Dividing plant cells perform a remarkable task of building a new cell wall within the cytoplasm in a few minutes. A long-standing paradigm claims that this primordial cell wall, known as the cell plate, is generated by delivery of newly synthesized material from Golgi apparatus-originated secretory vesicles. Here, we show that, in diverse plant species, cell surface material, including plasma membrane proteins, cell wall components, and exogenously applied endocytic tracers, is rapidly delivered to the forming cell plate. Importantly, this occurs even when de novo protein synthesis is blocked. In addition, cytokinesis-specific syntaxin KNOLLE as well as plasma membrane (PM) resident proteins localize to endosomes that fuse to initiate the cell plate. The rate of endocytosis is strongly enhanced during cell plate formation, and its genetic or pharmacological inhibition leads to cytokinesis defects. Our results reveal that endocytic delivery of cell surface material significantly contributes to cell plate formation during plant cytokinesis.

(D.W.M.) Brefeldin A (BFA) inhibits exocytosis but allows endocytosis, making it a valuable agent to identify molecules that recycle at cell peripheries. In plants, formation of large intracellular compartments in response to BFA treatment is a unique feature of some, but not all, cells. Here, we have analyzed assembly and distribution of BFA compartments in development-and tissue-specific contexts of growing maize (Zea mays) root apices. Surprisingly, these unique compartments formed only in meristematic cells of the root body. On the other hand, BFA compartments were absent from secretory cells of root cap periphery, metaxylem cells, and most elongating cells, all of which are active in exocytosis. We report that cell wall pectin epitopes counting rhamnogalacturonan II dimers cross-linked by borate diol diester, partially esterified (up to 40%) homogalacturonan pectins, and (134)-␤-d-galactan side chains of rhamnogalacturonan I were internalized into BFA compartments. In contrast, Golgi-derived secretory (esterified up to 80%) homogalacturonan pectins localized to the cytoplasm in control cells and did not accumulate within characteristic BFA compartments. Latrunculin B-mediated depolymerization of F-actin inhibited internalization and accumulation of cell wall pectins within intracellular BFA compartments. Importantly, cold treatment and protoplasting prevented internalization of wall pectins into root cells upon BFA treatment. These observations suggest that cell wall pectins of meristematic maize root cells undergo rapid endocytosis in an F-actin-dependent manner.Eukaryotic cells perform endomembrane flow accomplished by vesicles shuttling among endoplasmic reticulum (ER), Golgi apparatus (GA), the plasma membrane (PM), and endosomes (for plants see Robinson et al., 1998; Hawes et al., 1999). These compartments and pathways of endomembrane flow are highly conserved in unicellular yeast, higher plants, and animals (for plant cells, see Robinson et al., 1998; Hawes et al., 1999). A major breakthrough in our current understanding of this complex endomembrane flow was provided by rediscovery of the fungal metabolite brefeldin A (BFA; Fujiwara et al., 1988). BFA action prevents vesicle formation in the exocytosis pathway by stabilizing abortive complexes between conserved ADP ribosylation factor 1 (ARF1) and the Sec7 domain of its guanine nucleotide exchange factor during the assembly of coat protein complexes of budding vesicles (for plants see, Pimpl et al., 2000;Robineau et al., 2000). Because of this action, BFA inhibits anterograde vesicular pathways while allowing endocytosis and some retrograde pathways to proceed further (Miller et al., 1992; Gaynor et al., 1998; Belanger and Quatrano, 2000). Moreover, BFA inhibits the endosome to vacuole transport in budding yeast (Gaynor et al., 1998).The introduction of BFA to investigate the cell biology of endomembrane flow in plant cells occurred some years later (Satiat-Jeunemaitre and Hawes, 1992), but most of the major findings concerning the effects of...

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The Arabidopsis homolog of trithorax, ATX1, regulates numerous functions in Arabidopsis beyond the homeotic genes. Here, we identified genome-wide targets of ATX1 and showed that ATX1 is a receptor for a lipid messenger, phosphatidylinositol 5-phosphate, PI5P. PI5P negatively affects ATX1 activity, suggesting a regulatory pathway connecting lipid-signaling with nuclear functions. We propose a model to illustrate how plants may respond to stimuli (external or internal) that elevate cellular PI5P levels by altering expression of ATX1-controlled genes.epigenetic regulation ͉ lipid signaling P roteins of the trithorax family activate the early homeotic genes that regulate animal development and embryonic pattern formation (1, 2). A major difference in the developmental process in plants is that organ formation is not restricted to the embryonic state, differentiation and organogenesis occurring throughout the lifespan of the organism. In plants, as in animals, homeosis is a consequence of a mutation of a homeotic gene. Usually, homeotic genes encode transcription factors. Unlike the animal counterparts, however, many of the plant homeotic genes belong to the MADSbox family (3). Despite the difference in structure, plant homeotic genes, like animal counterparts, are controlled by factors belonging to the trithorax family (4). Mutation of the Arabidopsis homolog of trithorax, ATX1, causes numerous developmental defects in the formation, placement, and identity of flower organs: Petals (second-whorl organs) were seen to develop stems, a third-whorl feature; stamens (third-whorl organs) developed ovules, a fourthwhorl characteristic (4).The signature feature of all trithorax proteins is the presence of the highly conserved SET [SuVar (3-9)-E(z)-trithorax] domain. The discovery that the SET domain peptides carry histone methyltransferase activity (5) provided critical evidence that chromatinmodifying activities function as epigenetic regulators. Certain lysines at the histone tails can be either acetylated or methylated, creating recognition sites for cellular repressive or activating complexes (6). SET domains of the trithorax family can methylate lysine 4 of histone H3, a modification associated with transcriptional activation (7). The SET domain of ATX1 has histone H3-K4 methyltransferase activity and can activate expression of Arabidopsis genes (4, 8). Thus, biochemical and genetic evidence define ATX1 as a functional homolog of the animal trithorax genes.Regulation of homeotic genes is only one possible role for trithorax (9, 10). In Arabidopsis, atx1 mutants displayed stem-, root-, and leaf-growth defects, indicating that the plant homolog of trithorax has pleiotropic roles (4). By whole genome expression profiling, we determined that Ϸ1,700 genes changed robust expression as a result of ATX1 loss of function. The altered expression of these genes provides a probable molecular basis underlying the pleiotropic functions of ATX1.The most important result of the study reported here is the finding that ATX1 can specifi...

The fluorescent dye Lucifer Yellow (LY) is a well-known and widely-used marker for fluid-phase endocytosis. In this paper, both light and electron microscopy revealed that LY was internalized into transition zone cells of the inner cortex of intact maize root apices. The internalized LY was localized within tubulo-vesicular compartments invaginating from the plasma membrane at actomyosin-enriched pit-fields and individual plasmodesmata, as well as within adjacent small peripheral vacuoles. The internalization of LY was blocked by pretreating the roots with the F-actin depolymerizing drug latrunculin B, but not with the F-actin stabilizer jasplakinolide. F-actin enriched plasmodesmata and pit-fields of the inner cortex also contain abundant plant-specific unconventional class VIII myosin(s). In addition, 2,3 butanedione monoxime, a general inhibitor of myosin ATPases, partially inhibited the uptake of LY into cells of the inner cortex. Conversely, loss of microtubules did not inhibit fluid-phase endocytosis of LY into these cells. In conclusion, specialized actin- and myosin VIII-enriched membrane domains perform a tissue-specific form of fluid-phase endocytosis in maize root apices. The possible physiological relevance of this process is discussed.