Dynamics of agricultural expansion in Uruguayan territories

Uruguay experimenta desde hace dos décadas un proceso de transformaciones agrarias basado en la expansión de la forestación primero y de la agricultura de secano después, que impacta fuertemente sobre la estructura productiva, los actores de las cadenas productivas, las formas de organización de la producción, el trabajo, los territorios y el paisaje rural. La faceta más evidente de este proceso de transformaciones involucra el uso de la tierra y es claramente visible a través de la expansión de plantaciones en territorios con tradiciones productivas diferentes. Los cambios se desarrollan en el marco del avance del agronegocio, entendido como un modelo productivo que despliega una nueva agricultura con lógicas diferentes a las del empresariado agricola clásico, menos vinculada con actores y territorios locales y con un mayor grado de conexión con redes globales. El capital transnacional que invierte en la agricultura uruguaya reconfigura el paisaje, redefiniendo la identidad local del territorio y las relaciones de éste con quienes lo habitan y lo producen. Este articulo se propone poner en diálogo el análisis de los fenómenos sociales que emergen por el avance del modelo del agronegocio con los procesos especificos que ocurren a nivel territorial a partir de múltiples evidencias empiricas. Con base en el análisis de imágenes satelitales, recorridas de campo y entrevistas, se concentra en analizar las nuevas configuraciones de la agricultura de secano en territorios uruguayos, para dimensionar y comprender los procesos sociales desencadenados por el despliegue del agronegocio a partir de la década del 2000.For two decades, Uruguay has experienced a process of agrarian transformations based on forest expansion first and rainfed agriculture later, with strong impacts on the productive structure, the actors in the productive chains, the organizational forms of production and labor, territories and rural landscapes. The most evident side of this transformation process involves land use and is clearly visible through the expansion of crops in territories with different productive traditions. These changes are taking place together with the advance of agribusiness, understood as a productive model that deploys a new agriculture with different logics from those of the classic agricultural entrepreneurship, less linked to local actors and territories and with a greater degree of connection with global networks. By investing in Uruguayan agriculture, transnational capital reconfigures the landscape, redefining the local identity of territories and the relationships with those who inhabit and produce them. This article puts into dialogue the analysis of the social phenomena that emerge from the advance of the agribusiness model with the specific processes that occur at the territorial level based on multiple empirical evidence. Based on the analysis of satellite images, field trips and interviews, it focuses on analyzing the new configurations of rainfed agriculture in Uruguayan territories, in order to determine the dimension and understand the social processes triggered by the deployment of the agribusiness model since the 2000s

Characterization of Rpn10 monoubiquitination as a novel way of proteasome regulation

[eng] Targeted protein degradation plays a central role in eukaryotic cell regulation and homeostasis. The ubiquitin proteasome system (UPS) is involved in a large number of cellular events, being, day by day, more difficult to find pathways without links with the system. Proteins are marked for degradation by ubiquitin ligases that append polyubiquitin signals, which can recruit the targeted protein to the 26S proteasome for degradation. In the process, polyubiquitin chains appended to the target protein are disassembled into free monoubiquitin for subsequent rounds of substrate tagging by means of deubiquitinating enzymes. Rpn10 is a proteasome receptor that recognizes the Lys48-linked polyubiquitin degradation signal by means of a ubiquitin-interacting motif (UIM). Mutation of the UIM domain of Rpn10 significantly ablates the proteolytic capacity of the proteasome. Rpn10 also contains a Von Willebrand factor A (VWA) domain which is responsible of Rpn10 interactions inside the proteasome. Rpn10 is in equilibrium with a pool of free protein so that it functions as both, a receptor at the proteasome, and an extraproteasomal adaptor. In the present work, we have shown that Rpn10 is monoubiquitinated (Rpn10-mUb) in vivo and that Rpn10-mUb is found in both proteasomal and non-proteasomal pools. Levels of Rpn10-mUb are regulated in vivo by Rps5, a NEDD4 ubiquitin-ligase protein family, and Ubp2, a deubiquitinating enzyme. Our observations link for first time monoubiquitin signal with proteasome regulation. Monoubiquitination strongly inhibits Rpn10 to interact with ubiquitin conjugates by a specific interaction in cis between Rpn10 UIM domain and the linked monoubiquitin. By means of genetic and proteomics tools we found that four sites of Rpn10 sequence are subjected to monoubiquitination by Rsp5 (Lys71, Lys84, Lys99 and Lys268), but Lys84 within its VWA domain is the preferred one. Ubiquitination of Rpn10 inhibits its ability to bind polyubiquitinated substrates, thus functioning as mechanism of inactivation of this receptor. Interestingly, our findings suggest that Rpn10 monoubiquitination could decrease proteasome activity. The UIM of Rpn10 also strongly interacts with the ubiquitin-like domain (Ubl) of Dsk2, a polyubiquitin-binding protein. Furthermore, extraproteasomal Rpn10 plays a critical role in filtering Dsk2 and its substrates from the proteasome. We evaluated the affinity of several forms of Rpn10-mUb to Dsk2 and found that Ub-UIM interaction in cis of Rpn10-mUb impairs the binding of Dsk2 in trans. These results suggest that in an extraproteasomal context, Rpn10 monoubiquitination could regulate the Dsk2-Rpn10 interaction. Notably, the proteasomal pool of Rpn10-mUb is suppressed under perturbations that promote the proteolytic pathway, such as stress by temperature and oxidative stress, whereas the extraproteasomal pool remains fairly constant. To assess whether the distinct behavior of the Rpn10-mUb pools correlated with the modification of different lysine residues of Rpn10, we set up assays of absolute quantification of the two major ubiquitination sites, Lys84 and Lys268, by mass spectrometry (AQUA). The obtained results clearly indicate that Lys84 is the main target of Rsp5 ligase in both pools of Rpn10, enhancing the role of the VWA domain in Rpn10 ubiquitination. Finally, we have combined ubiquitin remnant profiling with quantitative proteomic approaches to identify ubiquitinated species that are increased in RPN10-RAD23 deletion, under standard growth conditions and cold-shock stress. Enriched conjugates have been distributed across different biological functions being proteins involved in RNA processing and transport metabolism accounting for the biggest fractions. Further molecular biology approaches and informatic analysis are required to address which proteins are likely to be true UPS substrates as opposed to proteins regulated by ubiquitination in a non-proteolytic manner. Overall, our results provide a novel evidence of involvement of monoubiquitin signals in the regulation of the availability of substrates to bind proteasomal surfaces. Considering that substrate binding is the first event in the multicatalytic process promoted by the proteasome, the control of the accessibility of proteasome receptors is a crucial level of proteasome regulation. Thus, if the NEDD4 enzyme catalyzed monoubiquitination of Rpn10 inhibits proteasome activity, the control of this reaction could be an interesting target. Classical proteasome inhibitors, such as Bortezomib, which target the proteolytic activity of the core particle of the proteasome, are used as anticancer drugs. We propose that Rpn10 monoubiquitination could be considered as a new target in cancer research.[cat] La degradació de proteïnes juga un rol essencial en la regulació i homeostasi de les cèl•lules eucariotes. Les proteïnes són marcades per a la seva degradació per ubicuitina lligases les quals annexen cadenes de poliubicuitina enviant de manera específica la proteïna al 26S proteasoma on serà degradada. Rpn10 és un receptor del proteasoma que reconeix cadenes d’ubicuitina a través del seu motiu UIM (ubiquitin-interacting motif). Rpn10 també conté un domini VWA el qual és responsable de les interaccions de Rpn10 en el seu context proteasomal. Rpn10 també es troba en equilibri amb una fracció no proteasomal. En el present treball hem demostrat que Rpn10 està monoubicuitinat (Rpn10-mUb) in vivo i que aquesta modificació es troba en ambdues fraccions proteasomal i extraproteasomal. Els nivells de Rpn10-mUb estan regulats in vivo per Rsp5, una ubicuitina lligasa de la família de les NEDD4, i Ubp2, una deubicuitinasa. Mitjançant mètodes genètics i de proteòmica es van trobar quatre lisines modificades per ubicuitina en la seqüència de Rpn10 (Lys71, Lys84, Lys99 and Lys268) éssent la Lys84 la diana preferent de Rsp5. La monoubicuitinació inhibeix la capacitat de Rpn10 d’unir substrats poliubicuitinats i en, conseqüència, és un mecanisme d’innactivació d’aquest receptor. També s’ha trobat que Rpn10-mUb disminuiex l’activitat del proteasoma. Els nostres resultats relacionen per primera vegada la monoubicuitinació amb la regulació del proteasoma, procés que imita la droga Bortezomib. Es proposa que Rpn10-mUb podria considerar-se com a nova diana en la teràpia contra el càncer

Characterization of rpn10 monoubiquitination as a novel way of proteasome regulation

Peer Reviewe

Defective in mitotic arrest 1 (Dma1) ubiquitin ligase controls G 1 cyclin degradation

Progression through the G1 phase of the cell cycle is controlled by diverse cyclin-dependent kinases (CDKs) that might be associated to numerous cyclin isoforms. Given such complexity, regulation of cyclin degradation should be crucial for coordinating progression through the cell cycle. In Saccharomyces cerevisiae, SCF is the only E3 ligase known to date to be involved in G1 cyclin degradation. Here, we report the design of a genetic screening that uncovered Dma1 as another E3 ligase that targets G1 cyclins in yeast. We show that the cyclin Pcl1 is ubiquitinated in vitro and in vivo by Dma1, and accordingly, is stabilized in dma1 mutants. We demonstrate that Pcl1 must be phosphorylated by its own CDK to efficiently interact with Dma1 and undergo degradation. A nonphosphorylatable version of Pcl1 accumulates throughout the cell cycle, demonstrating the physiological relevance of the proposed mechanism. Finally, we present evidence that the levels of Pcl1 and Cln2 are independently controlled in response to nutrient availability. This new previously unknown mechanism for G1 cyclin degradation that we report here could help elucidate the specific roles of the redundant CDK-cyclin complexes in G1.This work was supported by Spanish Ministry of Science and Innovation Grant BFU 2009-09728, now known as the Ministry of Economy and Competitiveness.Peer Reviewe

A new map to understand deubiquitination

Deubiquitination is a crucial mechanism in ubiquitin-mediated signalling networks. The importance of Dubs (deubiquitinating enzymes) as regulators of diverse cellular processes is becoming ever clearer as new roles are elucidated and new pathways are shown to be affected by this mechanism. Recent work, reviewed in the present paper, provides new perspective on the widening influence of Dubs and a new tool to focus studies of not only Dub interactions, but also potentially many more cellular systems. © The Authors Journal compilation.This work was supported by the Spanish Government Ministerio de Educación y Ciencia (grant number BFU2006-02928).Peer Reviewe

Monoubiquitination of RPN10 regulates substrate recruitment to the proteasome

El pdf del artículo es al manuscrito de autor (PubMed).-- et al.The proteasome recognizes its substrates via a diverse set of ubiquitin receptors, including subunits Rpn10/S5a and Rpn13. In addition, shuttling factors, such as Rad23, recruit substrates to the proteasome by delivering ubiquitinated proteins. Despite the increasing understanding of the factors involved in this process, the regulation of substrate delivery remains largely unexplored. Here we report that Rpn10 is monoubiquitinated in vivo and that this modification has profound effects on proteasome function. Monoubiquitination regulates the capacity of Rpn10 to interact with substrates by inhibiting Rpn10's ubiquitin-interacting motif (UIM). We show that Rsp5, a member of NEDD4 ubiquitin-protein ligase family, and Ubp2, a deubiquitinating enzyme, control the levels of Rpn10 monoubiquitination in vivo. Notably, monoubiquitination of Rpn10 is decreased under stress conditions, suggesting a mechanism of control of receptor availability mediated by the Rsp5-Ubp2 system. Our results reveal an unanticipated link between monoubiquitination signal and regulation of proteasome function.This work was performed in the Institute of Molecular Biology of Barcelona (IBMB), supported by Spanish Government (MICINN) grant BFU2006-02928. MS analysis was performed at Harvard Medical School (HMS). We acknowledge National Institutes of Health (NIH) grants GM065592 (D.F.) and GM67945 (S.G).Peer reviewe

Rpn10 monoubiquitination orchestrates the association of the ubiquilin-Type DSK2 receptor with the proteasome

Despite the progress made in understanding the roles of proteasome polyubiquitin receptors, such as the subunits Rpn10 (regulatory particle non-ATPase 10) and Rpn13, and the transient interactors Rad23 (radiation sensitivity abnormal 23) and Dsk2 (dual-specificity protein kinase 2), the mechanisms involved in their regulation are virtually unknown. Rpn10, which is found in the cell in proteasome-bound and-unbound pools, interacts with Dsk2, and this interaction has been proposed to regulate the amount of Dsk2 that gains access to the proteasome. Rpn10 monoubiquitination has emerged as a conserved mechanism with a strong effect on Rpn10 function. In the present study, we show that functional yeast proteasomes have the capacity to associate and dissociate with Rpn10 and that Rpn10 monoubiquitination decreases the Rpn10-proteasome and Rpn10-Dsk2 associations. Remarkably, this process facilitates the formation of Dsk2-proteasomes in vivo. Therefore, Rpn10 monoubiquitination acts as mechanism that serves to switch the proteasome from an 'Rpn10 high/Dsk2 low' state to an 'Rpn10 low/Dsk2 high' state. Interestingly, Rpn10-ubiquitin, with an inactivated ubiquitin-interacting motif (UIM), and Dsk2, with an inactive ubiquitin-like domain (UBL), show temperaturedependent phenotypes with multiple functional interactions.Peer Reviewe

Cold temperature induces the reprogramming of proteolytic pathways in yeast

Despite much evidence of the involvement of the proteasomeubiquitin signaling system in temperature stress response, the dynamics of the ubiquitylome during cold response has not yet been studied. Here, we have compared quantitative ubiquitylomes from a strain deficient in proteasome substrate recruitment and a reference strain during cold response. We have observed that a large group of proteins showing increased ubiquitylation in the proteasome mutant at low temperature is comprised by reverses suppressor of Ty-phenotype 5 (Rsp5)-regulated plasma membrane proteins. Analysis of internalization and degradation of plasma membrane proteins at low temperature showed that the proteasome becomes determinant for this process, whereas, at 30 °C, the proteasome is dispensable. Moreover, our observations indicate that proteasomes have increased capacity to interact with lysine 63-polyubiquitylated proteins during low temperature in vivo. These unanticipated observations indicate that, during cold response, there is a proteolytic cellular reprogramming in which the proteasome acquires a role in the endocytic-vacuolar pathway.This work was supported by Spanish Government (Ministry of Economy and Competitiveness) Grants BFU2009-06985 and BFU2012-35716, Agència de Gestió d'Ajuts Universitaris i de Recerca Grants 2009SGR-1482 and FI-DGR 2013, and European Molecular Biology Organization Fellowship ASTF 363.00-2010Peer Reviewe

Multiplexed, Proteome-Wide Protein Expression Profiling: Yeast Deubiquitylating Enzyme Knockout Strains

Characterizing a protein’s function often requires a description of the cellular state in its absence. Multiplexing in mass spectrometry-based proteomics has now achieved the ability to globally measure protein expression levels in yeast from 10 cell states simultaneously. We applied this approach to quantify expression differences in wild type and nine deubiquitylating enzyme (DUB) knockout strains with the goal of creating “information networks” that might provide deeper, mechanistic insights into a protein’s biological role. In total, more than 3700 proteins were quantified with high reproducibility across three biological replicates (30 samples in all). DUB mutants demonstrated different proteomics profiles, consistent with distinct roles for each family member. These included differences in total ubiquitin levels and specific chain linkages. Moreover, specific expression changes suggested novel functions for several DUB family members. For instance, the <i>ubp3Δ</i> mutant showed large expression changes for members of the cytochrome <i>C</i> oxidase complex, consistent with a role for Ubp3 in mitochondrial regulation. Several DUBs also showed broad expression changes for phosphate transporters as well as other components of the inorganic phosphate signaling pathway, suggesting a role for these DUBs in regulating phosphate metabolism. These data highlight the potential of multiplexed proteome-wide analyses for biological investigation and provide a framework for further study of the DUB family. Our methods are readily applicable to the entire collection of yeast deletion mutants and may help facilitate systematic analysis of yeast and other organisms