Iron (Fe) is widely suspected as a key controlling factor of N2 fixation due to the high Fe content of nitrogenase and photosynthetic enzymes complex, and to its low concentrations in oceanic surface seawaters. The influence of Fe limitation on the recently discovered unicellular diazotrophic cyanobacteria (UCYN) is poorly understood despite their biogeochemical importance in the carbon and nitrogen cycles. To address this knowledge gap, we conducted culture experiments on Crocosphaera watsonii WH8501 growing under a range of dissolved Fe concentrations (from 3.3 to 403 nM). Overall, severe Fe limitation led to significant decreases in growth rate (2.6-fold), C, N and chlorophyll a contents per cell (up to 4.1-fold), N2 and CO2 fixation rates per cell (17- and 7-fold) as well as biovolume (2.2-fold). We highlighted a two phased response depending on the degree of limitation: (i) under a moderate Fe limitation, the biovolume of C. watsonii was strongly reduced, allowing the cells to keep sufficient energy to maintain an optimal growth, volume-normalized contents and N2 and CO2 fixation rates; (ii) with increasing Fe deprivation, biovolume remained unchanged but the entire cell metabolism was affected, as shown by a strong decrease in the growth rate, volume-normalized contents and N2 and CO2 fixation rates. The half-saturation constant for growth of C. watsonii with respect to Fe is twice as low as that of the filamentous Trichodesmium indicating a better adaptation of C. watsonii to poor Fe environments than filamentous diazotrophs. The physiological response of C. watsonii to Fe limitation was different from that previously shown on the UCYN Cyanothece sp, suggesting potential differences in Fe requirements and/or Fe acquisition within the UCYN community. These results contribute to a better understanding of how Fe bioavailability can control the activity of UCYN and explain the biogeography of diverse N2 fixers in ocean. 相似文献
The tetrodotoxin‐resistant (TTX‐R) voltage‐gated sodium channel Nav1.8 is predominantly expressed in peripheral afferent neurons, but in case of neuronal injury an ectopic and detrimental expression of Nav1.8 occurs in neurons of the CNS. In CNS neurons, Nav1.2 and Nav1.6 channels accumulate at the axon initial segment, the site of the generation of the action potential, through a direct interaction with the scaffolding protein ankyrin G (ankG). This interaction is regulated by protein kinase CK2 phosphorylation. In this study, we quantitatively analyzed the interaction between Nav1.8 and ankG. GST pull‐down assay and surface plasmon resonance technology revealed that Nav1.8 strongly and constitutively interacts with ankG, in comparison to what observed for Nav1.2. An ion channel bearing the ankyrin‐binding motif of Nav1.8 displaced the endogenous Nav1 accumulation at the axon initial segment of hippocampal neurons. Finally, Nav1.8 and ankG co‐localized in skin nerves fibers. Altogether, these results indicate that Nav1.8 carries all the information required for its localization at ankG micro‐domains. The constitutive binding of Nav1.8 with ankG could contribute to the pathological aspects of illnesses where Nav1.8 is ectopically expressed in CNS neurons.
Two-dimensional gel electrophoresis (2DE) is a powerful tool to uncover proteome modifications potentially related to different physiological or pathological conditions. Basically, this technique is based on the separation of proteins according to their isoelectric point in a first step, and secondly according to their molecular weights by SDS polyacrylamide gel electrophoresis (SDS-PAGE). In this report an optimized sample preparation protocol for little amount of human post-mortem and mouse brain tissue is described. This method enables to perform both two-dimensional fluorescence difference gel electrophoresis (2D-DIGE) and mini 2DE immunoblotting. The combination of these approaches allows one to not only find new proteins and/or protein modifications in their expression thanks to its compatibility with mass spectrometry detection, but also a new insight into markers validation. Thus, mini-2DE coupled to western blotting permits to identify and validate post-translational modifications, proteins catabolism and provides a qualitative comparison among different conditions and/or treatments. Herein, we provide a method to study components of protein aggregates found in AD and Lewy body dementia such as the amyloid-beta peptide and the alpha-synuclein. Our method can thus be adapted for the analysis of the proteome and insoluble proteins extract from human brain tissue and mice models too. In parallel, it may provide useful information for the study of molecular and cellular pathways involved in neurodegenerative diseases as well as potential novel biomarkers and therapeutic targets. 相似文献
Gap junctions (GJ) represent a cellular communication system known to influence neuronal differentiation and survival. To
assess a putative role of this system for neural effects of tamoxifen (TAM) and raloxifene (RAL), we used the human teratocarcinoma
cell line NTera2/D1, retinoic acid (RA)-dependent neuronal differentiation of which is regulated by gap junctions formed of
connexin43 (Cx43). As demonstrated by Western blot analysis, concentrations above 1 μmol/l for TAM, and 0.1 μmol/l for RAL
lead to a temporary time- and concentration-dependent increase in Cx43 immunoreactivity, which reached a peak for TAM after
1 day and for RAL after 2 days. Immunocytochemical stainings revealed the increase in Cx43 immunoreactivity to result from
an accumulation in intracellular compartments such as the Golgi apparatus or lysosomes. In addition, TAM and RAL were able
to prevent the RA-dependent decrease of Cx43 immunoreactivity in NTera2/D1 cells, normally observed during neuronal differentiation.
This suggested a suppression of neuronal differentiation to result from these substances. According to this, treatment of
NTera2/D1 cells with 10 μmol/l TAM or RAL during weeks 1 and 2 of a 6 weeks RA-driven differentiation schedule impaired, whereas
treatment during weeks 5 and 6 did not impair, neuronal differentiation of these cells. Modulation of GJ coupling between
NTera2/D1 cells by TAM and RAL seems therefore to perturb early neuronal differentiation, whereas differentiated neurons in
the mature brain seem to be not affected. These effects could be of importance for actions of TAM and RAL on early embryonic
steps of nervous system formation. 相似文献
Vascular endothelial growth factor (VEGF) induces angiogenesis and regulates endothelial function via production and release of nitric oxide (NO), an important signaling molecule. The molecular basis leading to NO production involves phosphatidylinositiol-3 kinase (PI3K), Akt, and endothelial nitric-oxide synthase (eNOS) activation. In this study, we have examined whether small GTP-binding proteins of the ADP-ribosylation factor (ARF) family act as molecular switches to regulate signaling cascades activated by VEGF in endothelial cells. Our results show that this growth factor can promote the rapid and transient activation of ARF1. In endothelial cells, this GTPase is present on dynamic plasma membrane ruffles. Inhibition of ARF1 expression, using RNA interference, markedly impaired VEGF-dependent eNOS phosphorylation and NO production by preventing the activation of the PI3K/Akt signaling axis. Furthermore, our data indicate that phosphorylation of Tyr801, on VEGF receptor 2, is essential for activating Src- and ARF1-dependent signaling events leading to NO release from endothelial cells. Lastly, this mediator is known to regulate a broad variety of endothelial cell functions. Depletion of ARF1 markedly inhibits VEGF-dependent increase of vascular permeability as well as capillary tubule formation, a process important for angiogenesis. Taken together, our data indicate that ARF1 is a novel modulator of VEGF-stimulated NO release and signaling in endothelial cells. 相似文献
A vast majority of dyslexic children exhibit a phonological deficit, particularly noticeable in phonemic identification or discrimination tasks. The gap in performance between dyslexic and normotypical listeners appears to decrease into adulthood, suggesting that some individuals with dyslexia develop compensatory strategies. Some dyslexic adults however remain impaired in more challenging listening situations such as in the presence of background noise. This paper addresses the question of the compensatory strategies employed, using the recently developed Auditory Classification Image (ACI) methodology. The results of 18 dyslexics taking part in a phoneme categorization task in noise were compared with those of 18 normotypical age-matched controls. By fitting a penalized Generalized Linear Model on the data of each participant, we obtained his/her ACI, a map of the time-frequency regions he/she relied on to perform the task. Even though dyslexics performed significantly less well than controls, we were unable to detect a robust difference between the mean ACIs of the two groups. This is partly due to the considerable heterogeneity in listening strategies among a subgroup of 7 low-performing dyslexics, as confirmed by a complementary analysis. When excluding these participants to restrict our comparison to the 11 dyslexics performing as well as their average-reading peers, we found a significant difference in the F3 onset of the first syllable, and a tendency of difference on the F4 onset, suggesting that these listeners can compensate for their deficit by relying upon additional allophonic cues. 相似文献
Sex chromosomes often carry large nonrecombining regions that can extend progressively over time, generating evolutionary strata of sequence divergence. However, some sex chromosomes display an incomplete suppression of recombination. Large genomic regions without recombination and evolutionary strata have also been documented around fungal mating-type loci, but have been studied in only a few fungal systems. In the model fungus Podospora anserina (Ascomycota, Sordariomycetes), the reference S strain lacks recombination across a 0.8-Mb region around the mating-type locus. The lack of recombination in this region ensures that nuclei of opposite mating types are packaged into a single ascospore (pseudohomothallic lifecycle). We found evidence for a lack of recombination around the mating-type locus in the genomes of ten P. anserina strains and six closely related pseudohomothallic Podospora species. Importantly, the size of the nonrecombining region differed between strains and species, as indicated by the heterozygosity levels around the mating-type locus and experimental selfing. The nonrecombining region is probably labile and polymorphic, differing in size and precise location within and between species, resulting in occasional, but infrequent, recombination at a given base pair. This view is also supported by the low divergence between mating types, and the lack of strong linkage disequilibrium, chromosomal rearrangements, transspecific polymorphism and genomic degeneration. We found a pattern suggestive of evolutionary strata in P. pseudocomata. The observed heterozygosity levels indicate low but nonnull outcrossing rates in nature in these pseudohomothallic fungi. This study adds to our understanding of mating-type chromosome evolution and its relationship to mating systems. 相似文献
In this study, we synthesized a new congener series of N-sulphonylhydrazones designed as candidate ROCK inhibitors using the molecular hybridization of the clinically approved drug fasudil (1) and the IKK-β inhibitor LASSBio-1524 (2). Among the synthesized compounds, the N-methylated derivative 11 (LASSBio-2065) showed the best inhibitory profile for both ROCK isoforms, with IC50 values of 3.1 and 3.8?µM for ROCK1 and ROCK2, respectively. Moreover, these compounds were also active in the scratch assay performed in human breast cancer MDA-MB 231 cells and did not display toxicity in MTT and LDH assays. Molecular modelling studies provided insights into the possible binding modes of these N-sulphonylhydrazones, which present a new molecular architecture capable of being optimized and developed as therapeutically useful ROCK inhibitors. 相似文献
We conducted a comprehensive analysis of virulence in the fungal wheat pathogen Zymoseptoria tritici using quantitative trait locus (QTL) mapping. High‐throughput phenotyping based on automated image analysis allowed the measurement of pathogen virulence on a scale and with a precision that was not previously possible. Across two mapping populations encompassing more than 520 progeny, 540 710 pycnidia were counted and their sizes and grey values were measured. A significant correlation was found between pycnidia size and both spore size and number. Precise measurements of percentage leaf area covered by lesions provided a quantitative measure of host damage. Combining these large and accurate phenotypic datasets with a dense panel of restriction site‐associated DNA sequencing (RADseq) genetic markers enabled us to genetically dissect pathogen virulence into components related to host damage and those related to pathogen reproduction. We showed that different components of virulence can be under separate genetic control. Large‐ and small‐effect QTLs were identified for all traits, with some QTLs specific to mapping populations, cultivars and traits and other QTLs shared among traits within the same mapping population. We associated the presence of four accessory chromosomes with small, but significant, increases in several virulence traits, providing the first evidence for a meaningful function associated with accessory chromosomes in this organism. A large‐effect QTL involved in host specialization was identified on chromosome 7, leading to the identification of candidate genes having a large effect on virulence. 相似文献