Multigene families encoding class XI myosins are conserved in higher plants, however, little information is available on specific functions of these ubiquitous molecular motors. We isolated gene knockout mutants for all 13 class XI myosins present in Arabidopsis (Arabidopsis thaliana) genome. Inactivation of 11 myosin genes resulted in no discernible phenotypes under the normal growth conditions. In contrast, the knockouts of the remaining two myosin genes, XI-2 (formerly MYA2) and XI-K, exhibited similar defects in root hair elongation suggesting that the myosin-driven motility plays a significant role in a polar tip growth. Strikingly, inactivation of each of these myosins also reduced trafficking of Golgi stacks, peroxisomes, and mitochondria in root hairs and in leaf epidermal cells. These results indicate that myosins XI-K and XI-2 play major and overlapping roles in the cell dynamics in Arabidopsis and highlight the redundant nature of myosin function in plants.Myosins are signature molecular motors of eukaryotes that are involved in a broad spectrum of actin cytoskeleton-associated types of cellular dynamics (Vale, 2003). Comparative genomics revealed that myosins are conserved throughout the eukaryotic domain of life (Richards and Cavalier-Smith, 2005;Foth et al., 2006). Land plants possess two myosin classes, XI and VIII, each of which is evolutionary, related to animal and fungal class V myosins (Desnos et al., 2007), suggesting their origin was from a common ancestor that antedated the divergence of Plantae and Opisthokonts (Foth et al., 2006). Subsequent evolution of myosins was dominated by gene duplication and diversification that resulted in the presence of more than 10 myosin genes in all plant genomes sequenced so far (see accompanying article Avisar et al., 2008). In particular, Arabidopsis (Arabidopsis thaliana) encodes 13 class XI and four class VIII myosins (Reddy and Day, 2001). The studies that mostly involved cytoskeletal inhibitors have demonstrated the principal role of actomyosin motilty in plant cell dynamics including organelle trafficking, remodeling, and inheritance (Boevink et al., 1998;Nebenfuhr et al., 1999;Sheahan et al., 2004;Kim et al., 2005;Runions et al., 2006). Some of the class XI myosins were found in association with organelles suggesting their involvement in organelle transport (Wang and Pesacreta, 2004;Hashimoto et al., 2005;Li and Nebenfuhr, 2007;Reisen and Hanson, 2007). However, information on functional profiles of individual myosin motors is very limited. Two recent publications have demonstrated the role of Arabidopsis myosin XI-K in root hair growth (Ojangu et al., 2007), and implicated rice (Oryza sativa) myosin XI-B in pollen development (Jiang et al., 2007).Here we screen the gene knockouts of all 13 class XI myosins of Arabidopsis to show that, in addition to myosin XI-K, myosin XI-2 (MYA2) is also required for root hair development. Furthermore, we demonstrate that each of these two highly expressed myosins functions in the rapid movement of the Golgi stac...

A prominent feature of plant cells is the rapid, incessant movement of the organelles traditionally defined as cytoplasmic streaming and attributed to actomyosin motility. We sequenced six complete Nicotiana benthamiana cDNAs that encode class XI and class VIII myosins. Phylogenetic analysis indicates that these two classes of myosins diverged prior to the radiation of green algae and land plants from a common ancestor and that the common ancestor of land plants likely possessed at least seven myosins. We further report here that movement of Golgi stacks, mitochondria, and peroxisomes in the leaf cells of N. benthamiana is mediated mainly by myosin XI-K. Suppression of myosin XI-K function using dominant negative inhibition or RNA interference dramatically reduced movement of each of these organelles. When similar approaches were used to inhibit functions of myosin XI-2 or XI-F, only moderate to marginal effects were observed. Organelle trafficking was virtually unaffected in response to inhibition of each of the three class VIII myosins. Interestingly, none of the tested six myosins appears to be involved in light-induced movements of chloroplasts. Taken together, these data strongly suggest that myosin XI-K has a major role in trafficking of Golgi stacks, mitochondria, and peroxisomes, whereas myosins XI-2 and XI-F might perform accessory functions in this process. In addition, our analysis of thousands of individual organelles revealed independent movement patterns for Golgi stacks, mitochondria, and peroxisomes, indicating that the notion of coordinated cytoplasmic streaming is not generally applicable to higher plants.

Gene families with multiple members are predicted to have individuals with overlapping functions. We examined all of the Arabidopsis (Arabidopsis thaliana) myosin family members for their involvement in Golgi and other organelle motility. Truncated fragments of all 17 annotated Arabidopsis myosins containing either the IQ tail or tail domains only were fused to fluorescent markers and coexpressed with a Golgi marker in two different plants. We tracked and calculated Golgi body displacement rate in the presence of all myosin truncations and found that tail fragments of myosins MYA1, MYA2, XI-C, XI-E, XI-I, and XI-K were the best inhibitors of Golgi body movement in the two plants. Tail fragments of myosins XI-B, XI-F, XI-H, and ATM1 had an inhibitory effect on Golgi bodies only in Nicotiana tabacum, while tail fragments of myosins XI-G and ATM2 had a slight effect on Golgi body motility only in Nicotiana benthamiana. The best myosin inhibitors of Golgi body motility were able to arrest mitochondrial movement too. No exclusive colocalization was found between these myosins and Golgi bodies in our system, although the excess of cytosolic signal observed could mask myosin molecules bound to the surface of the organelle. From the preserved actin filaments found in the presence of enhanced green fluorescent protein fusions of truncated myosins and the motility of myosin punctae, we conclude that global arrest of actomyosin-derived cytoplasmic streaming had not occurred. Taken together, our data suggest that the above myosins are involved, directly or indirectly, in the movement of Golgi and mitochondria in plant cells.The Arabidopsis (Arabidopsis thaliana) myosin gene family contains 17 members: myosin group XI, which includes 13 members (myosins XI-

The Hsp70 homolog (Hsp70h) of Beet yellows virus (BYV) functions in virion assembly and cell-to-cell movement and is autonomously targeted to plasmodesmata in association with the actomyosin motility system (A. I. Prokhnevsky, V. V. Peremyslov, and V. V. Dolja, J. Virol. 79:14421-14428, 2005). Myosins are a diverse category of molecular motors that possess a motor domain and a tail domain involved in cargo binding. Plants have two classes of myosins, VIII and XI, whose specific functions are poorly understood. We used dominant negative inhibition to identify myosins required for Hsp70h localization to plasmodesmata. Six full-length myosin cDNAs from the BYV host plant Nicotiana benthamiana were sequenced and shown to encode apparent orthologs of the Arabidopsis thaliana myosins VIII-1, VIII-2, VIII-B, XI-2, XI-F, and XI-K. We found that the ectopic expression of the tail domains of each of the class VIII, but not the class XI, myosins inhibited the plasmodesmatal localization of Hsp70h. In contrast, the overexpression of the motor domains or the entire molecules of the class VIII myosins did not affect Hsp70h targeting. Further mapping revealed that the minimal cargo-binding part of the myosin VIII tails was both essential and sufficient for the inhibition of the proper Hsp70h localization. Interestingly, plasmodesmatal localization of the Tobacco mosaic virus movement protein and Arabidopsis protein RGP2 was not affected by myosin VIII tail overexpression. Collectively, our data implicate class VIII myosins in protein delivery to plasmodesmata and suggest that more than one mechanism of such delivery exist in plants.

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The current study investigated the sorption of sulfadimethoxine (SMT), sulfamethoxazole (SMX), tetracycline (TET), and oxytetracycline (OTC) to Narich montmorillonite clay in synthetic effluent (SE) and field wastewater effluent (FE). Both SMT and SMX showed a low sorption capacity and are therefore likely to be highly mobile in the environment, while the sorption of TET to clay in environmental pH range (6.5-7.5) showed similarly high adsorption capacity. Differences in sorption capacities of TET and OTC to SE or FE were attributed to the various concentrations of divalent cations in the effluents. In addition, differences in sorption of OTC or TET to SE were attributed to their different molecular structure. Moreover, the adsorption of TET in SE and FE showed linear adsorption isotherms and fitted to Freundlich model. Further experiments showed that addition of humic acid or SE to TET sorbed to clay did not enhance or suppress the sorption of TET to clay.

It has recently been found that among the 17 Arabidopsis myosins, six (XIC, XIE, XIK, XI-I, MYA1, and MYA2) have a major role in the motility of Golgi bodies and mitochondria in Nicotiana benthamiana and Nicotiana tabacum. Here, the same dominant negative tail fragments were also found to arrest the movement of Gogi bodies when transiently expressed in Arabidopsis plants. However, when a Golgi marker was transiently expressed in plants knocked out in these myosins, its movement was dramatically inhibited only in the xik mutant. In addition, a tail fragment of myosin XIK could inhibit the movement of several post-Golgi organelles, such as the trans-Golgi network, pre-vacuolar compartment, and endosomes, as well as total cytoplasmic streaming, suggesting that myosin XIK is a major player in cytoplasm kinetics. However, no co-localization of myosin tails with the arrested organelles was observed. Several deletion truncations of the myosin XIK tail were generated to corroborate function with localization. All deletion mutants possessing an inhibitory effect on organelle movement exhibited a diffuse cytoplasmic distribution. Point mutations in the tail of myosin XIK revealed that Arg1368 and Arg1443 are essential for its activity. These residues correspond to Lys1706 and Lys1779 from mouse myosin Va, which mediate the inhibitory head–tail interaction in this myosin. Therefore, such an interaction might underlie the dominant negative effect of truncated plant myosin tails and explain the mislocalization with target organelles.

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