Effect of oocyte diameter on meiotic competence, embryo development, p34 (cdc2) expression and MPF activity in prepubertal goat oocytes

The aim of this study was to analyze the relationship between oocyte diameter, meiotic and embryo developmental competence and the expression of the catalytic subunit of MPF, the p34(cdc2), at mRNA, RNA and protein level, as well as its kinase activity, in prepubertal (1-2 months old) goat oocytes. MPF is the main meiotic regulator and a possible regulator of cytoplasmic maturation; therefore, it could be a key factor in understanding the differences between competent and incompetent oocytes. Oocytes were classified according to oocyte diameter in four categories: <110, 110-125, 125-135 and >135 microm and matured, fertilized and cultured in vitro. The p34(cdc2) was analyzed in oocytes at the time of collection (0 h) and after 27 h of IVM (27 h) in each of the oocyte diameter categories. The oocyte diameter was positively related to the percentage of oocytes at MII after IVM (0, 20.7, 58 and 78%, respectively) and the percentage of blastocysts obtained at 8 days postinsemination (0, 0, 1.95 and 12.5%, respectively). The expression of RNA and mRNA p34(cdc2) did not vary between oocyte diameters at 0 and 27h. Protein expression of p34(cdc2) increased in each oocyte category after 27 h of maturation. MPF activity among diameter groups did not vary at 0h but after IVM there was a clear and statistically significant increase of MPF activity in the biggest oocytes.     

Epidemiology and Prognosis of Coagulase-Negative Staphylococcal Endocarditis: Impact of Vancomycin Minimum Inhibitory Concentration

This study describes coagulase-negative staphylococcal (CoNS) infective endocarditis (IE) epidemiology at our institution, the antibiotic susceptibility profile, and the influence of vancomycin minimum inhibitory concentration (MIC) on patient outcomes. One hundred and three adults with definite IE admitted to an 850-bed tertiary care hospital in Barcelona from 1995-2008 were prospectively included in the cohort. We observed that CoNS IE was an important cause of community-acquired and healthcare-associated IE; one-third of patients involved native valves. Staphylococcus epidermidis was the most frequent species, methicillin-resistant in 52% of patients. CoNS frozen isolates were available in 88 patients. Vancomycin MICs of 2.0 μg/mL were common; almost all cases were found among S. epidermidis isolates and did not increase over time. Eighty-five patients were treated either with cloxacillin or vancomycin: 38 patients (Group 1) were treated with cloxacillin, and 47 received vancomycin; of these 47, 27 had CoNS isolates with a vancomycin MIC <2.0 μg/mL (Group 2), 20 had isolates with a vancomycin MIC ≥2.0 μg/mL (Group 3). One-year mortality was 21%, 48%, and 65% in Groups 1, 2, and 3, respectively (P=0.003). After adjusting for confounders and taking Group 2 as a reference, methicillin-susceptibility was associated with lower 1-year mortality (OR 0.12, 95% CI 0.02-0.55), and vancomycin MIC ≥2.0 μg/mL showed a trend to higher 1-year mortality (OR 3.7, 95% CI 0.9-15.2; P=0.069). Other independent variables associated with 1-year mortality were heart failure (OR 6.2, 95% CI 1.5-25.2) and pacemaker lead IE (OR 0.1, 95%CI 0.02-0.51). In conclusion, methicillin-resistant S.epidermidis was the leading cause of CoNS IE, and patients receiving vancomycin had higher mortality rates than those receiving cloxacillin; mortality was higher among patients having isolates with vancomycin MICs ≥2.0 μg/mL.     

Analysis and Dynamics of the Chromosomal Complements of Wild Sparkling-Wine Yeast Strains

We isolated Saccharomyces cerevisiae yeast strains that are able to carry out the second fermentation of sparkling wine from spontaneously fermenting musts in El Penedès (Spain) by specifically designed selection protocols. All of them (26 strains) showed one of two very similar mitochondrial DNA (mtDNA) restriction patterns, whereas their karyotypes differed. These strains showed high rates of karyotype instability, which were dependent on both the medium and the strain, during vegetative growth. In all cases, the mtDNA restriction pattern was conserved in strains kept under the same conditions. Analysis of different repetitive sequences in their genomes suggested that ribosomal DNA repeats play an important role in the changes in size observed in chromosome XII, whereas SUC genes or Ty elements did not show amplification or transposition processes that could be related to rearrangements of the chromosomes showing these sequences. Karyotype changes also occurred in monosporidic diploid derivatives. We propose that these changes originated mainly from ectopic recombination between repeated sequences interspersed in the genome. None of the rearranged karyotypes provided a selective advantage strong enough to allow the strains to displace the parental strains. The nature and frequency of these changes suggest that they may play an important role in the establishment and maintenance of the genetic diversity observed in S. cerevisiae wild populations.     

Phytoplankton across Tropical and Subtropical Regions of the Atlantic,Indian and Pacific Oceans

We examine the large-scale distribution patterns of the nano- and microphytoplankton collected from 145 oceanic stations, at 3 m depth, the 20% light level and the depth of the subsurface chlorophyll maximum, during the Malaspina-2010 Expedition (December 2010-July 2011), which covered 15 biogeographical provinces across the Atlantic, Indian and Pacific oceans, between 35°N and 40°S. In general, the water column was stratified, the surface layers were nutrient-poor and the nano- and microplankton (hereafter phytoplankton, for simplicity, although it included also heterotrophic protists) community was dominated by dinoflagellates, other flagellates and coccolithophores, while the contribution of diatoms was only important in zones with shallow nutriclines such as the equatorial upwelling regions. We applied a principal component analysis to the correlation matrix among the abundances (after logarithmic transform) of the 76 most frequent taxa to synthesize the information contained in the phytoplankton data set. The main trends of variability identified consisted of: 1) A contrast between the community composition of the upper and the lower parts of the euphotic zone, expressed respectively by positive or negative scores of the first principal component, which was positively correlated with taxa such as the dinoflagellates Oxytoxum minutum and Scrippsiella spp., and the coccolithophores Discosphaera tubifera and Syracosphaera pulchra (HOL and HET), and negatively correlated with taxa like Ophiaster hydroideus (coccolithophore) and several diatoms, 2) a general abundance gradient between phytoplankton-rich regions with high abundances of dinoflagellate, coccolithophore and ciliate taxa, and phytoplankton-poor regions (second principal component), 3) differences in dominant phytoplankton and ciliate taxa among the Atlantic, the Indian and the Pacific oceans (third principal component) and 4) the occurrence of a diatom-dominated assemblage (the fourth principal component assemblage), including several pennate taxa, Planktoniella sol, Hemiaulus hauckii and Pseudo-nitzschia spp., in the divergence regions. Our findings indicate that consistent assemblages of co-occurring phytoplankton taxa can be identified and that their distribution is best explained by a combination in different degrees of both environmental and historical influences.     

Molecular characterization of the endoplasmic reticulum: insights from proteomic studies

The endoplasmic reticulum (ER) is a multifunctional intracellular organelle responsible for the synthesis, processing and trafficking of a wide variety of proteins essential for cell growth and survival. Therefore, comprehensive characterization of the ER proteome is of great importance to the understanding of its functions and has been actively pursued in the past decade by scientists in the proteomics field. This review summarizes major proteomic studies published in the past decade that focused on the ER proteome. We evaluate the data sets obtained from two different organs, liver and pancreas each of which contains a primary cell type (hepatocyte and acinar cell) with specialized functions. We also discuss how the nature of the proteins uncovered is related to the methods of organelle purification, organelle purity and the techniques used for protein separation prior to MS. In addition, this review also puts emphasis on the biological insights gained from these studies regarding the molecular functions of the ER including protein synthesis and translocation, protein folding and quality control, ER-associated degradation and ER stress, ER export and membrane trafficking, calcium homeostasis and detoxification and drug metabolism.     

Novel effect of C75 on carnitine palmitoyltransferase I activity and palmitate oxidation

C75 is a potential drug for the treatment of obesity. It was first identified as a competitive, irreversible inhibitor of fatty acid synthase (FAS). It has also been described as a malonyl-CoA analogue that antagonizes the allosteric inhibitory effect of malonyl-CoA on carnitine palmitoyltransferase I (CPT I), the main regulatory enzyme involved in fatty acid oxidation. On the basis of MALDI-TOF analysis, we now provide evidence that C75 can be transformed to its C75-CoA derivative. Unlike the activation produced by C75, the CoA derivative is a potent competitive inhibitor that binds tightly but reversibly to CPT I. IC50 values for yeast-overexpressed L- or M-CPT I isoforms, as well as for purified mitochondria from rat liver and muscle, were within the same range as those observed for etomoxiryl-CoA, a potent inhibitor of CPT I. When a pancreatic INS(823/13), muscle L6E9, or kidney HEK293 cell line was incubated directly with C75, fatty acid oxidation was inhibited. This suggests that C75 could be transformed in the cell to its C75-CoA derivative, inhibiting CPT I activity and consequently fatty acid oxidation. In vivo, a single intraperitoneal injection of C75 in mice produced short-term inhibition of CPT I activity in mitochondria from the liver, soleus, and pancreas, indicating that C75 could be transformed to its C75-CoA derivative in these tissues. Finally, in silico molecular docking studies showed that C75-CoA occupies the same pocket in CPT I as palmitoyl-CoA, suggesting an inhibiting mechanism based on mutual exclusion. Overall, our results describe a novel role for C75 in CPT I activity, highlighting the inhibitory effect of its C75-CoA derivative.