Transcriptome changes and cAMP oscillations in an archaeal cell cycle

Background The cell cycle of all organisms includes mass increase by a factor of two, replication of the genetic material, segregation of the genome to different parts of the cell, and cell division into two daughter cells. It is tightly regulated and typically includes cell cycle-specific oscillations of the levels of transcripts, proteins, protein modifications, and signaling molecules. Until now cell cycle-specific transcriptome changes have been described for four eukaryotic species ranging from yeast to human, but only for two prokaryotic species. Similarly, oscillations of small signaling molecules have been identified in very few eukaryotic species, but not in any prokaryote. Results A synchronization procedure for the archaeon Halobacterium salinarum was optimized, so that nearly 100% of all cells divide in a time interval that is 1/4th of the generation time of exponentially growing cells. The method was used to characterize cell cycle-dependent transcriptome changes using a genome-wide DNA microarray. The transcript levels of 87 genes were found to be cell cycle-regulated, corresponding to 3% of all genes. They could be clustered into seven groups with different transcript level profiles. Cluster-specific sequence motifs were detected around the start of the genes that are predicted to be involved in cell cycle-specific transcriptional regulation. Notably, many cell cycle genes that have oscillating transcript levels in eukaryotes are not regulated on the transcriptional level in H. salinarum. Synchronized cultures were also used to identify putative small signaling molecules. H. salinarum was found to contain a basal cAMP concentration of 200 uM, considerably higher than that of yeast. The cAMP concentration is shortly induced directly prior to and after cell division, and thus cAMP probably is an important signal for cell cycle progression. Conclusions The analysis of cell cycle-specific transcriptome changes of H. salinarum allowed to identify a strategy of transcript level regulation that is different from all previously characterized species. The transcript levels of only 3% of all genes are regulated, a fraction that is considerably lower than has been reported for four eukaryotic species (6% - 28%) and for the bacterium C. crescentus (19%). It was shown that cAMP is present in significant concentrations in an archaeon, and the phylogenetic profile of the adenylate cyclase indicates that this signaling molecule is widely distributed in archaea. The occurrence of cell cycle-dependent oscillations of the cAMP concentration in an archaeon and in several eukaryotic species indicates that cAMP level changes might be a phylogenetically old signal for cell cycle progression

Quantification of ploidy in proteobacteria revealed the existence of monoploid, (mero-)oligoploid and polyploid species

Bacteria are generally assumed to be monoploid (haploid). This assumption is mainly based on generalization of the results obtained with the most intensely studied model bacterium, Escherichia coli (a gamma-proteobacterium), which is monoploid during very slow growth. However, several species of proteobacteria are oligo- or polyploid, respectively. To get a better overview of the distribution of ploidy levels, genome copy numbers were quantified in four species of three different groups of proteobacteria. A recently developed Real Time PCR approach, which had been used to determine the ploidy levels of halophilic archaea, was optimized for the quantification of genome copy numbers of bacteria. Slow-growing (doubling time 103 minutes) and fast-growing (doubling time 25 minutes) E. coli cultures were used as a positive control. The copy numbers of the origin and terminus region of the chromosome were determined and the results were in excellent agreement with published data. The approach was also used to determine the ploidy levels of Caulobacter crescentus (an alpha-proteobacterium) and Wolinella succinogenes (an epsilon-proteobacterium), both of which are monoploid. In contrast, Pseudomonas putida (a gamma-proteobacterium) contains 20 genome copies and is thus polyploid. A survey of the proteobacteria with experimentally-determined genome copy numbers revealed that only three to four of 11 species are monoploid and thus monoploidy is not typical for proteobacteria. The ploidy level is not conserved within the groups of proteobacteria, and there are no obvious correlations between the ploidy levels with other parameters like genome size, optimal growth temperature or mode of life

EU Smart City Lighthouse Projects between Top-Down Strategies and Local Legitimation: The Case of Hamburg

The concept of the smart city has become increasingly popular in recent years and a large number of cities globally follow smart city strategies. By awarding subsidies in the Horizon 2020 programme, the European Union (EU) has taken on an influential role in how smart city projects are conceived and implemented in European municipalities. Using the example of the smart city pilot project mySMARTLife in Hamburg, the purpose of this article is to examine the area of tension between strategically pursuing own objectives and adjustment to external provisions of the EU funding framework. In a qualitative single case study, the article analyses what implications the project mySMARTLife has on urban development practice and local governance arrangements in Hamburg. Examining current literature on smart cities from the perspective of multi-level governance and presenting the current state of research dealing with EU smart city projects, a theoretical framework is developed. The analysis reveals that, due to the EU funding framework, precise project contents are contractually defined at an early stage when local stakeholders have limited involvement in this process. Furthermore, the analysis shows that the EU smart city funding in the project mySMARTLife is more limited to the implementation of individual interventions than to a comprehensive smart city strategy. As a result, this article considers EU-funded smart city initiatives as experimental fields that enable cities to gain experiences that can be incorporated into local strategic development objectives

Smart City Governance in Practice – the Example of mySMARTLife Project in Hamburg

The concept of the “smart city” has become increasingly popular in recent years and a large number of cities globally follow smart city strategies. The Free and Hanseatic City of Hamburg has also adopted the aim to become a smart city. With the project mySMARTLife under the framework of the European Union’s Horizon 2020 programme Hamburg received the status of a smart city lighthouse project, and local actors from politics, administration, private economy, and civil society began collaborating to transform the district of Hamburg-Bergedorf towards a smart city. By experimenting with innovative technologies on-site, the project follows an applied and implementation-oriented approach. The purpose of this paper is to analyse under which institutional framework conditions the smart city project mySMARTLife is implemented in Hamburg. In particular, it is examined which forms of governance characterise the implementation process of the project in the district of Bergedorf. Based on a study of current literature on smart city governance an analytical framework will be developed and applied on the example of the mySMARTLife project. The empirical findings of the paper elaborate institutional and actor-related challenges of putting a smart city initiative into practice and identify which factors promote and hinder these intentions. Being part of the mySMARTLife consortium the authors gathered knowledge from the inside perspective of the project as well as from interviews with relevant stakeholders

TESTS ON 10.9 BOLTS UNDER COMBINED TENSION AND SHEAR

Prior investigations of the load-bearing capacity of bolts during fire have shown differing behaviour between bolts that have been loaded by shear or by tensile loads. A combination of the two loads has not yet been examined under fire conditions. This paper describes a series of tests on high-strength bolts of property class 10.9 both during and after fire under a combined shear and tensile load

TESTS ON 10.9 BOLTS UNDER COMBINED TENSION AND SHEAR DURING AND AFTER FIRE

Prior investigations of the load bearing capacity of bolts during fire have shown differing behaviour between bolts that were loaded by shear or by tensile loads. The interaction of the two loads has not yet been examined under fire conditions. This paper describes a preliminary test series on the post-fire performance of high-strength bolts of the property class 10.9 under combined tension and shear. The results show that how the bolt is loaded influences the load bearing capacity. It is assumed that this is also true at elevated temperatures. Further, atest set-up for experiments at elevated temperatures and a more detailed test series on the post-fire performance under combined tension and shear is presented

a methodology for understanding path dependence and path creation

Although an increasing number of studies of technological, institutional and organizational change refer to the concepts of path dependence and path creation, few attempts have been made to consider these concepts explicitly in their methodological accounts. This paper addresses this gap and contributes to the literature by developing a comprehensive methodology that originates from the concepts of path dependence and path creation – path constitution analysis (PCA) – and allows for the integration of multi-actor constellations on multiple levels of analysis within a process perspective. Based upon a longitudinal case study in the field of semiconductors, we illustrate PCA ‘in action’ as a template for other researchers and critically examine its adequacy. We conclude with implications for further path-oriented inquiries

Stress and perceived partnership quality of parents to children with autism spectrum disorder: a random intercept cross-lagged panel approach

Parents to children with autism spectrum disorder (ASD) experience increased levels of stress compared to other parents. However, it has been difficult to clarify how this parental stress affects the quality of parent partnership, and how stress and partnership quality predict each other longitudinally. Stress and perceived partnership quality of 160 parents to children with ASD who started therapy in an autism therapy center were surveyed at three measurement points. To find out about within-person effects, a random intercept cross-lagged panel model was used. The results showed that a decrease in stress level over the course of therapy led to a decrease in partnership quality. The theory of the parenting alliance can be used to explain these results: coping with demands in parenthood during stressful times seems to tie parents together and strengthen their relationship. Implications for therapeutic practice, such as the use of marriage and family counselors to support the parents’ partnership during their child's autism therapy are presented

Estimating Absolute Configurational Entropies of Macromolecules: The Minimally Coupled Subspace Approach

We develop a general minimally coupled subspace approach (MCSA) to compute absolute entropies of macromolecules, such as proteins, from computer generated canonical ensembles. Our approach overcomes limitations of current estimates such as the quasi-harmonic approximation which neglects non-linear and higher-order correlations as well as multi-minima characteristics of protein energy landscapes. Here, Full Correlation Analysis, adaptive kernel density estimation, and mutual information expansions are combined and high accuracy is demonstrated for a number of test systems ranging from alkanes to a 14 residue peptide. We further computed the configurational entropy for the full 67-residue cofactor of the TATA box binding protein illustrating that MCSA yields improved results also for large macromolecular systems