Environmental Regulation of Heterosis in the Allopolyploid Arabidopsis suecica.

Allopolyploids are organisms possessing more than two complete sets of chromosomes from two or more species and are frequently more vigorous than their progenitors. To address the question why allopolyploids display hybrid vigor, we compared the natural allopolyploid Arabidopsis suecica to its progenitor species Arabidopsis thaliana and Arabidopsis arenosa. We measured chlorophyll content, CO2 assimilation, and carbohydrate production under varying light conditions and found that the allopolyploid assimilates more CO2 per unit chlorophyll than either of the two progenitor species in high intensity light. The increased carbon assimilation corresponds with greater starch accumulation, but only in strong light, suggesting that the strength of hybrid vigor is dependent on environmental conditions. In weaker light A. suecica tends to produce as much primary metabolites as the better progenitor. We found that gene expression of LIMIT DEXTRINASE1, a debranching enzyme that cleaves branch points within starch molecules, is at the same level in the allopolyploid as in the maternal progenitor A. thaliana and significantly more expressed than in the paternal progenitor A. arenosa. However, expression differences of ß-amylases and GLUCAN-WATER DIKINASE1 were not statistically significantly elevated in the allopolyploid over progenitor expression levels. In contrast to allopolyploids, autopolyploid A. thaliana showed the same photosynthetic rate as diploids, indicating that polyploidization alone is likely not the reason for enhanced vigor in the allopolyploid. Taken together, our data suggest that the magnitude of heterosis in A. suecica is environmentally regulated, arises from more efficient photosynthesis, and, under specific conditions, leads to greater starch accumulation than in its progenitor species.

Synergistic interaction of CLAVATA1, CLAVATA2, and RECEPTOR-LIKE PROTEIN KINASE 2 in cyst nematode parasitism of Arabidopsis.

Plant-parasitic cyst nematodes secrete CLAVATA3 (CLV3)/ENDOSPERM SURROUNDING REGION (CLE)-like effector proteins. These proteins act as ligand mimics of plant CLE peptides and are required for successful nematode infection. Previously, we showed that the CLV2/CORYNE (CRN) heterodimer receptor complex is required for nematode CLE signaling. However, there was only a partial reduction in nematode infection when this signaling was disrupted, indicating that there might be additional nematode CLE receptors. In this study, we demonstrate that CLV1 and RECEPTOR-LIKE PROTEIN KINASE 2/TOADSTOOL2 (RPK2), two additional receptors that can transmit the CLV3 signal independent of CLV2/CRN for shoot apical meristem maintenance, also play a role in nematode CLE perception. Localization studies showed that both receptors are expressed in nematode-induced syncytia. Infection assays with clv1 and rpk2 single mutants revealed a decrease in both nematode infection and syncytium size. Significantly, further reduction in nematode infection was observed when rpk2 was combined with clv1 and clv2 mutants. Taken together, our results indicate that parallel signaling pathways involving CLV1, CLV2, and RPK2 are important for nematode parasitism.

Nematode CLE signaling in Arabidopsis requires CLAVATA2 and CORYNE.

Plant-parasitic cyst nematodes secrete CLAVATA3 (CLV3)/ESR (CLE)-like effector proteins. These proteins have been shown to act as ligand mimics of plant CLE peptides and are required for successful nematode infection; however, the receptors for nematode CLE-like peptides have not been identified. Here we demonstrate that CLV2 and CORYNE (CRN), members of the receptor kinase family, are required for nematode CLE signaling. Exogenous peptide assays and overexpression of nematode CLEs in Arabidopsis demonstrated that CLV2 and CRN are required for perception of nematode CLEs. In addition, promoter-reporter assays showed that both receptors are expressed in nematode-induced syncytia. Lastly, infection assays with receptor mutants revealed a decrease in both nematode infection and syncytium size. Taken together, our results indicate that perception of nematode CLEs by CLV2 and CRN is not only required for successful nematode infection but is also involved in the formation and/or maintenance of nematode-induced syncytia.

Unique N-Terminal Interactions Connect F-BOX STRESS INDUCED (FBS) Proteins to a WD40 Repeat-like Protein Pathway in Arabidopsis.

SCF-type E3 ubiquitin ligases provide specificity to numerous selective protein degradation events in plants, including those that enable survival under environmental stress. SCF complexes use F-box (FBX) proteins as interchangeable substrate adaptors to recruit protein targets for ubiquitylation. FBX proteins almost universally have structure with two domains: A conserved N-terminal F-box domain interacts with a SKP protein and connects the FBX protein to the core SCF complex, while a C-terminal domain interacts with the protein target and facilitates recruitment. The F-BOX STRESS INDUCED (FBS) subfamily of plant FBX proteins has an atypical structure, however, with a centrally located F-box domain and additional conserved regions at both the N- and C-termini. FBS proteins have been linked to environmental stress networks, but no ubiquitylation target(s) or biological function has been established for this subfamily. We have identified two WD40 repeat-like proteins in Arabidopsis that are highly conserved in plants and interact with FBS proteins, which we have named FBS INTERACTING PROTEINs (FBIPs). FBIPs interact exclusively with the N-terminus of FBS proteins, and this interaction occurs in the nucleus. FBS1 destabilizes FBIP1, consistent with FBIPs being ubiquitylation targets SCFFBS1 complexes. This work indicates that FBS proteins may function in stress-responsive nuclear events, and it identifies two WD40 repeat-like proteins as new tools with which to probe how an atypical SCF complex, SCFFBS, functions via FBX protein N-terminal interaction events.

Dual roles for the variable domain in protein trafficking and host-specific recognition of Heterodera glycines CLE effector proteins.

*Soybean cyst nematodes (Heterodera glycines) produce secreted effector proteins that function as peptide mimics of plant CLAVATA3/ESR (CLE)-like peptides probably involved in the developmental reprogramming of root cells to form specialized feeding cells called syncytia. *The site of action and mechanism of delivery of CLE effectors to host plant cells by the nematode, however, have not been established. In this study, immunologic, genetic and biochemical approaches were used to reveal the localization and site of action of H. glycines-secreted CLE proteins in planta. *We present evidence indicating that the nematode CLE propeptides are delivered to the cytoplasm of syncytial cells, but ultimately function in the apoplast, consistent with their proposed role as ligand mimics of plant CLE peptides. We determined that the nematode 12-amino-acid CLE motif peptide is not sufficient for biological activity in vivo, pointing to an important role for sequences upstream of the CLE motif in function. *Genetic and biochemical analysis confirmed the requirement of the variable domain in planta for host-specific recognition and revealed a novel role in trafficking cytoplasmically delivered CLEs to the apoplast in order to function as ligand mimics.

A Novel Phagomyxid Parasite Produces Sporangia in Root Hair Galls of Eelgrass (Zostera marina).

The objective of this study was to identify the parasite causing the formation of root hair galls on eelgrass (Zostera marina) in Puget Sound, WA. Microscopic and molecular analyses revealed that a novel protist formed plasmodia that developed into sporangia in root hair tip galls and released biflagellate swimming zoospores. Root hair galls were also observed in the basal section of root hairs, and contained plasmodia or formed thick-walled structures filled with cells (resting spores). Phylogenetic analyses of 18S rDNA sequence data obtained from cells in sporangia indicated that the closest relative of the parasite with a known taxonomic identification was Plasmodiophora diplantherae (86.9% sequence similarity), a phagomyxid parasite that infects the seagrass Halodule spp. To determine the local geographic distribution of the parasite, root and soil samples were taken from four eelgrass populations in Puget Sound and analyzed for root hair galls and parasite DNA using a newly designed qPCR protocol. The percent of root hairs with galls and amount of parasite DNA in roots and sediment varied among the four eelgrass populations. Future studies are needed to establish the taxonomy of the parasite, its effects on Z. marina, and the factors that determine its distribution and abundance.

Can consumers self-select for appropriate use of an over-the-counter statin? The Self Evaluation of Lovastatin to Enhance Cholesterol Treatment Study.

Access to over-the-counter (OTC) statins has the potential to improve public health by reducing cardiovascular events. The Self Evaluation of Lovastatin to Enhance Cholesterol Treatment (SELECT) Study was designed to assess consumers' ability to self-select for treatment with lovastatin in an unsupervised setting. Subjects examined proposed OTC lovastatin cartons with labels that detailed an algorithm for self-selection based on age, lipid profile, and cardiovascular risk factors. Subjects viewed a carton with either a low-density lipoprotein cholesterol-based self-selection algorithm or one based on total cholesterol. Labels also contained warnings against use based on health conditions that might increase the risk of adverse events. Subjects were asked if the drug was appropriate for their use (self-assessment) and whether they would like to purchase the drug (purchase decision). A total of 1,326 consumers provided self-assessment decisions. After viewing the low-density lipoprotein cholesterol-based label, 82%, 36%, and 82% of those who self-assessed that the drug was appropriate for their use were correct with respect to the age, lipid, and risk-factor criteria, respectively. Corresponding numbers for the total cholesterol algorithm were 85%, 50% and 75%. Almost 90% of women aged <55 years who evaluated the drug indicated the drug was not right for them, and women in this age group made up only 9% of the total group of subjects who believed the drug was appropriate for their use. The label was also effective in discouraging use by women who were or may become pregnant, consumers with liver disease, and those with potential drug interactions. In conclusion, SELECT showed that consumers could use an OTC drug label in an unsupervised setting to appropriately self-select for self-management of their cholesterol with lovastatin.

Hepatic effects of lovastatin exposure in patients with liver disease: a retrospective cohort study.

BACKGROUND: Little is known about the potential adverse hepatic effects of HMG-CoA reductase inhibitors ('statins') in patients with existing liver disease; therefore, we examined the risk of liver toxicity with lovastatin exposure in these patients. METHODS: A retrospective cohort study was performed using data from a large integrated health plan in Northern California, USA. Patients with laboratory or clinical evidence of liver disease were identified and their exposure to lovastatin was determined. The primary outcome was a pattern of liver-test abnormalities associated with a poor prognosis among patients with drug-induced liver disease, based on Hy's Rule. Secondary outcomes included liver injury (defined as moderate or severe, depending on the degree of ALT level elevations) or the development of either clinical cirrhosis or liver failure. Incidence rate ratios (IRRs) were calculated and multivariate analyses conducted using extended Cox models. RESULTS: A total of 93 106 patients met the entry criteria. Lovastatin exposure was associated with a lower incidence of all endpoints, including the primary outcome (IRR = 0.28, 95% CI 0.12, 0.55), moderate liver injury (IRR = 0.56, 95% CI 0.47, 0.65), severe liver injury (IRR = 0.50, 95% CI 0.29, 0.81) and the occurrence of either cirrhosis or liver failure (IRR = 0.29, 95% CI 0.21, 0.38); adjustment for age and sex resulted in some attenuation of this reduction in incidence. The observed effects were generally consistent across a range of baseline liver-disease diagnoses and greater cumulative lovastatin exposure was associated with fewer outcome events for some endpoints. CONCLUSIONS: In this retrospective analysis, exposure to lovastatin was not associated with an increased risk of adverse hepatic outcomes. These results do not support concern regarding lovastatin-related hepatotoxicity in patients with existing liver disease.

Identification of potential host plant mimics of CLAVATA3/ESR (CLE)-like peptides from the plant-parasitic nematode Heterodera schachtii.

In this article, we present the cloning of two CLAVATA3/ESR (CLE)-like genes, HsCLE1 and HsCLE2, from the beet cyst nematode Heterodera schachtii, a plant-parasitic cyst nematode with a relatively broad host range that includes the model plant Arabidopsis. CLEs are small secreted peptide ligands that play important roles in plant growth and development. By secreting peptide mimics of plant CLEs, the nematode can developmentally reprogramme root cells for the formation of unique feeding sites within host roots for its own benefit. Both HsCLE1 and HsCLE2 encode small secreted polypeptides with a conserved C-terminal CLE domain sharing highest similarity to Arabidopsis CLEs 1-7. Moreover, HsCLE2 contains a 12-amino-acid CLE motif that is identical to AtCLE5 and AtCLE6. Like all other plant and nematode CLEs identified to date, HsCLEs caused wuschel-like phenotypes when overexpressed in Arabidopsis, and this activity was abolished when the proteins were expressed without the CLE motif. HsCLEs could also function in planta without a signal peptide, highlighting the unique, yet conserved function of nematode CLE variable domains in trafficking CLE peptides for secretion. In a direct comparison of HsCLE2 overexpression phenotypes with those of AtCLE5 and AtCLE6, similar shoot and root phenotypes were observed. Exogenous application of 12-amino-acid synthetic peptides corresponding to the CLE motifs of HsCLEs and AtCLE5/6 suggests that the function of this class of CLEs may be subject to complex endogenous regulation. When seedlings were grown on high concentrations of peptide (10 µm), root growth was suppressed; however, when seedlings were grown on low concentrations of peptide (0.1 µm), root growth was stimulated. Together, these findings indicate that AtCLEs1-7 may be the target peptides mimicked by HsCLEs to promote parasitism.

The tobacco Cel7 gene promoter is auxin-responsive and locally induced in nematode feeding sites of heterologous plants.

SUMMARY: Emerging evidence suggests that plant cell-wall-modifying enzymes induced by root-parasitic nematodes play important roles in feeding cell formation. We previously identified a tobacco endo-beta-1,4-glucanase (cellulase) gene, NtCel7, that was strongly induced in both root-knot and cyst nematode feeding cells. To characterize further the developmental and nematode-responsive regulation of NtCel7, we isolated the NtCel7 promoter and analysed its expression over a time course of nematode infection and in response to auxin, gibberellin, ethylene and sucrose in soybean and tomato hairy roots and in Arabidopsis containing the NtCel7 promoter fused to the beta-glucuronidase (GUS) reporter gene. Histochemical analyses of transgenic plant materials revealed that the NtCel7 promoter exhibited a unique organ-specific expression pattern during plant development suggestive of important roles for NtCel7 in both vegetative and reproductive growth. In all plant species tested, strong GUS expression was observed in root tips and lateral root primordia of uninfected roots with weaker expression in the root vasculature. Further analyses of transgenic Arabidopsis plants revealed expression in shoot and root meristems and the vasculature of most organs during plant development. We also determined that the NtCel7 promoter was induced by auxin, but not gibberellin, ethylene or sucrose. Moreover, strong GUS activity was observed in both cyst and root-knot nematode-induced feeding sites in transgenic roots of soybean, tomato and Arabidopsis. The conserved developmental and nematode-responsive expression of the NtCel7 promoter in heterologous plants indicates that motifs of this regulatory element play a fundamental role in regulating NtCel7 gene expression within nematode feeding sites and that this regulation may be mediated by auxin.