A seesaw model for intermolecular gating in the kinesin motor protein

Recent structural observations of kinesin-1, the founding member of the kinesin group of motor proteins, have led to substantial gains in our understanding of this molecular machine. Kinesin-1, similar to many kinesin family members, assembles to form homodimers that use alternating ATPase cycles of the catalytic motor domains, or “heads”, to proceed unidirectionally along its partner filament (the microtubule) via a hand-over-hand mechanism. Cryo-electron microscopy has now revealed 8-Å resolution, 3D reconstructions of kinesin-1•microtubule complexes for all three of this motor’s principal nucleotide-state intermediates (ADP-bound, no-nucleotide, and ATP analog), the first time filament co-complexes of any cytoskeletal motor have been visualized at this level of detail. These reconstructions comprehensively describe nucleotide-dependent changes in a monomeric head domain at the secondary structure level, and this information has been combined with atomic-resolution crystallography data to synthesize an atomic-level "seesaw" mechanism describing how microtubules activate kinesin’s ATP-sensing machinery. The new structural information revises or replaces key details of earlier models of kinesin’s ATPase cycle that were based principally on crystal structures of free kinesin, and demonstrates that high-resolution characterization of the kinesin–microtubule complex is essential for understanding the structural basis of the cycle. I discuss the broader implications of the seesaw mechanism within the cycle of a fully functional kinesin dimer and show how the seesaw can account for two types of "gating" that keep the ATPase cycles of the two heads out of sync during processive movement

Proteomic Analysis of Pathways Involved in Estrogen-Induced Growth and Apoptosis of Breast Cancer Cells

Estrogen is a known growth promoter for estrogen receptor (ER)-positive breast cancer cells. Paradoxically, in breast cancer cells that have been chronically deprived of estrogen stimulation, re-introduction of the hormone can induce apoptosis.Here, we sought to identify signaling networks that are triggered by estradiol (E2) in isogenic MCF-7 breast cancer cells that undergo apoptosis (MCF-7:5C) versus cells that proliferate upon exposure to E2 (MCF-7). The nuclear receptor co-activator AIB1 (Amplified in Breast Cancer-1) is known to be rate-limiting for E2-induced cell survival responses in MCF-7 cells and was found here to also be required for the induction of apoptosis by E2 in the MCF-7:5C cells. Proteins that interact with AIB1 as well as complexes that contain tyrosine phosphorylated proteins were isolated by immunoprecipitation and identified by mass spectrometry (MS) at baseline and after a brief exposure to E2 for two hours. Bioinformatic network analyses of the identified protein interactions were then used to analyze E2 signaling pathways that trigger apoptosis versus survival. Comparison of MS data with a computationally-predicted AIB1 interaction network showed that 26 proteins identified in this study are within this network, and are involved in signal transduction, transcription, cell cycle regulation and protein degradation.G-protein-coupled receptors, PI3 kinase, Wnt and Notch signaling pathways were most strongly associated with E2-induced proliferation or apoptosis and are integrated here into a global AIB1 signaling network that controls qualitatively distinct responses to estrogen

Unfertilized Xenopus Eggs Die by Bad-Dependent Apoptosis under the Control of Cdk1 and JNK

Ovulated eggs possess maternal apoptotic execution machinery that is inhibited for a limited time. The fertilized eggs switch off this time bomb whereas aged unfertilized eggs and parthenogenetically activated eggs fail to stop the timer and die. To investigate the nature of the molecular clock that triggers the egg decision of committing suicide, we introduce here Xenopus eggs as an in vivo system for studying the death of unfertilized eggs. We report that after ovulation, a number of eggs remains in the female body where they die by apoptosis. Similarly, ovulated unfertilized eggs recovered in the external medium die within 72 h. We showed that the death process depends on both cytochrome c release and caspase activation. The apoptotic machinery is turned on during meiotic maturation, before fertilization. The death pathway is independent of ERK but relies on activating Bad phosphorylation through the control of both kinases Cdk1 and JNK. In conclusion, the default fate of an unfertilized Xenopus egg is to die by a mitochondrial dependent apoptosis activated during meiotic maturation

Ocean acidification influences the biomass and diversity of reef-associated turf algal communities

Projected ocean acidification will have a detrimental impact on coral reef ecosystems, where fleshy algae are expected to replace corals. Of particular importance to reef ecosystems are fleshy turf algal communities, which have the potential to overgrow corals; few studies have investigated the community structure and diversity of turfs to climate change. Here, we assessed the response of reef-associated turf algal communities from the Great Barrier Reef, Australia to three levels of ocean acidification. Biomass of turf communities was positively affected by increases in carbon dioxide (CO2), where turf communities grown under high CO2 had the greatest biomass. No effect of CO2 was found on mean turf organic content or genus richness. By contrast, turf community evenness and diversity (H′) increased under medium and high CO2 treatments. Our results indicate that increased turf growth under high CO2 will aid the overall expansion and growth of fleshy macroalgae in coral reef ecosystems, as opportunistic algae may have an advantage over other reef-associated species. Changes in turf community diversity will help provide insight into how macroalgal communities may be structured in the future, highlighting genera primed to take advantage of the changes in ocean chemistry associated with ocean acidification.No Full Tex

Identification of a strong binding site for kinesin on the microtubule using mutant analysis of tubulin

The kinesin-binding site on the microtubule has not been identified because of the technical difficulties involved in the mutant analyses of tubulin. Exploiting the budding yeast expression system, we succeeded in replacing the negatively charged residues in the α-helix 12 of β-tubulin with alanine and analyzed their effect on kinesin–microtubule interaction in vitro. The microtubule gliding assay showed that the affinity of the microtubules for kinesin was significantly reduced in E410A, D417A, and E421A, but not in E412A mutant. The unbinding force measurement revealed that in the former three mutants, the kinesin–microtubule interaction in the adenosine 5′-[β,γ-imido]triphosphate state (AMP-PNP state) became less stable when a load was imposed towards the microtubule minus end. In parallel with this decreased stability, the stall force of kinesin was reduced. Our results implicate residues E410, D417, and E421 as crucial for the kinesin–microtubule interaction in the strong binding state, thereby governing the size of kinesin stall force

Long-term impacts of season of grazing on soil carbon sequestration and selected soil properties in the arid Eastern Cape, South Africa

BACKGROUND AND AIMS : The karoo biomes of South Africa are major feed resources, yet soil nutrient depletion and degradation is a major problem. The objective of this study was to assess impacts of long-term (>75 years) grazing during spring (SPG), summer (SUG), winter (WG) and exclosure (non-grazed control) treatments on soil nutrients, penetration resistance and infiltration tests. METHODS : A soil sampling campaign was carried out to collect soil to a depth of 60 cm to analyse bulk density, soil physical and chemical parameters as well as soil compaction and infiltration. RESULTS : Generally, grazing treatments reduced soil organic C (SOC) stocks and C:N ratios, and modified soil properties. There was higher SOC stock (0.128 Mg ha-1 yr-1) in the exclosure than in the SPG (0.096 Mg ha-1 yr- 1), SUG (0.099 Mg ha-1 yr-1) and WG (0.105 Mg ha-1 yr-1). The C:N ratios exhibited similar pattern to that of C. From the grazing treatments, the WG demonstrated 7 to 10% additional SOC stock over the SPG and SUG, respectively. CONCLUSIONS : Short period animal exclusion could be an option to be considered to improve plant nutrients in sandy soils of South Africa. However, this may require a policy environment which supports stock exclusion from such areas vulnerable to land degradation, nutrient and C losses by grazing-induced vegetation and landscape changes.Department of Science and Technology University of Pretoria) and the European Communities, 7th framework program under the grant agreement No. 266018, ANIMALCHANGE project.http://link.springer.com/journal/111042016-12-31hb201