PCAF regulates the stability of the transcriptional regulator and cyclin-dependent kinase inhibitor p27Kip1

P27(Kip1) (p27) is a member of the Cip/Kip family of cyclin-dependent kinase inhibitors. Recently, a new function of p27 as transcriptional regulator has been reported. It has been shown that p27 regulates the expression of target genes mostly involved in splicing, cell cycle, respiration and translation. We report here that p27 directly binds to the transcriptional coactivator PCAF by a region including amino acids 91-120. PCAF associates with p27 through its catalytic domain and acetylates p27 at lysine 100. Our data showed that overexpression of PCAF induces the degradation of p27 whereas in contrast, the knockdown of PCAF stabilizes the protein. A p27 mutant in which K100 was substituted by arginine (p27-K100R) cannot be acetylated by PCAF and has a half-life much higher than that of p27WT. Moreover, p27-K100R remains stable along cell-cycle progression. Ubiquitylation assays and the use of proteasome inhibitors indicate that PCAF induces p27 degradation via proteasome. We also observed that knockdown of skp2 did not affect the PCAF induced degradation of p27. In conclusion, our data suggest that the p27 acetylation by PCAF regulates its stability.

Oligomerization and DNA binding of Ler, a master regulator of pathogenicity of enterohemorrhagic and enteropathogenic Escherichia coli

Ler is a DNA-binding, oligomerizable protein that regulates pathogenicity islands in enterohemorrhagic and enteropathogenic Escherichia coli strains. Ler counteracts the transcriptional silencing effect of H-NS, another oligomerizable nucleoid-associated protein. We studied the oligomerization of Ler in the absence and presence of DNA by atomic force microscopy. Ler forms compact particles with a multimodal size distribution corresponding to multiples of 35 units of Ler. DNA wraps around Ler particles that contain more than 1516 Ler monomers. The resulting shortening of the DNA contour length is in agreement with previous measurements of the length of DNA protected by Ler in footprinting assays. We propose that the repetition unit corresponds to the number of monomers per turn of a tight helical Ler oligomer. While the repressor (H-NS) and anti-repressor (Ler) have similar DNA-binding domains, their oligomerization domains are unrelated. We suggest that the different oligomerization behavior of the two proteins explains the opposite results of their interaction with the same or proximal regions of DNA.

Essential residues in the H-NS binding site of Hha, a co-regulator of horizontally acquired genes in Enterobacteria

Proteins of the Hha/YmoA family co-regulate with H-NS the expression of horizontally acquired genes in Enterobacteria. Systematic mutations of conserved acidic residues in Hha have allowed the identification of D48 as an essential residue for H-NS binding and the involvement of E25. Mutations of these residues resulted in deregulation of sensitive genes in vivo. D48 is only partially solvent accessible, yet it defines the functional binding interface between Hha and H-NS confirming that Hha has to undergo a conformational change to bind H-NS. Exposed acidic residues, such as E25, may electrostatically facilitate and direct the approach of Hha to the positively charged region of H-NS enabling the formation of the final complex when D48 becomes accessible by a conformational change of Hha.

Association of MMP-2 polymorphisms with severe and very severe COPD: a case control study of MMPs-1, 9 and 12 in a European population.

BACKGROUND: Genetic factors play a role in chronic obstructive pulmonary disease (COPD) but are poorly understood. A number of candidate genes have been proposed on the basis of the pathogenesis of COPD. These include the matrix metalloproteinase (MMP) genes which play a role in tissue remodelling and fit in with the protease--antiprotease imbalance theory for the cause of COPD. Previous genetic studies of MMPs in COPD have had inadequate coverage of the genes, and have reported conflicting associations of both single nucleotide polymorphisms (SNPs) and SNP haplotypes, plausibly due to under-powered studies. METHODS: To address these issues we genotyped 26 SNPs, providing comprehensive coverage of reported SNP variation, in MMPs- 1, 9 and 12 from 977 COPD patients and 876 non-diseased smokers of European descent and evaluated their association with disease singly and in haplotype combinations. We used logistic regression to adjust for age, gender, centre and smoking history. RESULTS: Haplotypes of two SNPs in MMP-12 (rs652438 and rs2276109), showed an association with severe/very severe disease, corresponding to GOLD Stages III and IV. CONCLUSIONS: Those with the common A-A haplotype for these two SNPs were at greater risk of developing severe/very severe disease (p = 0.0039) while possession of the minor G variants at either SNP locus had a protective effect (adjusted odds ratio of 0.76; 95% CI 0.61 - 0.94). The A-A haplotype was also associated with significantly lower predicted FEV1 (42.62% versus 44.79%; p = 0.0129). This implicates haplotypes of MMP-12 as modifiers of disease severity.

The SUD1 Gene Encodes a Putative E3 Ubiquitin Ligase and Is a Positive Regulator of 3-Hydroxy-3-Methylglutaryl Coenzyme A Reductase Activity in Arabidopsis

The 3-hydroxy-3-methylglutaryl-CoA reductase (HMGR) enzyme catalyzes the major rate-limiting step of the mevalonic acid (MVA) pathway from which sterols and other isoprenoids are synthesized. In contrast with our extensive knowledge of the regulation of HMGR in yeast and animals, little is known about this process in plants. To identify regulatory components of the MVA pathway in plants, we performed a genetic screen for second-site suppressor mutations of the Arabidopsis thaliana highly drought-sensitive drought hypersensitive2 (dry2) mutant that shows decreased squalene epoxidase activity. We show that mutations in SUPPRESSOR OF DRY2 DEFECTS1 (SUD1) gene recover most developmental defects in dry2 through changes in HMGR activity. SUD1 encodes a putative E3 ubiquitin ligase that shows sequence and structural similarity to yeast Degradation of a factor (Doa10) and human TEB4, components of the endoplasmic reticulum-associated degradation C (ERAD-C) pathway. While in yeast and animals, the alternative ERAD-L/ERAD-M pathway regulates HMGR activity by controlling protein stability, SUD1 regulates HMGR activity without apparent changes in protein content. These results highlight similarities, as well as important mechanistic differences, among the components involved in HMGR regulation in plants, yeast, and animals.

TP53 induced glycolysis and apoptosis regulator (TIGAR) knockdown results in radiosensitization of glioma cells

Background and purpose: The TP53 induced glycolysis and apoptosis regulator (TIGAR) functions to lower fructose-2,6-bisphosphate (Fru-2,6-P2) levels in cells, consequently decreasing glycolysis and leading to the scavenging of reactive oxygen species (ROS), which correlate with a higher resistance to cell death. The decrease in intracellular ROS levels in response to TIGAR may also play a role in the ability of p53 to protect from the accumulation of genomic lesions. Given these good prospects of TIGAR for metabolic regulation and p53-response modulation, we analyzed the effects of TIGAR knockdown in U87MG and T98G glioblastoma-derived cell lines. Methods/results: After TIGAR-knockdown in glioblastoma cell lines, different metabolic parameters were assayed, showing an increase in Fru-2,6-P2, lactate and ROS levels, with a concomitant decrease in reduced glutathione (GSH) levels. In addition, cell growth was inhibited without evidence of apoptotic or autophagic cell death. In contrast, a clear senescent phenotype was observed. We also found that TIGAR protein levels were increased shortly after irradiation. In addition, avoiding radiotherapy-triggered TIGAR induction by gene silencing resulted in the loss of capacity of glioblastoma cells to form colonies in culture and the delay of DNA repair mechanisms, based in c-H2AX foci, leading cells to undergo morphological changes compatible with a senescent phenotype. Thus, the results obtained raised the possibility to consider TIGAR as a therapeutic target to increase radiotherapy effects. Conclusion: TIGAR abrogation provides a novel adjunctive therapeutic strategy against glial tumors by increasing radiation-induced cell impairment, thus allowing the use of lower radiotherapeutic doses.

Mergers, investment decisions and internal organisation

We analyse the effects of investment decisions and firms' internal organisation on the efficiency and stability of horizontal mergers. In our framework economies of scale are endogenous and there might be internal conflict within merged firms. We show that often stable mergers do not lead to more efficiency and may even lead to efficiency losses. These mergers lead to lower total welfare, suggesting that a regulator should be careful in assuming that possible efficiency gains of a merger will be effiectively realised. Moreover, the paper offers a possible explanation for merger failures.

Pressure groups and experts in environmental regulation

We study a problem of adverse selection in the context of environmental regulation, where the firm may suffer from a certain degree of ignorance about its own type. In a framework like the construction of a certain infrastructure project, the presence of ignorance about its impact on the environment, can play an important role in the determination of the regulatory policy. First, an optimal contract is constructed for any exogenous level of ignorance. Second, the presence of potentially informed third-parties is studied from the perspective of the regulator, which allows us to analyze the impact on the efficiency of the contract, of the presence of environmentalists and of experts. Then, we obtain some insights on how the problem differs when the degree of ignorance is a choice variable for the firm. We finally use our results to derive policy implications concerning the existing envoronmental regulation, and the potential role of interested parties as information providers.

A Retail benchmarking approach to eficient two-way access pricing

We study a retail benchmarking approach to determine access prices for interconnected networks. Instead of considering fixed access charges as in the existing literature, we study access pricing rules that determine the access price that network i pays to network j as a linear function of the marginal costs and the retail prices set by both networks. In the case of competition in linear prices, we show that there is a unique linear rule that implements the Ramsey outcome as the unique equilibrium, independently of the underlying demand conditions. In the case of competition in two-part tariffs, we consider a class of access pricing rules, similar to the optimal one under linear prices but based on average retail prices. We show that firms choose the variable price equal to the marginal cost under this class of rules. Therefore, the regulator (or the competition authority) can choose one among the rules to pursue additional objectives such as consumer surplus, network covera.

Regulació del factor de transcripció NFAT5 per les quinases WNK1, SPAK i OSR1 i cerca de quinases activadores de la pròpia WNK1 en resposta a estrés osmòtic

Estudi elaborat a partir d’una estada a la School of Life Sciences de la University of Dundee, Gran Bretanya, entre gener i març del 2007.L'estrès osmòtic causa rà pidament l'activació de la quinasa WNK1, que fosforila i activa a continuació les quinases SPAK i OSR1, que alhora regulen canals i transportadors d’ions preexistents a la membrana cel•lular. El factor de transcripció NFAT5 és el principal regulador de la resposta cel•lular transcripcional secundà ria a hipertonicitat i s’ha descrit que les quinases p38, Fyn, PKA, ERK/MEK i ATM estan involucrades en la seva regulació post-traduccional. No obstant, com que la funció d’aquestes quinases no explica totalment els mecanismes d'activació de NFAT5, s’ha estudiat si l’activitat transcripcional de NFAT5 pot estar regulada per WNK1, SPAK o OSR1. Així doncs, es va observar que l’activitat d’un reporter dependent de NFAT5 no es veu afectada per la presència de cap de les quinases anteriors, en la seva forma wild-type o dominant negatiu. D’altra banda, es va estudiar quin domini de WNK1 és necessari per a que pugui respondre a hipertonicitat i quines quinases poden estar involucrades en la fosforilació de la serina 382 de WNK1. En conclusió, les dades obtingudes apunten que l’activació de WNK1 en resposta a estrès osmòtic requereix la seva fosforilació en la serina 382 per quinases upstream com PAK2 o RSK i que també és necessari un dels seus dominis coiled-coil, almenys els aminoà cids 558 i 561. Aquests processos, però, semblen ser independents de l’activació de NFAT5 en resposta a hipertonicitat.   

Mobile termination and mobile penetration

In this paper, we study how access pricing affects network competition when subscription demand is elastic and each network uses non-linear prices and can apply termination-based price discrimination. In the case of a fixed per minute termination charge, we find that a reduction of the termination charge below cost has two opposing effects: it softens competition but helps to internalize network externalities. The former reduces mobile penetration while the latter boosts it. We find that firms always prefer termination charge below cost for either motive while the regulator prefers termination below cost only when this boosts penetration. Next, we consider the retail benchmarking approach (Jeon and Hurkens, 2008) that determines termination charges as a function of retail prices and show that this approach allows the regulator to increase penetration without distorting call volumes.

Estudi de la degradació anaeròbia de tres contaminants emergents: àcid clofíbric, carbamazepina i ibuprofè

En el present estudi s’ha fet un seguiment baromètric de la producció de biogàs de tres compostos farmacèutics mitjançant tests de degradació anaeròbia. El llot del digestor anaerobi de l’estació depuradora d’aigües residuals (EDAR) de Granollers s’ha emprat com a inòcul. En un primer experiment, s’ha fet l’estudi de l’etapa metanogènica, amb metanol com a substrat i les concentracions de 5, 10 i 20 mg·L-1 d’àcid clofíbric (CLOFI, regulador lípid), ibuprofè (IBU, anti-inflamatori) i carbamazepina (CARBA, antiepilèptic/analgèsic). En un segon experiment, s’ha estudiat l’etapa acetogènica, amb àcid propiònic com a substrat i amb concentracions de 20 i 100 mg·L-1 dels mateixos fàrmacs. Per una banda, en el primer experiment, no s’ha observat degradació dels fàrmacs i cap d’ells presenta toxicitat pel llot anaerobi. CARBA i IBU s’adsorbeixen majoritàriament a la fase sòlida. Per altra banda, en l’etapa acetogènica ni CARBA ni IBU s’han degradat i tampoc presenten toxicitat en cap de les concentracions provades. Com en el cas anterior, la majoria d’aquests fàrmacs queda adsorbit en el llot. En canvi, CLOFI es degrada en 100 mg·L-1 de concentració donant com a producte intermedi un metabòlit.

The Role of model uncertainty and learning in the U.S. postwar policy response to oil prices

This paper studies optimal monetary policy in a framework that explicitly accounts for policymakers' uncertainty about the channels of transmission of oil prices into the economy. More specfically, I examine the robust response to the real price of oil that US monetary authorities would have been recommended to implement in the period 1970 2009; had they used the approach proposed by Cogley and Sargent (2005b) to incorporate model uncertainty and learning into policy decisions. In this context, I investigate the extent to which regulator' changing beliefs over different models of the economy play a role in the policy selection process. The main conclusion of this work is that, in the specific environment under analysis, one of the underlying models dominates the optimal interest rate response to oil prices. This result persists even when alternative assumptions on the model's priors change the pattern of the relative posterior probabilities, and can thus be attributed to the presence of model uncertainty itself.

On the joint production of research and training

Universities and research institutions have the responsibility to produce science and to provide training to new generations of researchers. In this paper, we propose a model to analyze the determinants of a senior scientist's decisions about allocating time between these tasks. The results of this decision depend upon the characteristics of the research project, the senior scientist's concern for training and the expected innate ability of the junior scientist involved. We analyze the role that a regulator can play in defining both the value of scientific projects and the future population of independent scientists.

Progresión tumoral y búsqueda de nuevas dianas terapéuticas en la familia de tumores del sarcoma de Ewing: función biológica de la caveolina-1

El sarcoma de Ewing es el segundo tumor óseo infantil más frecuente y presenta una alta incidencia de enfermedad metastática. Este tipo de tumores presentan una traslocación génica característica que da origen a una proteína de fusión, normalmente EWS/FLI1. Esta proteína de fusión actúa como factor de transcripción aberrante regulando la expresión de diferentes genes implicados en la iniciación, mantenimiento y progresión del tumor. Nuestro grupo describió como uno de estos genes diana a la caveolina 1 (CAV1) describiendo además su papel determinante en el fenotipo maligno del sarcoma de Ewing, en la tumorigénesis y en la resistencia a apoptosis inducida por quimioterapia. Para investigar el papel concreto de CAV1 en el proceso metastático de este sarcoma, creamos un modelo de baja expresión de CAV1 en líneas celulares de sarcoma de Ewing y determinamos cambios en su capacidad migratoria, invasiva y metastática. En los ensayos in vitro hallamos una menor capacidad migratoria de las células knockdown de CAV1 y una reducción en la expresión de MMP9 y en la actividad de MMP2. La regulación de la actividad de MMP2 parece estar relacionada con la posible regulación que ejerce CAV1 en la función de MT1-MMP, proteína fundamental para la activación de MMP2. Por otro lado, en este estudio proponemos que CAV1 promueve la expresión de MMP9 tanto transcripcionalmente, regulando la vía de señalización ERK1/2, como a nivel post-transcripcional regulando la vía RSK1/rpS6. Además, en los ensayos de metástasis experimental in vivo las células knockdown de CAV1 presentaron una menor incidencia de metástasis pulmonar, hecho que correlacionó con una disminución en la expresión de SPARC, una proteína de adhesión importante en procesos metastáticos. En resumen, nuestros resultados evidencian la importancia de CAV1 en el proceso metastático del sarcoma de Ewing.