Association of Candidate Gene Polymorphisms With Chronic Kidney Disease: Results of a Case-Control Analysis in the Nefrona Cohort

Chronic kidney disease (CKD) is a major risk factor for end-stage renal disease, cardiovascular disease and premature death. Despite classical clinical risk factors for CKD and some genetic risk factors have been identified, the residual risk observed in prediction models is still high. Therefore, new risk factors need to be identified in order to better predict the risk of CKD in the population. Here, we analyzed the genetic association of 79 SNPs of proteins associated with mineral metabolism disturbances with CKD in a cohort that includes 2, 445 CKD cases and 559 controls. Genotyping was performed with matrix assisted laser desorption ionizationtime of flight mass spectrometry. We used logistic regression models considering different genetic inheritance models to assess the association of the SNPs with the prevalence of CKD, adjusting for known risk factors. Eight SNPs (rs1126616, rs35068180, rs2238135, rs1800247, rs385564, rs4236, rs2248359, and rs1564858) were associated with CKD even after adjusting by sex, age and race. A model containing five of these SNPs (rs1126616, rs35068180, rs1800247, rs4236, and rs2248359), diabetes and hypertension showed better performance than models considering only clinical risk factors, significantly increasing the area under the curve of the model without polymorphisms. Furthermore, one of the SNPs (the rs2248359) showed an interaction with hypertension, being the risk genotype affecting only hypertensive patients. We conclude that 5 SNPs related to proteins implicated in mineral metabolism disturbances (Osteopontin, osteocalcin, matrix gla protein, matrix metalloprotease 3 and 24 hydroxylase) are associated to an increased risk of suffering CKD

A Family of Plasmodesmal Proteins with Receptor-Like Properties for Plant Viral Movement Proteins

Plasmodesmata (PD) are essential but poorly understood structures in plant cell walls that provide symplastic continuity and intercellular communication pathways between adjacent cells and thus play fundamental roles in development and pathogenesis. Viruses encode movement proteins (MPs) that modify these tightly regulated pores to facilitate their spread from cell to cell. The most striking of these modifications is observed for groups of viruses whose MPs form tubules that assemble in PDs and through which virions are transported to neighbouring cells. The nature of the molecular interactions between viral MPs and PD components and their role in viral movement has remained essentially unknown. Here, we show that the family of PD-located proteins (PDLPs) promotes the movement of viruses that use tubule-guided movement by interacting redundantly with tubule-forming MPs within PDs. Genetic disruption of this interaction leads to reduced tubule formation, delayed infection and attenuated symptoms. Our results implicate PDLPs as PD proteins with receptor-like properties involved the assembly of viral MPs into tubules to promote viral movement

Arabidopsis Plasmodesmal Proteome

The multicellular nature of plants requires that cells should communicate in order to coordinate essential functions. This is achieved in part by molecular flux through pores in the cell wall, called plasmodesmata. We describe the proteomic analysis of plasmodesmata purified from the walls of Arabidopsis suspension cells. Isolated plasmodesmata were seen as membrane-rich structures largely devoid of immunoreactive markers for the plasma membrane, endoplasmic reticulum and cytoplasmic components. Using nano-liquid chromatography and an Orbitrap ion-trap tandem mass spectrometer, 1341 proteins were identified. We refer to this list as the plasmodesmata- or PD-proteome. Relative to other cell wall proteomes, the PD-proteome is depleted in wall proteins and enriched for membrane proteins, but still has a significant number (35%) of putative cytoplasmic contaminants, probably reflecting the sensitivity of the proteomic detection system. To validate the PD-proteome we searched for known plasmodesmal proteins and used molecular and cell biological techniques to identify novel putative plasmodesmal proteins from a small subset of candidates. The PD-proteome contained known plasmodesmal proteins and some inferred plasmodesmal proteins, based upon sequence or functional homology with examples identified in different plant systems. Many of these had a membrane association reflecting the membranous nature of isolated structures. Exploiting this connection we analysed a sample of the abundant receptor-like class of membrane proteins and a small random selection of other membrane proteins for their ability to target plasmodesmata as fluorescently-tagged fusion proteins. From 15 candidates we identified three receptor-like kinases, a tetraspanin and a protein of unknown function as novel potential plasmodesmal proteins. Together with published work, these data suggest that the membranous elements in plasmodesmata may be rich in receptor-like functions, and they validate the content of the PD-proteome as a valuable resource for the further uncovering of the structure and function of plasmodesmata as key components in cell-to-cell communication in plants

Dissecting the proteome dynamics of the early heat stress response leading to plant survival or death in Arabidopsis

In many plant species, an exposure to a sublethal temperature triggers an adaptative response called acclimation. This response involves an extensive molecular reprogramming that allows the plant to further survive to an otherwise lethal increase of temperature. A related response is also launched under an abrupt and lethal heat stress that, in this case, is unable to successfully promote thermotolerance and therefore ends up in plant death. Although these molecular programmes are expected to have common players, the overlapping degree and the specific regulators of each process are currently unknown. We have carried out a high-throughput comparative proteomics analysis during acclimation and during the early stages of the plant response to a severe heat stress that lead Arabidopsis seedlings either to survival or death. This analysis dissects these responses, unravels the common players and identifies the specific proteins associated with these different fates. Thermotolerance assays of mutants in genes with an uncharacterized role in heat stress demonstrate the relevance of this study to uncover both positive and negative heat regulators and pinpoint a pivotal role of JR1 and BAG6 in heat tolerance.The research leading to these results has received funding from the European Research Council under the European Union's Seventh Framework Programme (FP/2007-2013)/ERC Grant Agreement 260468 to M. Mar Castellano and from the grants RTA2013-00027-00-00 and S2013/ABI-2734-CM. We thank Dr. A. Munoz for critical reading of the manuscript, Dr. R. Toribio for his help in the annotation of gene descriptions and Dr. Hara-Nishimura and Dr. Hans-Peter Mock for kindly providing the OLEO2 and HSP90.1 antibodies, respectively. Plant Response is also thanked for its support in the project. The authors declare no conflict of interest.S

Tobacco rattle virus 16K silencing suppressor binds AGO4 and inhibits formation of RNA silencing complexes

35 p.-5 fig.1 tab. supl.The cysteine-rich 16K protein of tobacco rattle virus (TRV), the type member of the genus Tobravirus, is known to suppress RNA silencing. However, the mechanism of action of the 16K suppressor is not well understood. In this study, we used a GFP-based sensor strategy and an Agrobacterium-mediated transient assay in Nicotiana benthamiana to show that 16K was unable to inhibit the activity of existing small interfering RNA (siRNA)- and microRNA (miRNA)-programmed RNA-induced silencing effector complexes (RISCs). In contrast, 16K efficiently interfered with de novo formation of miRNA- and siRNA-guided RISCs, thus preventing cleavage of target RNA. Interestingly, we found that transiently expressed endogenous miR399 and miR172 directed sequence-specific silencing of complementary sequences of viral origin. 16K failed to bind small RNAs, although it interacted with ARGONAUTE 4, as revealed by bimolecular fluorescence complementation and immunoprecipitation assays. Site-directed mutagenesis demonstrated that highly conserved cysteine residues within the N-terminal and central regions of the 16K protein are required for protein stability and/or RNA silencing suppression.This work was supported by grants BIO2006-13107 and BIO2009-12004 from the Spanish Ministry of Science and Innovation (MICINN-FEDER funding program) (CL), grants OTKA NN 107787 and NN 111024 from the Hungarian Scientific Research Fund and (LL), and grant AGL2008-03482 from MICINN (TC). LFC was the recipient of a JAE-Doc Contract from CSIC. IGB and IG were supported by graduate fellowships from MICINN. LL was supported from TÁMOP-4.2.2.A11/1/KONV-2012-0035.Peer reviewe