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921.
Poornima Sukumar Karin S. Edwards Abidur Rahman Alison DeLong Gloria K. Muday 《Plant physiology》2009,150(2):722-735
Reversible protein phosphorylation is a key regulatory mechanism governing polar auxin transport. We characterized the auxin transport and gravitropic phenotypes of the pinoid-9 (pid-9) mutant of Arabidopsis (Arabidopsis thaliana) and tested the hypothesis that phosphorylation mediated by PID kinase and dephosphorylation regulated by the ROOTS CURL IN NAPHTHYLPHTHALAMIC ACID1 (RCN1) protein might antagonistically regulate root auxin transport and gravity response. Basipetal indole-3-acetic acid transport and gravitropism are reduced in pid-9 seedlings, while acropetal transport and lateral root development are unchanged. Treatment of wild-type seedlings with the protein kinase inhibitor staurosporine phenocopies the reduced auxin transport and gravity response of pid-9, while pid-9 is resistant to inhibition by staurosporine. Staurosporine and the phosphatase inhibitor, cantharidin, delay the asymmetric expression of DR5∷revGFP (green fluorescent protein) at the root tip after gravistimulation. Gravity response defects of rcn1 and pid-9 are partially rescued by treatment with staurosporine and cantharidin, respectively. The pid-9 rcn1 double mutant has a more rapid gravitropic response than rcn1. These data are consistent with a reciprocal regulation of gravitropism by RCN1 and PID. Furthermore, the effect of staurosporine is lost in pinformed2 (pin2). Our data suggest that reduced PID kinase function inhibits gravitropism and basipetal indole-3-acetic acid transport. However, in contrast to PID overexpression studies, we observed wild-type asymmetric membrane distribution of the PIN2 protein in both pid-9 and wild-type root tips, although PIN2 accumulates in endomembrane structures in pid-9 roots. Similarly, staurosporine-treated plants expressing a PIN2∷GFP fusion exhibit endomembrane accumulation of PIN2∷GFP, but no changes in membrane asymmetries were detected. Our data suggest that PID plays a limited role in root development; loss of PID activity alters auxin transport and gravitropism without causing an obvious change in cellular polarity.A variety of important growth and developmental processes, including gravity response, embryo and vascular development, and the branching of roots and shoots, are controlled by the directional and regulated transport of auxin in higher plants. Reversible protein phosphorylation is an important regulatory strategy that may modulate auxin transport and dependent processes such as root gravitropism, perhaps through action of the PINOID (PID) kinase (for review, see DeLong et al., 2002; Galvan-Ampudia and Offringa, 2007). PID is an AGC family Ser/Thr kinase (Christensen et al., 2000) and belongs to an AGC kinase clade containing WAG1, WAG2, AGC3-4, and D6PK/AGC1-1 (Santner and Watson, 2006; Galvan-Ampudia and Offringa, 2007; Zourelidou et al., 2009). PID activity has been demonstrated in vitro and in vivo (Christensen et al., 2000; Michniewicz et al., 2007), and several pid mutant alleles exhibit altered auxin transport in the inflorescence and a floral development defect resembling that of auxin transport mutants (Bennett et al., 1995). Overexpression of the PID gene results in profound alterations in root development and responses to auxin transport inhibitors, reduced gravitropism and auxin accumulation at the root tip (Christensen et al., 2000; Benjamins et al., 2001; Michniewicz et al., 2007), as well as enhanced indole-3-acetic acid (IAA) efflux in tobacco (Nicotiana tabacum) cell cultures (Lee and Cho, 2006) and altered PINFORMED1 (PIN1), PIN2, and PIN4 localization patterns (Friml et al., 2004; Michniewicz et al., 2007), consistent with PID being a positive regulator of IAA efflux. However, the effects of pid loss-of-function mutations on auxin transport activities and gravitropic responses in roots have not yet been reported (Robert and Offringa, 2008).In contrast, auxin transport and gravitropism defects of a mutant with reduced protein phosphatase activity have been characterized in detail. The roots curl in naphthylphthalamic acid1 (rcn1) mutation, which ablates the function of a protein phosphatase 2A regulatory subunit, causes reduced PP2A activity in vivo and in vitro (Deruère et al., 1999). Roots and hypocotyls of rcn1 seedlings have elevated basipetal auxin transport (Deruère et al., 1999; Rashotte et al., 2001; Muday et al., 2006), and rcn1 roots exhibit a significant delay in gravitropism, consistent with altered auxin transport (Rashotte et al., 2001; Shin et al., 2005). These data indicate that PP2A is a negative regulator of basipetal transport and suggest that if PID-dependent phosphorylation regulates root auxin transport and gravitropism, then it may act in opposition to PP2A-dependent dephosphorylation.In roots, auxin transport is complex, with distinct sets of influx and efflux carriers that define tissue-specific and opposing directional polarities (for review, see Leyser, 2006). IAA moves acropetally, from the shoot toward the root apex, through the central cylinder (Tsurumi and Ohwaki, 1978), and basipetally, from the root apex toward the base, through the outer layer of cells (for review, see Muday and DeLong, 2001). When plants are reoriented relative to the gravity vector, auxin becomes asymmetrically distributed across the root tip, as a result of a process termed lateral auxin transport (for review, see Muday and Rahman, 2008). Several carriers that mediate root basipetal IAA transport have been clearly defined and include the influx carrier AUXIN-INSENSITIVE1 (AUX1; Marchant et al., 1999; Swarup et al., 2004; Yang et al., 2006) and efflux carriers of two classes, PIN2 (Chen et al., 1998; Müller et al., 1998; Rashotte et al., 2000) and ATP-BINDING CASSETTE TYPE B TRANSPORTER4/MULTIDRUG-RESISTANT4/P-GLYCOPROTEIN4 (ABCB4/MDR4/PGP4; Geisler et al., 2005; Terasaka et al., 2005; Lewis et al., 2007). Lateral transport at the root tip may be mediated by PIN3, an efflux carrier with a gravity-dependent localization pattern (Friml et al., 2002; Harrison and Masson, 2007).Gravitropic curvature of Arabidopsis (Arabidopsis thaliana) roots requires changes in IAA transport at the root tip (for review, see Muday and Rahman, 2008). Auxin transport inhibitors (Rashotte et al., 2000) and mutations in genes encoding basipetal transporters, including aux1 (Bennett et al., 1996), pin2/agr1 (Chen et al., 1998; Müller et al., 1998), and abcb4/mdr4/pgp4 (Lin and Wang, 2005; Lewis et al., 2007), alter gravitropism. Auxin-inducible reporters exhibit asymmetric expression across the root tip prior to differential growth, and this asymmetry is abolished by treatment with auxin transport inhibitors that prevent gravitropic curvature (Rashotte et al., 2001; Ottenschläger et al., 2003). Additionally, the pin3 mutant exhibits slightly reduced rates of gravitropic curvature (Harrison and Masson, 2007), and PIN3 is expressed in the columella cells, which are the site of gravity perception (Blancaflor et al., 1998; Friml et al., 2002). The PIN3 protein relocates to membranes on the lower side of columella cells after gravitropic reorientation, consistent with a role in facilitating asymmetric IAA transport at the root tip (Friml et al., 2002; Harrison and Masson, 2007).The available data suggest a model in which PID and RCN1 antagonistically regulate basipetal transport and gravitropic response in root tips (Fig. 1). In this model, the regions with the highest IAA concentrations in the epidermal and cortical cell layers are indicated by shading, and the arrows indicate the direction and relative amounts of basipetal auxin transport. Our previous work suggests that elevated basipetal IAA transport in rcn1 roots impairs gravitropic response, presumably due to the inability of roots either to form or to perceive a lateral auxin gradient in the context of a stronger polar IAA transport stream (Rashotte et al., 2001). Enhanced basipetal transport may increase the initial auxin concentration along the upper side of the root, impeding the establishment or perception of a gradient in rcn1 and cantharidin-treated wild-type roots (Fig. 1, right). Based on the published pid inflorescence transport data (Bennett et al., 1995), we hypothesize that pid seedling roots and staurosporine-treated wild-type roots have reduced basipetal auxin transport (Fig. 1, left). Upon reorientation of roots relative to the gravity vector, the reduced basipetal IAA transport in pid may lead to slower establishment of an auxin gradient across the root. This model then predicts that cantharidin treatment of pid-9 or staurosporine treatment of rcn1 seedlings would enhance or restore gravitropism in these mutants. Similarly, a double mutant might be expected to exhibit a corrected gravitropic response relative to the single mutants.Open in a separate windowFigure 1.Auxin transport defects in pid-9 and rcn1 mutants alter auxin redistribution after reorientation relative to the gravity vector. This model predicts that differences in basipetal auxin transport activities of wild-type, pid-9, and rcn1 roots will affect the formation of lateral auxin gradients. The shaded area in each root represents the region of highest IAA concentration in epidermal and cortical cells, with darker shading in the central columella cells, believed to be the auxin maxima. The direction and amount of basipetal IAA transport are indicated by arrows. The region of differential growth during gravitropic bending is indicated by the shaded rectangle. If auxin transport is reduced (as shown in the pid-9 mutant or in staurosporine-treated seedlings), this would lead to a slower formation of an auxin gradient in root tips. The rcn1 mutation (or treatment with cantharidin) has already been shown to lead to increased basipetal transport and a reduced rate of gravitropic bending, consistent with altered formation or perception of an auxin gradient. The antagonistic effects of kinase and phosphatase inhibition are predicted to lead to normal gravity responses in the pid-9 rcn1 double mutant as well as in pid-9 and rcn1 single mutants treated with the “reciprocal” inhibitor.The experiments described here were designed to test this model by examining gravitropism and root basipetal IAA transport in pid and staurosporine-treated seedlings. We investigated the regulation of gravity response by PID kinase and RCN1-dependent PP2A activities and observed antagonistic interactions between the rcn1 and pid-9 loss-of-function phenotypes that are consistent with reciprocal kinase/phosphatase regulation. We found that loss of kinase activity in the pid mutant and in staurosporine-treated wild-type plants inhibits basipetal auxin transport and the dependent physiological process of root gravitropism. Our results suggest that staurosporine acts to regulate these processes through inhibition of PID kinase and that PID effects are PIN2 dependent. In both wild-type and pid-9 roots, we observed polar membrane distribution of the PIN2 protein; unlike wild-type roots, though, pid-9 roots exhibited modest accumulation of PIN2 in endomembrane structures. Similarly, we detected asymmetric distribution and endomembrane accumulation of PIN2∷GFP in staurosporine-treated roots. Our data suggest that PID plays a limited role in root development; loss of PID activity alters PIN2 trafficking, auxin transport, and gravitropism without causing an obvious loss of cellular polarity. Together, these experiments provide insight into phosphorylation-mediated control of the gravity response and auxin transport in Arabidopsis roots. 相似文献
922.
Ilham Bouimetarhan Fabienne Marret Lydie Dupont Karin Zonneveld 《Marine Micropaleontology》2009,71(3-4):113-130
An organic-walled dinoflagellate cyst analysis was carried out on 53 surface sediment samples from West Africa (17–6°N) to obtain insight in the relationship between their spatial distribution and hydrological conditions in the upper water column as well as marine productivity in the study area.Multivariate analysis of the dinoflagellate cyst relative abundances and environmental parameters of the water column shows that sea-surface temperature, salinity, marine productivity and bottom water oxygen are the factors that relate significantly to the distribution patterns of individual species in the region.The composition of cyst assemblages and dinoflagellate cyst concentrations allows the identification of four hydrographic regimes; 1) the northern regime between 17 and 14°N characterized by high productivity associated with seasonal coastal upwelling, 2) the southern regime between 12 and 6°N associated with high-nutrient waters influenced by river discharge 3) the intermediate regime between 14 and 12°N influenced mainly by seasonal coastal upwelling additionally associated with fluvial input of terrestrial nutrients and 4) the offshore regime characterized by low chlorophyll-a concentrations in upper waters and high bottom water oxygen concentrations.Our data show that cysts of Polykrikos kofoidii, Selenopemphix quanta, Dubridinium spp., Echinidinium species, cysts of Protoperidinium monospinum and Spiniferites pachydermus are the best proxies to reconstruct the boundary between the NE trade winds and the monsoon winds in the subtropical eastern Atlantic Ocean. The association of Bitectatodinium spongium, Lejeunecysta oliva, Quinquecuspis concreta, Selenopemphix nephroides, Trinovantedinium applanatum can be used to reconstruct past river outflow variations within this region. 相似文献
923.
924.
Karin E. van Straaten Claudio F. Gonzalez Ricardo B. Valladares Xiaohui Xu Alexei V. Savchenko David A. R. Sanders 《Protein science : a publication of the Protein Society》2009,18(10):2196-2202
The structure of the Atu1476 protein from Agrobacterium tumefaciens was determined at 2 Å resolution. The crystal structure and biochemical characterization of this enzyme support the conclusion that this protein is an S-formylglutathione hydrolase (AtuSFGH). The three-dimensional structure of AtuSFGH contains the α/β hydrolase fold topology and exists as a homo-dimer. Contacts between the two monomers in the dimer are formed both by hydrogen bonds and salt bridges. Biochemical characterization reveals that AtuSFGH hydrolyzes C—O bonds with high affinity toward short to medium chain esters, unlike the other known SFGHs which have greater affinity toward shorter chained esters. A potential role for Cys54 in regulation of enzyme activity through S-glutathionylation is also proposed. 相似文献
925.
Through lysis of bacterioplankton cells, viruses mediate an important, but poorly understood, pathway of carbon and nutrients from the particulate to the dissolved form. Via this activity, nutrient-rich cell lysates may become available to noninfected cells and support significant growth. However, the nutritional value of lysates for noninfected bacteria presumably depends on the prevailing nutrient limitation. In the present study, we examined dynamics of dissolved DNA (D-DNA) and viruses along a transect in the phosphorus (P)-limited Ore Estuary, northern Baltic Sea. We found that viruses were an important mortality factor for bacterioplankton and that their activity mediated a significant recycling of carbon and especially of P. Uptake of dissolved DNA accounted for up to 70% of the bacterioplankton P demand, and about a quarter of the D-DNA pool was supplied through viral lysis of bacterial cells. Generally, the importance of viral lysates and uptake of D-DNA was highest at the estuarine and offshore stations and was positively correlated with P limitation measured as alkaline phosphatase activity. Our results highlight the importance of viral activity for the internal recycling of principal nutrients and pinpoints D-DNA as a particularly relevant compound in microbial P dynamics. 相似文献
926.
Gustafsson E Forsberg C Haraldsson K Lindman S Ljung L Furebring C 《Protein expression and purification》2009,63(2):95-101
The Chemotaxis Inhibitory Protein of Staphylococcus aureus (CHIPS) binds and blocks the C5a receptor (C5aR) and formyl-peptide receptor (FPR). This way, CHIPS is a potent inhibitor of the immune cell recruitment associated with inflammation. Truncation of the protein and the introduction of mutations, shifts the expression towards the insoluble fraction of Escherichia coli, whereas the wild-type protein can be solubly expressed. A protocol for expression and tag independent purification of biologically active CHIPS variants has been established to enable further characterization of an improved CHIPS variant, called ADC-1004. The CHIPS variants were purified by washing of E. coli inclusion bodies followed by refolding and gel filtration. New techniques were utilized to optimize the purification process. Expression in inclusion bodies was increased by the use of Ultra Yield™ flasks and optimal refolding conditions were determined by the use of the iFOLD Refolding System 2™.The folding and biological activity of the purified proteins were analyzed by circular dichroism (CD) spectroscopy and flow cytometry, respectively, and compared to solubly produced CHIPS31–113 and wild-type CHIPS1–121. We show that the CHIPS variants produced in inclusion bodies can be refolded and purified to achieve equal biological activity as solubly produced CHIPS31–113 and wild-type CHIPS1–121. The truncation causes minor structural changes while purification from inclusion bodies or the soluble fraction does not further affect the structure. 相似文献
927.
Glutamate receptors modulate sodium-dependent and calcium-independent vitamin C bidirectional transport in cultured avian retinal cells 总被引:1,自引:1,他引:0
Camila Cabral Portugal Vivian Sayuri Miya Karin da Costa Calaza Rochelle Alberto Martins Santos Roberto Paes-de-Carvalho 《Journal of neurochemistry》2009,108(2):507-520
Vitamin C is transported in the brain by sodium vitamin C co‐transporter 2 (SVCT‐2) for ascorbate and glucose transporters for dehydroascorbate. Here we have studied the expression of SVCT‐2 and the uptake and release of [14C] ascorbate in chick retinal cells. SVCT‐2 immunoreactivity was detected in rat and chick retina, specially in amacrine cells and in cells in the ganglion cell layer. Accordingly, SVCT‐2 was expressed in cultured retinal neurons, but not in glial cells. [14C] ascorbate uptake was saturable and inhibited by sulfinpyrazone or sodium‐free medium, but not by treatments that inhibit dehydroascorbate transport. Glutamate‐stimulated vitamin C release was not inhibited by the glutamate transport inhibitor l ‐β‐threo‐benzylaspartate, indicating that vitamin C release was not mediated by glutamate uptake. Also, ascorbate had no effect on [3H] d ‐aspartate release, ruling out a glutamate/ascorbate exchange mechanism. 2‐Carboxy‐3‐carboxymethyl‐4‐isopropenylpyrrolidine (Kainate) or NMDA stimulated the release, effects blocked by their respective antagonists 6,7‐initroquinoxaline‐2,3‐dione (DNQX) or (5R,2S)‐(1)‐5‐methyl‐10,11‐dihydro‐5H‐dibenzo[a,d]cyclohepten‐5,10‐imine hydrogen maleate (MK‐801). However, DNQX, but not MK‐801 or 2‐amino‐5‐phosphonopentanoic acid (APV), blocked the stimulation by glutamate. Interestingly, DNQX prevented the stimulation by NMDA, suggesting that the effect of NMDA was mediated by glutamate release and stimulation of non‐NMDA receptors. The effect of glutamate was neither dependent on external calcium nor inhibited by 1,2‐bis (2‐aminophenoxy) ethane‐N′,N′,N′,N′,‐tetraacetic acid tetrakis (acetoxy‐methyl ester) (BAPTA‐AM), an internal calcium chelator, but was inhibited by sulfinpyrazone or by the absence of sodium. In conclusion, retinal cells take up and release vitamin C, probably through SVCT‐2, and the release can be stimulated by NMDA or non‐NMDA glutamate receptors. 相似文献
928.
929.
The bioavailability of chromium from Cr-picolinate (CrPic3) and Cr-chloride (CrCl3) was studied in rats using 51Cr-labelled compounds and whole-body-counting. The intestinal absorption of Cr was twice as high from CrPic3 (1.16% vs 0.55%) than from CrCl3, however most of the absorbed 51Cr from CrPic3 was excreted into the urine within 24 h. After i.v. or i.p. injection, the whole-body retention curves fitted well to a multiexponential
function, demonstrating that plasma chromium is in equilibrium with three pools. For CrPic3, a large pool exists with a very rapid exchange (T
1/2 = <0.5 days), suggesting that CrPic3 is absorbed as intact molecule, from which the main part is directly excreted by the kidney before degradation of the chromium
complex in the liver can occur. CrCl3 is less well absorbed but the rapid exchange pool is much smaller, resulting in even higher Cr concentrations in tissue such
as muscle and fat. However, 1–3 days after application, the relative distribution of 51Cr from both compounds was similar in all tissues studied, indicating that both compounds contribute to the same storage pool.
In summary, the bioavailability of CrPic3 in rats is not superior compared to CrCl3. 相似文献
930.
Pablo Sabat Sandra Gonzalez-Vejares Karin Maldonado 《Comparative biochemistry and physiology. Part A, Molecular & integrative physiology》2009,152(3):322-326
The urine field osmolality in Zonotrichia capensis along a latitudinal gradient in rainfall and temperature in Chile was examined. We also investigated latitudinal variation in the renal traits that mediate how these birds cope with dehydration. We used the δ15N of this species' tissue to investigate whether the reliance on animals and seeds varied among birds and if it had any effect on excretion and renal traits. We found a significant latitudinal variation in urine osmolality, a variable that was correlated with habitat aridity. We also found that the kidney size and proportion of kidney devoted to medullary tissue differed between birds from arid and mesic localities, but not in a lineal fashion with aridity. The increment in the position in the food web, as measured by δ15N, led to an increment in urine osmolality, without changes in kidney features. Our data suggested that differences in dietary habits in the field could be not extended enough to cause changes in the kidney structure in Rufous-collared sparrows. 相似文献