting a feedback between DRP1A function and lipid order establishment. Finally, it is shown that sterols are also present at the tonoplast of dividing and elongated root cells. Taken together, the results reveal that formation of the cell plate in Arabidopsis thaliana depends on an intricate interplay between cytokinetic vesicle fusion, sterol-dependent lateral membrane and high-lipid-order domain organization as well as endocytic machinery function

Dissecting sterol function during clathrin-dependent endocytosis and cytokinesis in Arabidopsis thaliana

Sterols are lipid components of eukaryotic membranes. Alterations of membrane sterol composition perturb the execution of cell division, which in diverse eukaryotes can have severe consequences for development of the organism. Partitioning of the cytoplasm during cell division occurs at the final stage of cell division named cytokinesis. In somatic plant cells, cytokinesis is initiated by fusion of membrane vesicles in the plane of cell division resulting in a transient compartment termed the cell plate. Cell plate maturation relies on temporal and spatial orchestration of membrane fusion and endocytosis. Impaired vesicle fusion or defects in endocytosis result in cytokinetic defects. In Arabidopsis thaliana, the KNOLLE and DYNAMIN-RELATED PROTEIN 1A (DRP1A) contribute to cytokinesis. KNOLLE mediates fusion of vesicles at the plane of cell division while DRP1A appears to be involved in cell plate maturation through its role in clathrin-mediated endocytosis. This thesis shows that KNOLLE is specifically restricted to the cell division plane through sterol-dependent endocytosis that involves a clathrin- and DRP1A-mediated mechanism. Sterols affect internalization of KNOLLE through their role in lateral membrane organization by keeping diffusion of KNOLLE to lateral membranes in check via its endocytic removal. It is shown that the cell plate represents a high-lipid-order membrane domain that depends on the correct composition and the right concentration of sterols. Accumulation of DRP1A at the cell plate requires correct sterol concentration and composition similar to high-lipid order. Conversely, high-lipid-order at the cell plate relies on DRP1A activity suggesting a feedback between DRP1A function and lipid order establishment. Finally, it is shown that sterols are also present at the tonoplast of dividing and elongated root cells. Taken together, the results reveal that formation of the cell plate in Arabidopsis thaliana depends on an intricate interplay between cytokinetic vesicle fusion, sterol-dependent lateral membrane and high-lipid-order domain organization as well as endocytic machinery function