Gene regulation by CcpA and catabolite repression explored by RNA-Seq in Streptococcus mutans

A bacterial transcriptome of the primary etiological agent of human dental caries, Streptococcus mutans, is described here using deep RNA sequencing. Differential expression profiles of the transcriptome in the context of carbohydrate source, and of the presence or absence of the catabolite control protein CcpA, revealed good agreement with previously-published DNA microarrays. In addition, RNA-seq considerably expanded the repertoire of DNA sequences that showed statistically-significant changes in expression as a function of the presence of CcpA and growth carbohydrate. Novel mRNAs and small RNAs were identified, some of which were differentially expressed in conditions tested in this study, suggesting that the function of the S. mutans CcpA protein and the influence of carbohydrate sources has a more substantial impact on gene regulation than previously appreciated. Likewise, the data reveal that the mechanisms underlying prioritization of carbohydrate utilization are more diverse than what is currently understood. Collectively, this study demonstrates the validity of RNA-seq as a potentially more-powerful alternative to DNA microarrays in studying gene regulation in S. mutans because of the capacity of this approach to yield a more precise landscape of transcriptomic changes in response to specific mutations and growth conditions

Genetic Analysis of the Functions and Interactions of Components of the LevQRST Signal Transduction Complex of Streptococcus mutans

Transcription of the genes for a fructan hydrolase (fruA) and a fructose/mannose sugar:phosphotransferase permease (levDEFG) in Streptococcus mutans is activated by a four-component regulatory system consisting of a histidine kinase (LevS), a response regulator (LevR) and two carbohydrate-binding proteins (LevQT). The expression of the fruA and levD operons was at baseline in a levQ mutant and substantially decreased in a levT null mutant, with lower expression with the cognate inducers fructose or mannose, but slightly higher expression in glucose or galactose. A strain expressing levQ with two point mutations (E170A/F292S) did not require inducers to activate gene expression and displayed altered levD expression when growing on various carbohydrates, including cellobiose. Linker-scanning (LS) mutagenesis was used to generate three libraries of mutants of levQ, levS and levT that displayed various levels of altered substrate specificity and of fruA/levD gene expression. The data support that LevQ and LevT are intimately involved in the sensing of carbohydrate signals, and that LevQ appears to be required for the integrity of the signal transduction complex, apparently by interacting with the sensor kinase LevS

Script development as a ‘wicked problem’

© 2018 Intellect Ltd Article. Both a process and a set of products, influenced by policy as well as people, and incorporating objective agendas at the same time as subjective experiences, script development is a core practice within the screen industry –yet one that is hard to pin down and, to some extent, define. From an academic research perspective, we might say that script development is a ‘wicked problem’ precisely because of these complex and often contradictory aspects. Following on from a recent Journal of Screenwriting special issue on script development (2017, vol. 8:3), and in particular an article therein dedicated to reviewing the literature and ‘defining the field’, an expanded team of researchers follow up on those ideas and insights. In this article, then, we attempt to theorize script development as a ‘wicked problem’ that spans a range of themes and disciplines. As a ‘wicked’ team of authors, our expertise encompasses screenwriting theory, screenwriting practice, film and television studies, cultural policy, ethnography, gender studies and comedy. By drawing on these critical domains and creative practices, we present a series of interconnected themes that we hope not only suggests the potential for script development as a rich and exciting scholarly pursuit, but that also inspires and encourages other researchers to join forces in an attempt to solve the script development ‘puzzle’

Modification of the visual response properties of cerebellar neurons by norepinephrine

Extracellular recordings were conducted in the paraflocculus of anesthetized Long-Evans pigmented rats to determine how iontophoresis of norepinephrine (NE) affects the responsiveness of individual Purkinje cells and interneurons to presentations of visual stimuli within their visual receptive fields. Presentations of moving or stationary visual stimuli during the control (pre-NE) period elicited simple spike excitations or inhibitory responses in slightly more than one-half (55%, n = 32) of the cells tested (20 of 38 Purkinje cells, 12 of 20 interneurons). The predominant effect of NE iontophoresis was to improve visually evoked responses in those neurons which showed modulations in their simple spike discharge to control presentations of visual stimuli. A clear enhancement of visual responses by NE (i.e., absolute increase over control) was observed in 18 of the units, and in 12 of the 14 remaining cells, reductions in stimulus-bound discharge during catecholamine iontophoresis were accompanied by much larger depressions in background activity, resulting in a net enhancement in the ratio of signal-to-noise. NE differentially affected responses to stimulus movement in the preferred and non-preferred direction in one-third of these neurons, such that directional selectivity was increased. However, the orientation bias of individual units was unchanged by NE. Iontophoretic application of the [beta]-adrenergic antagonist sotalol but not the [alpha]-adrenergic antagonist phentolamine blocked these facilitating noradrenergic effects. An additional feature of noradrenergic action was revealed in tests conducted in 26 cells which did not respond to control presentations of visual stimuli. Iontophoresis of NE resulted in the elicitation of visual responses in 11 of these units, suggesting the possibility that NE might act in some cases to gate the efficacy of subliminal synaptic input conveyed by classical afferent channels. It is proposed that an important aspect of noradrenergic action within local cerebellar circuits might be to refine the receptive field properties of individual neuronal elements and thereby improve information flow through the cerebellum.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/28612/1/0000424.pd

Space Weather in the Saturn-Titan System

New evidence based on Cassini magnetic field and plasma data has revealed that the discovery of Titan outside Saturn’s magnetosphere during the T96 flyby on 2013 December 1 was the result of the impact of two consecutive interplanetary coronal mass ejections (ICMEs) that left the Sun in 2013 early November and interacted with the moon and the planet. We study the dynamic evolution of Saturn's magnetopause and bow shock, which evidences a magnetospheric compression from late November 28 to December 4 (at least), under prevailing solar wind dynamic pressures of 0.16-0.3 nPa. During this interval, transient disturbances associated with the two ICMEs are observed, allowing for the identification of their magnetic structures. By analyzing the magnetic field direction, and the pressure balance in Titan’s induced magnetosphere, we show that Cassini finds Saturn’s moon embedded in the second ICME after being swept by its interplanetary shock and amid a shower of solar energetic particles that may have caused dramatic changes in the moon’s lower ionosphere. Analyzing a list of Saturn's bow shock crossings during 2004-2016, we find that the magnetospheric compression needed for Titan to be in the supersonic solar wind can be generally associated with the presence of an ICME or a corotating interaction region. This leads to the conclusion that Titan would rarely face the pristine solar wind, but would rather interact with transient solar structures under extreme space weather conditions

Shocklets and Short Large Amplitude Magnetic Structures (SLAMS) in the high Mach foreshock of Venus

Shocklets and short large-amplitude magnetic structures (SLAMS) are steepened magnetic fluctuations commonly found in Earth's upstream foreshock. Here we present Venus Express observations from the 26th of February 2009 establishing their existence in the steady-state foreshock of Venus, building on a past study which found SLAMS during a substantial disturbance of the induced magnetosphere. The Venusian structures were comparable to those reported near Earth. The 2 Shocklets had magnetic compression ratios of 1.23 and 1.34 with linear polarization in the spacecraft frame. The 3 SLAMS had ratios between 3.22 and 4.03, two of which with elliptical polarization in the spacecraft frame. Statistical analysis suggests SLAMS coincide with unusually high solar wind Alfvén mach-number at Venus (12.5, this event). Thus, while we establish Shocklets and SLAMS can form in the stable Venusian foreshock, they may be rarer than at Earth. We estimate a lower limit of their occurrence rate of ≳14%

Memory consolidation in the cerebellar cortex

Several forms of learning, including classical conditioning of the eyeblink, depend upon the cerebellum. In examining mechanisms of eyeblink conditioning in rabbits, reversible inactivations of the control circuitry have begun to dissociate aspects of cerebellar cortical and nuclear function in memory consolidation. It was previously shown that post-training cerebellar cortical, but not nuclear, inactivations with the GABA(A) agonist muscimol prevented consolidation but these findings left open the question as to how final memory storage was partitioned across cortical and nuclear levels. Memory consolidation might be essentially cortical and directly disturbed by actions of the muscimol, or it might be nuclear, and sensitive to the raised excitability of the nuclear neurons following the loss of cortical inhibition. To resolve this question, we simultaneously inactivated cerebellar cortical lobule HVI and the anterior interpositus nucleus of rabbits during the post-training period, so protecting the nuclei from disinhibitory effects of cortical inactivation. Consolidation was impaired by these simultaneous inactivations. Because direct application of muscimol to the nuclei alone has no impact upon consolidation, we can conclude that post-training, consolidation processes and memory storage for eyeblink conditioning have critical cerebellar cortical components. The findings are consistent with a recent model that suggests the distribution of learning-related plasticity across cortical and nuclear levels is task-dependent. There can be transfer to nuclear or brainstem levels for control of high-frequency responses but learning with lower frequency response components, such as in eyeblink conditioning, remains mainly dependent upon cortical memory storage

βA3/A1-Crystallin controls anoikis-mediated cell death in astrocytes by modulating PI3K/AKT/mTOR and ERK survival pathways through the PKD/Bit1-signaling axis

During eye development, apoptosis is vital to the maturation of highly specialized structures such as the lens and retina. Several forms of apoptosis have been described, including anoikis, a form of apoptosis triggered by inadequate or inappropriate cell–matrix contacts. The anoikis regulators, Bit1 (Bcl-2 inhibitor of transcription-1) and protein kinase-D (PKD), are expressed in developing lens when the organelles are present in lens fibers, but are downregulated as active denucleation is initiated. We have previously shown that in rats with a spontaneous mutation in the Cryba1 gene, coding for βA3/A1-crystallin, normal denucleation of lens fibers is inhibited. In rats with this mutation (Nuc1), both Bit1 and PKD remain abnormally high in lens fiber cells. To determine whether βA3/A1-crystallin has a role in anoikis, we induced anoikis in vitro and conducted mechanistic studies on astrocytes, cells known to express βA3/A1-crystallin. The expression pattern of Bit1 in retina correlates temporally with the development of astrocytes. Our data also indicate that loss of βA3/A1-crystallin in astrocytes results in a failure of Bit1 to be trafficked to the Golgi, thereby suppressing anoikis. This loss of βA3/A1-crystallin also induces insulin-like growth factor-II, which increases cell survival and growth by modulating the phosphatidylinositol-3-kinase (PI3K)/AKT/mTOR and extracellular signal-regulated kinase pathways. We propose that βA3/A1-crystallin is a novel regulator of both life and death decisions in ocular astrocytes

High Diversity of the Saliva Microbiome in Batwa Pygmies

We describe the saliva microbiome diversity in Batwa Pygmies, a former hunter-gatherer group from Uganda, using next-generation sequencing of partial 16S rRNA sequences. Microbial community diversity in the Batwa is significantly higher than in agricultural groups from Sierra Leone and the Democratic Republic of Congo. We found 40 microbial genera in the Batwa, which have previously not been described in the human oral cavity. The distinctive composition of the salvia microbiome of the Batwa may have been influenced by their recent different lifestyle and diet

The Effects of Breeding Protocol in C57BL/6J Mice on Adult Offspring Behaviour

Animal experiments have demonstrated that a wide range of prenatal exposures can impact on the behaviour of the offspring. However, there is a lack of evidence as to whether the duration of sire exposure could affect such outcomes. We compared two widely used methods for breeding offspring for behavioural studies. The first involved housing male and female C57Bl/6J mice together for a period of time (usually 10–12 days) and checking for pregnancy by the presence of a distended abdomen (Pair-housed; PH). The second involved daily introduction of female breeders to the male homecage followed by daily checks for pregnancy by the presence of vaginal plugs (Time-mated; TM). Male and female offspring were tested at 10 weeks of age on a behavioural test battery including the elevated plus-maze, hole board, light/dark emergence, forced swim test, novelty-suppressed feeding, active avoidance and extinction, tests for nociception and for prepulse inhibition (PPI) of the acoustic startle response. We found that length of sire exposure (LSE) had no significant effects on offspring behaviour, suggesting that the two breeding protocols do not differentially affect the behavioural outcomes of interest. The absence of LSE effects on the selected variables examined does not detract from the relevance of this study. Information regarding the potential influences of breeding protocol is not only absent from the literature, but also likely to be of particular interest to researchers studying the influence of prenatal manipulations on adult behaviour