An essential role for katanin p80 and microtubule severing in male gamete production

Katanin is an evolutionarily conserved microtubule-severing complex implicated in multiple aspects of microtubule dynamics. Katanin consists of a p60 severing enzyme and a p80 regulatory subunit. The p80 subunit is thought to regulate complex targeting and severing activity, but its precise role remains elusive. In lower-order species, the katanin complex has been shown to modulate mitotic and female meiotic spindle dynamics and flagella development. The in vivo function of katanin p80 in mammals is unknown. Here we show that katanin p80 is essential for male fertility. Specifically, through an analysis of a mouse loss-of-function allele (the Taily line), we demonstrate that katanin p80, most likely in association with p60, has an essential role in male meiotic spindle assembly and dissolution and the removal of midbody microtubules and, thus, cytokinesis. Katanin p80 also controls the formation, function, and dissolution of a microtubule structure intimately involved in defining sperm head shaping and sperm tail formation, the manchette, and plays a role in the formation of axoneme microtubules. Perturbed katanin p80 function, as evidenced in the Taily mouse, results in male sterility characterized by decreased sperm production, sperm with abnormal head shape, and a virtual absence of progressive motility. Collectively these data demonstrate that katanin p80 serves an essential and evolutionarily conserved role in several aspects of male germ cell development.     

Troglitazone acutely inhibits protein synthesis in endothelial cells via a novel mechanism involving protein phosphatase 2A-dependent p70 S6 kinase inhibition

Thiazolidinediones (TZDs), synthetic peroxisome proliferator-activated receptor (PPAR) ligands, have been implicated in the inhibition of protein synthesis in a variety of cells, but the underlying mechanisms remain obscure. We report that troglitazone, the first TZD drug, acutely inhibited protein synthesis by decreasing p70 S6 kinase (p70S6K) activity in bovine aortic endothelial cells (BAEC). This inhibition was not accompanied by decreased phosphorylation status or in vitro kinase activity of mammalian target of rapamycin (mTOR). Furthermore, cotreatment with rapamycin, a specific mTOR inhibitor, and troglitazone additively inhibited both p70S6K activity and protein synthesis, suggesting that the inhibitory effects of troglitazone are not mediated by mTOR. Overexpression of the wild-type p70S6K gene significantly reversed the troglitazone-induced inhibition of protein synthesis, indicating an important role of p70S6K. Okadaic acid, a protein phosphatase 2A (PP2A) inhibitor, partially reversed the troglitazone-induced inhibition of p70S6K activity and protein synthesis. Although troglitazone did not alter total cellular PP2A activity, it increased the physical association between p70S6K and PP2A, suggesting an underlying molecular mechanism. GW9662, a PPAR antagonist, did not alter any of the observed inhibitory effects. Finally, we also found that the mTOR-independent inhibitory mechanism of troglitazone holds for the TZDs ciglitazone, pioglitazone, and rosiglitazone, in BAEC and other types of endothelial cells tested. In conclusion, our data demonstrate for the first time that troglitazone (and perhaps other TZDs) acutely decreases p70S6K activity through a PP2A-dependent mechanism that is independent of mTOR and PPAR, leading to the inhibition of protein synthesis in endothelial cells. protein synthesis     

Assessment of environmental stress by the micronucleus and comet assays on Limnoperna fortunei exposed to Guaíba hydrographic region samples (Brazil) under laboratory conditions

The Guaíba Basin is a source of drinking water for Porto Alegre (RS, Brazil). The water from this basin receives industrial, urban, and rural waste from many sources. The mussel species Limnoperna fortunei was chosen based on population data, distribution, and sensitivity. Previous tests with comet assay and micronuclei frequency in this freshwater mussel have shown to be successful in biomonitoring studies. The aim of this study was to evaluate the genotoxic contamination of the Guaíba Lake Hydrographic Region, through the determination of damage by the micronuclei and comet assays in L. fortunei (golden mussel). Nine sampling sites were evaluated in three different seasons: five sites in the mouths of the main rivers that flow into Guaíba lake; one site at the mouth of a stream; one major site of sewage discharge; two sites at Guaíba lake, near a sewage discharge; and the control site in a preservation area. DNA damage was detected by the single cell gel assay, as well as the frequency of micronuclei in hemocytes of mussels exposed under laboratory conditions for 7 days to water and sediment samples. Significant results were found in different seasons in almost all sampling sites (P<0.05, ANOVA Dunnet's test). Most of the positive results were found in samples affected mainly by urban effluents. It was possible to observe that there was a weak relation between mutagenic and genotoxic responses and mussels inorganic elements contents. Seasonal variation was observed at different sampling sites, but always indicating a huge contamination near urban sewage discharge. These results are consistent with previous studies, allowing us to infer that urban contamination is the biggest problem in this region. It is also possible to infer that L. fortunei is a good sentinel organism for the Guaíba Basin.     

Southern Cone Initiative for the elimination of domestic populations of Triatoma infestans and the interruption of transfusional Chagas disease. Historical aspects, present situation, and perspectives

Created in 1991 by the governments of Argentina, Bolivia, Brazil, Chile, Paraguay, and Uruguay, the Southern Cone Initiative (SCI) has been extremely important for Chagas disease control in this region. Its basic objective was to reach the interruption of this disease, chiefly by means of the elimination of the principal vector Triatoma infestans and by the selection of safe donors in the regional blood banks. After a summarized historic of SCI, the text shows the advance of technical and operative activities, emphasizing some factors for the initiative success, as well as some difficulties and constraints. The future of SCI will depend of the continuity of the actions and of political priority. Scientific community has been highly responsible for this initiative and its maintenance. At the side of this, national and international efforts must be involved and reinforced to assure the accomplishment of the final targets of SCI. Very specially, the Pan American Health Organization has cooperated with the Initiative in all its moments and activities,being the most important catalytic and technical factor for SCI success.     

Mitochondrial introgressions into the nuclear genome of the domestic cat

Translocation of mtDNA into the nuclear genome, also referred to as numt, was first reported in the domestic cat (Felis catus) by Lopez et al. (1994). The Lopez-numt consisted of a translocation of 7.9 kbp of mtDNA that inserted into the domestic cat chromosome D2 around 1.8 million years ago. More than a decade later, the release of the domestic cat whole-genome shotgun sequences (1.9x coverage) provides the resource to obtain more comprehensive insight into the extent of mtDNA transfer over time in the domestic cat genome. MegaBLAST searches revealed that the cat genome harbors a wide variety of numts (298 320 bp), one-third of which likely correspond to the Lopez-numt tandem repeat, whereas the remaining numts are probably derived from multiple independent insertions, which in some cases were followed by segmental duplication after insertion in the nucleus. Numts were detected across most cat chromosomes, but the number of numts assigned to chromosomes is underestimated due to the relatively high number of numt sequences with insufficient flanking sequence to map. The catalog of cat numts provides a valuable resource for future studies in Felidae species, including its use as a tool to avoid numt contaminations that may confound population genetics and phylogenetic studies.     

Biochemical characterization of ecto-nucleotide pyrophosphatase/phosphodiesterase (E-NPP, E.C. 3.1.4.1) from rat heart left ventricle

In the present study we investigate the biochemical properties of the members of NPP family in synaptosomes prepared from rat heart left ventricles. Using p-nitrophenyl-5′-thymidine monophosphate (p-Nph-5′-TMP) as substrate for E-NPPs in rat cardiac synaptosomes, we observed an alkaline pH dependence, divalent cation dependence and the K _M value corresponded to 91.42 ± 13.97 μM and the maximal velocity (V _max ) value calculated was 63.79 ± 3.59 nmol p-nitrophenol released/min/mg of protein (mean ± SD, n = 4). Levamisole (1 mM), was ineffective as inhibitor of p-Nph-5′-TMP hydrolysis in pH 8.9 (optimum pH for the enzyme characterized). Suramin (0.25 mM) strongly reduced the hydrolysis of p-Nph-5′-TMP by about 46%. Sodium azide (10 and 20 mM) and gadolinium chloride (0.3 and 0.5 mM), E-NTPases inhibitors, had no effects on p-Nph-5′-TMP hydrolysis. RT-PCR analysis of left ventricle demonstrated the expression of NPP2 and NPP3 enzymes, but excluded the presence of NPP1 member. By quantitative real-time PCR we identified the NPP3 as the enzyme with the highest expression in rat left ventricle. The demonstration of the presence of the E-NPP family in cardiac system, suggest that these enzymes could contribute with the fine-tuning control of the nucleotide levels at the nerve terminal endings of left ventricles that are involved in several cardiac pathologies.     

A neurobiological model of the recovery strategies from perturbed walking

This paper proposes a human mimetic neuro-musculo-skeletal model to simulate the recovery reactions from perturbations during walking. The computational model incorporates nonlinear viscoelastic muscular mechanics, supraspinal control of the center-of-mass, spinal pattern generator including muscle synergy network, spinal reflexes, and long-loop reflexes. Especially the long-loop reflexes specify recovery strategies based on the experimental observations [Schillings, A.M., van Wezel, B.M.H., Mulder, T.H., Duysen, J., 2000. Muscular responses and movement strategies during stumbling over obstacles. J. Neurophysiol. 83, 2093–2102; Eng, J.J., Winter, D.A., Patla, A.E., 1994. Strategies for recovery from a trip in early and late swing during human walking. Exp. Brain Res. 102, 339–349]. The model demonstrates two typical recovery strategies, i.e., elevating and lowering strategies against pulling over a swing leg. Sensed perturbation triggers a simple tonic pulse from the cortex. Depending on the swing phase, the tonic pulse activates a different compound of muscles over lower limbs. The compound induces corresponding recovery strategies. The reproduction of principal recovery behaviors may support the model's proposed functional and/or anatomical correspondence.