Suchergebnisse

Batrachochytrium dendrobatidis (Bd) is a fungal pathogen which causes the emerging infectious disease chytridiomycosis. Bd presents low host specificity and threatens amphibians worldwide, thus systematic inventory is the key in order to detect and mitigate the effects of the disease. Extensive data collection was conducted in Hungary in 2009-2015 from fourteen different areas. Combined data – recent field sampling on sixteen taxa and the examination of archived Bombina spp. specimens – from 1360 individuals were analysed with qPCR. Two sentinel taxa, Bombina variegata and the members of the Pelophylax esculentus complex were marked to monitor the occurrence of Bd in two core areas (Bakony Mts and Hortobágy National Park, respectively) of sampling. Climatic variables were also examined in core areas to test their effect on prevalence and infection intensity. Among the sixteen sampled amphibian taxa seven tested positive for Bd and the overall prevalence in Hungary was 7.46%. Among the ethanol-fixed Bombina spp. individuals Bd was not detected. In the first core area (Bakony Mts) the overall prevalence in B. variegata was 10.32% and juvenile individuals showed significantly higher prevalence than adults. On the other hand there was a significant negative relationship between infection prevalence and monthly mean air temperature. Finally, in the other core area (Hortobágy National Park) the overall prevalence in P. esculentus complex was 13.00%, and no differences were found in prevalence or infection intensity between sexes, sampling years or age classes. ; During the project JV was supported by the Hungarian Scientific Research Fund (OTKA K77841) and by the Bolyai János Research Scholarship of the Hungarian Academy of Sciences (BO/00579/14/8). DH was supported by the European Union and co-financed by the European Social Fund through the Social Renewal Operational Programme under the projects TÁMOP–4.2.2/B–10/1–2010–0024 and SROP-4.2.2.B-15/1/KONV-2015-0001. AF was sup-ported by the Hungarian National Research, Develop-ment and Innovation Office (OTKA grant no. K112527). Research permit was issued by the National Inspectorate of Environment, Nature Conservation and Water Man-agement (14/3535/2/2010) and the Tisza Region Inspec-torate of Environment, Nature Conservation and Water Management (4633/OH/2012). ; Peer Reviewed

mRNA export from the nucleus depends on the ATPase activity of the DEAD-box protein Dbp5/DDX19. Although Dbp5 has measurable ATPase activity alone, several regulatory factors (e.g., RNA, nucleoporin proteins, and the endogenous small molecule InsP6) modulate catalytic activity in vitro and in vivo to facilitate mRNA export. An analysis of the intrinsic and regulator-activated Dbp5 ATPase cycle is necessary to define how these factors control Dbp5 and mRNA export. Here, we report a kinetic and equilibrium analysis of the Saccharomyces cerevisiae Dbp5 ATPase cycle, including the influence of RNA on Dbp5 activity. These data show that ATP binds Dbp5 weakly in rapid equilibrium with a binding affinity (KT~4 mM) comparable to the KM for steady-state cycling, while ADP binds an order of magnitude more tightly (KD~0.4 mM). The overall intrinsic steady-state cycling rate constant (kcat) is limited by slow, near-irreversible ATP hydrolysis and even slower subsequent phosphate release. RNA increases kcat and rate-limiting Pi release 20-fold, although Pi release continues to limit steady-state cycling in the presence of RNA, in conjunction with RNA binding. Together, this work identifies RNA binding and Pi release as important biochemical transitions within the Dbp5 ATPase cycle and provides a framework for investigating the means by which Dbp5 and mRNA export is modulated by regulatory factors. ; E.V.W. is supported by National Science Foundation Graduate Research Fellowship No. DGE-1122492 and J.V. is supported by an Alberta Innovates Health Solutions Postdoctoral Fellowship. M.M. and Y.M. were supported by a Senior Research Fellowship from the Wellcome Trust (101908/Z/13/Z) and by National Institutes of Health grant R01 GM102869. Research support for B.M. was provided by the Natural Sciences and Engineering Research Council of Canada (RGPIN 435380), Canada Foundation for Innovation (31271), Government of Alberta Research Capacity Program, and Canada Research Chairs program.

mRNA export from the nucleus depends on the ATPase activity of the DEAD-box protein Dbp5/DDX19. Although Dbp5 has measurable ATPase activity alone, several regulatory factors (e.g., RNA, nucleoporin proteins, and the endogenous small molecule InsP6) modulate catalytic activity in vitro and in vivo to facilitate mRNA export. An analysis of the intrinsic and regulator-activated Dbp5 ATPase cycle is necessary to define how these factors control Dbp5 and mRNA export. Here, we report a kinetic and equilibrium analysis of the Saccharomyces cerevisiae Dbp5 ATPase cycle, including the influence of RNA on Dbp5 activity. These data show that ATP binds Dbp5 weakly in rapid equilibrium with a binding affinity (KT~4 mM) comparable to the KM for steady-state cycling, while ADP binds an order of magnitude more tightly (KD~0.4 mM). The overall intrinsic steady-state cycling rate constant (kcat) is limited by slow, near-irreversible ATP hydrolysis and even slower subsequent phosphate release. RNA increases kcat and rate-limiting Pi release 20-fold, although Pi release continues to limit steady-state cycling in the presence of RNA, in conjunction with RNA binding. Together, this work identifies RNA binding and Pi release as important biochemical transitions within the Dbp5 ATPase cycle and provides a framework for investigating the means by which Dbp5 and mRNA export is modulated by regulatory factors. ; E.V.W. is supported by National Science Foundation Graduate Research Fellowship No. DGE-1122492 and J.V. is supported by an Alberta Innovates Health Solutions Postdoctoral Fellowship. M.M. and Y.M. were supported by a Senior Research Fellowship from the Wellcome Trust (101908/Z/13/Z) and by National Institutes of Health grant R01 GM102869. Research support for B.M. was provided by the Natural Sciences and Engineering Research Council of Canada (RGPIN 435380), Canada Foundation for Innovation (31271), Government of Alberta Research Capacity Program, and Canada Research Chairs program. ; This is the final version of the article. It first appeared from Elsevier via https://doi.org/10.1016/j.jmb.2015.12.018