Oligomers of the ATPase EHD2 confine caveolae to the plasma membrane through association with actin

Caveolae are specialized domains present in the plasma membrane (PM) of most mammalian cell types. They function in signalling, membrane regulation, and endocytosis. We found that the Eps-15 homology domain-containing protein 2 (EHD2, an ATPase) associated with the static population of PM caveolae. Recruitment to the PM involved ATP binding, interaction with anionic lipids, and oligomerization into large complexes (60-75S) via interaction of the EH domains with intrinsic NPF/KPF motifs. Hydrolysis of ATP was essential for binding of EHD2 complexes to caveolae. EHD2 was found to undergo dynamic exchange at caveolae, a process that depended on a functional ATPase cycle. Depletion of EHD2 by siRNA or expression of a dominant-negative mutant dramatically increased the fraction of mobile caveolar vesicles coming from the PM. Overexpression of EHD2, in turn, caused confinement of cholera toxin B in caveolae. The confining role of EHD2 relied on its capacity to link caveolae to actin filaments. Thus, EHD2 likely plays a key role in adjusting the balance between PM functions of stationary caveolae and the role of caveolae as vesicular carriers.     

IGF-IR signaling attenuates the age-related decline of diastolic cardiac function

Insulin-like growth factor (IGF-I) signaling has been implicated to play an important role in regulation of cardiac growth, hypertrophy, and contractile function and has been linked to the development of age-related congestive heart failure. Here, we address the question to what extent cardiomyocyte-specific IGF-I signaling is essential for maintenance of the structural and functional integrity of the adult murine heart. To investigate the effects of IGF-I signaling in the adult heart without confounding effects due to IGF-I overexpression or adaptation during embryonic and early postnatal development, we inactivated the IGF-I receptor (IGF-IR) by a 4-hydroxytamoxifen-inducible Cre recombinase in adult cardiac myocytes. Efficient inactivation of the IGF-IR (iCMIGF-IRKO) as assessed by Western analysis and real-time PCR went along with reduced IGF-I-dependent Akt and GSK3β phosphorylation. Functional analysis by conductance manometry and MRI revealed no functional alterations in young adult iCMIGF-IRKO mice (age 3 mo). However, when induced in aging mice (11 mo) diastolic cardiac function was depressed. To address the question whether insulin signaling might compensate for the defective IGF-IR signaling, we inactivated β-cells by streptozotocin. However, the diabetes-associated functional depression was similar in control and iCMIGF-IRKO mice. Similarly, analysis of the cardiac gene expression profile on 44K microarrays did not reveal activation of overt adaptive processes. Endogenous IGF-IR signaling is required for conservation of cardiac function of the aging heart, but not for the integrity of cardiac structure and function of young hearts.     

COSA-1 reveals robust homeostasis and separable licensing and reinforcement steps governing meiotic crossovers

Crossovers (COs) between homologous chromosomes ensure their faithful segregation during meiosis. We identify C.?elegans COSA-1, a cyclin-related protein conserved in metazoa, as a key component required to convert meiotic double-strand breaks (DSBs) into COs. During late meiotic prophase, COSA-1 localizes to foci that correspond to the single CO site on each homolog pair and indicate sites of eventual concentration of other conserved CO proteins. Chromosomes gain and lose competence to load CO proteins during meiotic progression, with competence to load COSA-1 requiring prior licensing. Our data further suggest a self-reinforcing mechanism maintaining CO designation. Modeling of a nonlinear dose-response relationship between IR-induced DSBs and COSA-1 foci reveals efficient conversion of DSBs into COs when DSBs are limiting and a robust capacity to limit cytologically differentiated CO sites when DSBs are in excess. COSA-1 foci serve as a unique live cell readout for investigating CO formation and CO interference.     

Carbohydrate Hydrolysis and Transport in the Extreme Thermoacidophile Sulfolobus solfataricus

Extremely thermoacidophilic microbes, such as Sulfolobus solfataricus, are strict chemoheterotrophs despite their geologic niche. To clarify their ecophysiology, the overlapping roles of endoglucanases and carbohydrate transporters were examined during growth on soluble cellodextrins as the sole carbon and energy source. Strain-specific differences in genome structure implied a unique role for one of three endogenous endoglucanases. Plasmid-based endoglucanase expression promoted the consumption of oligosaccharides, including cellohexaose (G6) through cellonanaose (G9). Protein transporters required for cellodextrin uptake were identified through mutagenesis and complementation of an ABC transporter cassette, including a putative oligosaccharide binding protein. In addition, ablation of the binding protein compromised growth on glucose and alpha-linked oligosaccharides while inactivation of a previously described glucose transporter had no apparent impact. These data demonstrate that S. solfataricus employs a redundant mechanism for soluble cellodextrin catabolism having both substrate uptake and extracytoplasmic hydrolytic components.     

Draft genome sequence of Leucobacter chromiiresistens, an extremely chromium-tolerant strain

Here we present the draft genome of Leucobacter chromiiresistens. This is the first genome sequence of an organism belonging to the genus Leucobacter. L. chromiiresistens was sequenced due to its capability to tolerate up to 300 mM Cr(VI) in the medium, which is so far a unique feature for microorganisms.     

Cell Wall Amidase AmiC1 Is Required for Cellular Communication and Heterocyst Development in the Cyanobacterium Anabaena PCC 7120 but Not for Filament Integrity

Filamentous cyanobacteria of the order Nostocales display typical properties of multicellular organisms. In response to nitrogen starvation, some vegetative cells differentiate into heterocysts, where fixation of N(2) takes place. Heterocysts provide a micro-oxic compartment to protect nitrogenase from the oxygen produced by the vegetative cells. Differentiation involves fundamental remodeling of the Gram-negative cell wall by deposition of a thick envelope and by formation of a neck-like structure at the contact site to the vegetative cells. Cell wall-hydrolyzing enzymes, like cell wall amidases, are involved in peptidoglycan maturation and turnover in unicellular bacteria. Recently, we showed that mutation of the amidase homologue amiC2 gene in Nostoc punctiforme ATCC 29133 distorts filament morphology and function. Here, we present the functional characterization of two amiC paralogues from Anabaena sp. strain PCC 7120. The amiC1 (alr0092) mutant was not able to differentiate heterocysts or to grow diazotrophically, whereas the amiC2 (alr0093) mutant did not show an altered phenotype under standard growth conditions. In agreement, fluorescence recovery after photobleaching (FRAP) studies showed a lack of cell-cell communication only in the AmiC1 mutant. Green fluorescent protein (GFP)-tagged AmiC1 was able to complement the mutant phenotype to wild-type properties. The protein localized in the septal regions of newly dividing cells and at the neck region of differentiating heterocysts. Upon nitrogen step-down, no mature heterocysts were developed in spite of ongoing heterocyst-specific gene expression. These results show the dependence of heterocyst development on amidase function and highlight a pivotal but so far underestimated cellular process, the remodeling of peptidoglycan, for the biology of filamentous cyanobacteria.     

Inhibition of NAMPT pathway by FK866 activates the function of p53 in HEK293T cells

Inactivation of p53 protein by endogenous and exogenous carcinogens is involved in the pathogenesis of different human malignancies. In cancer associated with SV-40 DNA tumor virus, p53 is considered to be non-functional mainly due to its interaction with the large T-antigen. Using the 293T cell line (HEK293 cells transformed with large T antigen) as a model, we provide evidence that p53 is one of the critical downstream targets involved in FK866-mediated killing of 293T cells. A reduced rate of apoptosis and an increased number of cells in S-phase was accompanied after knockdown of p53 in these cells. Inhibition of NAMPT by FK866, or inhibition of SIRT by nicotinamide decreased proliferation and triggered death of 293T cells involving the p53 acetylation pathway. Additionally, knockdown of p53 attenuated the effect of FK866 on cell proliferation, apoptosis, and cell cycle arrest. The data presented here shed light on two important facts: (1) that p53 in 293T cells is active in the presence of FK866, an inhibitor of NAMPT pathway; (2) the apoptosis induced by FK866 in 293T cells is associated with increased acetylation of p53 at Lys382, which is required for the functional activity of p53.     

Polyphenol metabolism of developing apple skin of a scab resistant and a susceptible apple cultivar

During fruit development, the concentration of main polyphenols (flavonols, flavanols, dihydrochalcones, hydroxycinnamic acids, anthocyanins) and the activities of related enzymes (phenylalanine ammonia lyase, chalcone synthase/chalcone isomerase, flavanone 3-hydroxylase, dihydroflavonol 4-reductase, flavonol synthase, peroxidase) were monitored in apple (Malus domestica Borkh.). The seasonal survey was performed at five different sampling dates and included the healthy peel of the resistant cultivar ‘Florina’ and healthy peel, scab symptomatic spot and the tissue around the infected spot of the susceptible cultivar ‘Golden Delicious’. From all enzymes tested, chalcone synthase/chalcone isomerase had the highest activity in both cultivars, while phenylalanine ammonia lyase had the lowest. The healthy peels of the susceptible and the resistant cultivar did not show differences in the accumulation of the main polyphenol groups present in the apple skin. However, in the resistant cultivar ‘Florina’, an increase of polyphenol enzyme activities could be observed in late stages of fruit development, which seems to be related to the anthocyanin accumulation in ripe fruits. Significant differences in the polyphenol metabolism were observed in the three different tissues of the susceptible cultivar ‘Golden Delicious’. Increased concentrations of hydroxycinnamic acids, dihydrochalcones and flavan-3-ols were found in the scab symptomatic spots and surrounding tissues. Phenylalanine ammonia-lyase, dihydroflavonol 4-reductase, flavanone 3-hydroxylase and peroxidase showed higher activities in the scab symptomatic spot compared to other analysed tissues, whereas the activities of other enzymes remained unchanged. Highest induction of polyphenol accumulation after scab infection was observed in early developmental stages, whereas enzyme activities were increased in later stages.     

Scram1 is a modifier of spinal cord resistance for astrocytoma on mouse Chr 5

Tumor location can profoundly affect morbidity and patient prognosis, even for the same tumor type. Very little is known about whether tumor location is determined stochastically or whether genetic risk factors can affect where tumors arise within an organ system. We have taken advantage of the Nf1-/+;Trp53-/+cis mouse model of astrocytoma/glioblastoma to map genetic loci affecting whether astrocytomas are found in the spinal cord. We identify a locus on distal Chr 5, termed Scram1 for spinal cord resistance to astrocytoma modifier 1, with a LOD score of 5.0 and a genome-wide significance of P?相似文献