Demystifying the RAD fad

We are writing in response to the population and phylogenomics meeting review by Andrews & Luikart ( 2014 ) entitled ‘Recent novel approaches for population genomics data analysis’. Restriction‐site‐associated DNA (RAD) sequencing has become a powerful and useful approach in molecular ecology, with several different published methods now available to molecular ecologists, none of which can be considered the best option in all situations. A&L report that the original RAD protocol of Miller et al. ( 2007 ) and Baird et al. ( 2008 ) is superior to all other RAD variants because putative PCR duplicates can be identified (see Baxter et al. 2011 ), thereby reducing the impact of PCR artefacts on allele frequency estimates (Andrews & Luikart 2014 ). In response, we (i) challenge the assertion that the original RAD protocol minimizes the impact of PCR artefacts relative to that of other RAD protocols, (ii) present additional biases in RADseq that are at least as important as PCR artefacts in selecting a RAD protocol and (iii) highlight the strengths and weaknesses of four different approaches to RADseq which are a representative sample of all RAD variants: the original RAD protocol (mbRAD, Miller et al. 2007 ; Baird et al. 2008 ), double digest RAD (ddRAD, Peterson et al. 2012 ), ezRAD (Toonen et al. 2013 ) and 2bRAD (Wang et al. 2012 ). With an understanding of the strengths and weaknesses of different RAD protocols, researchers can make a more informed decision when selecting a RAD protocol.     

Increased fear learning,spatial learning as well as neophobia in Rgs2−/− mice

Anxiety disorders result from a complex interplay of genetic and environmental factors such as stress. On the level of cellular signaling, regulator of G protein signaling 2 (Rgs2) has been implicated in human and rodent anxiety. However, there is limited knowledge about the role of Rgs2 in fear learning and reactivity to stress. In this study, Rgs2^?/? mice showed increased fear learning, male mice displayed increased contextual and cued fear learning, while females showed selectively enhanced cued fear learning. Male Rgs2^?/? mice displayed increased long‐term‐contextual fear memory, but increased cued fear extinction. Learning in spatial non‐aversive paradigms was also increased in Rgs2^?/? mice. Female, but not male mice show increased spatial learning in the Barnes maze, while male mice showed enhanced place preference in the IntelliCage, rendering enhanced cognitive function non‐specific for aversive stimuli. Consistent with the previous results, Rgs2 deletion resulted in increased innate anxiety, including neophobic behavior expressed as hypolocomotion, in three different tests based on the approach‐avoidance conflict. Acute electric foot shock stress provoked hypolocomotion in several exploration‐based tests, suggesting fear generalization in both genotypes. Rgs2 deletion was associated with reduced monoaminergic neurotransmitter levels in the hippocampus and prefrontal cortex and disturbed corresponding GPCR expression of the adrenergic, serotonergic, dopaminergic and neuropeptide Y system. Taken together, Rgs2 deletion promotes improved cognitive function as well as increased anxiety‐like behavior, but has no effect on acute stress reactivity. These effects may be related to the observed disruption of the monoaminergic systems.     

Ail protein binds ninth type III fibronectin repeat (9FNIII) within central 120-kDa region of fibronectin to facilitate cell binding by Yersinia pestis

The Yersinia pestis adhesin molecule Ail interacts with the extracellular matrix protein fibronectin (Fn) on host cells to facilitate efficient delivery of cytotoxic Yop proteins, a process essential for plague virulence. A number of bacterial pathogens are known to bind to the N-terminal region of Fn, comprising type I Fn (FNI) repeats. Using proteolytically generated Fn fragments and purified recombinant Fn fragments, we demonstrated that Ail binds the centrally located 120-kDa fragment containing type III Fn (FNIII) repeats. A panel of monoclonal antibodies (mAbs) that recognize specific epitopes within the 120-kDa fragment demonstrated that mAb binding to (9)FNIII blocks Ail-mediated bacterial binding to Fn. Epitopes of three mAbs that blocked Ail binding to Fn were mapped to a similar face of (9)FNIII. Antibodies directed against (9)FNIII also inhibited Ail-dependent cell binding activity, thus demonstrating the biological relevance of this Ail binding region on Fn. Bacteria expressing Ail on their surface could also bind a minimal fragment of Fn containing repeats (9-10)FNIII, and this binding was blocked by a mAb specific for (9)FNIII. These data demonstrate that Ail binds to (9)FNIII of Fn and presents Fn to host cells to facilitate cell binding and delivery of Yops (cytotoxins of Y. pestis), a novel interaction, distinct from other bacterial Fn-binding proteins.     

The molecular basis of erectile dysfunction: from bench to bedside

Erectile dysfunction is a common problem affecting many men across all age groups. Its etiology is multifactorial. Hormonal, vascular, neurogenic, lifestyle, and psychological entities have all been implicated as causative agents. The molecular basis underlying its etiology and progression is complex and still challenges researchers in the field. Nonetheless, newly discovered common pathways and targets of its pathogenesis have opened a new era for both prevention and active treatment of the disease. This review describes some of the known molecular mechanisms contributing to erectile dysfunction and discusses the future of gene therapy for the disease.     

Anti-bat tiger moth sounds: Form and function

The night sky is the venue of an ancient acoustic battle between echolocating bats and their insect prey. Many tiger moths (Lepidoptera: Arctiidae) answer the attack calls of bats with a barrage of high frequency clicks. Some moth species use these clicks for acoustic aposematism and mimicry, and others for sonar jamming, however, most of the work on these defensive functions has been done on individual moth species. We here analyze the diversity of structure in tiger moth sounds from 26 spe-cies collected at three locations in North and South America. A principal components analysis of the anti-bat tiger moth sounds reveals that they vary markedly along three axes: (1) frequency, (2) duty cycle (sound production per unit time) and frequency modulation, and (3) modulation cycle (clicks produced during flexion and relaxation of the sound producing tymbal) structure. Tiger moth species appear to cluster into two distinct groups: one with low duty cycle and few clicks per modulation cycle that supports an acoustic aposematism function, and a second with high duty cycle and many clicks per modulation cycle that is con-sistent with a sonar jamming function. This is the first evidence from a community-level analysis to support multiple functions for tiger moth sounds. We also provide evidence supporting an evolutionary history for the development of these strategies. Further-more, cross-correlation and spectrogram correlation measurements failed to support a "phantom echo" mechanism underlying sonar jamming, and instead point towards echo interference.     

Changes in the ratio of free NEDD8 to ubiquitin triggers NEDDylation by ubiquitin enzymes

Ubiquitin and UBL (ubiquitin-like) modifiers are small proteins that covalently modify other proteins to alter their properties or behaviours. Ubiquitin modification (ubiquitylation) targets many substrates, often leading to their proteasomal degradation. NEDD8 (neural-precursor-cell-expressed developmentally down-regulated 8) is the UBL most closely related to ubiquitin, and its best-studied role is the activation of CRLs (cullin-RING ubiquitin ligases) by its conjugation to a conserved C-terminal lysine residue on cullin proteins. The attachment of UBLs requires three UBL-specific enzymes, termed E1, E2 and E3, which are usually well insulated from parallel UBL pathways. In the present study, we report a new mode of NEDD8 conjugation (NEDDylation) whereby the UBL NEDD8 is linked to proteins by ubiquitin enzymes in vivo. We found that this atypical NEDDylation is independent of classical NEDD8 enzymes, conserved from yeast to mammals, and triggered by an increase in the NEDD8 to ubiquitin ratio. In cells, NEDD8 overexpression leads to this type of NEDDylation by increasing the concentration of NEDD8, whereas proteasome inhibition has the same effect by depleting free ubiquitin. We show that bortezomib, a proteasome inhibitor used in cancer therapy, triggers atypical NEDDylation in tissue culture, which suggests that a similar process may occur in patients receiving this treatment.     

Prefrontal deep projection neurons enable cognitive flexibility via persistent feedback monitoring

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A rapid and sensitive method for the determination of serotonin turnover in discrete brain areas

Summary The present method permits rapid and sensitive study of the concentration levels of serotonin (5-HT) and 5-hydroxyindolacetic acid (5-HIAA) in the same tissue sample and the determination of the turnover of these substances in small brain parts.I wish to thank Merck Sharp and Dohme for supplies of probenecid.