Targeted therapy for LIMD1-deficient non-small cell lung cancer subtypes

An early event in lung oncogenesis is loss of the tumour suppressor gene LIMD1 (LIM domains containing 1); this encodes a scaffold protein, which suppresses tumorigenesis via a number of different mechanisms. Approximately 45% of non-small cell lung cancers (NSCLC) are deficient in LIMD1, yet this subtype of NSCLC has been overlooked in preclinical and clinical investigations. Defining therapeutic targets in these LIMD1 loss-of-function patients is difficult due to a lack of ‘druggable’ targets, thus alternative approaches are required. To this end, we performed the first drug repurposing screen to identify compounds that confer synthetic lethality with LIMD1 loss in NSCLC cells. PF-477736 was shown to selectively target LIMD1-deficient cells in vitro through inhibition of multiple kinases, inducing cell death via apoptosis. Furthermore, PF-477736 was effective in treating LIMD1^−/− tumours in subcutaneous xenograft models, with no significant effect in LIMD1^+/+ cells. We have identified a novel drug tool with significant preclinical characterisation that serves as an excellent candidate to explore and define LIMD1-deficient cancers as a new therapeutic subgroup of critical unmet need.Subject terms: Targeted therapies, Non-small-cell lung cancer     

A Computational Model of Lipopolysaccharide-Induced Nuclear Factor Kappa B Activation: A Key Signalling Pathway in Infection-Induced Preterm Labour

Preterm birth is the single biggest cause of significant neonatal morbidity and mortality, and the incidence is rising. Development of new therapies to treat and prevent preterm labour is seriously hampered by incomplete understanding of the molecular mechanisms that initiate labour at term and preterm. Computational modelling provides a new opportunity to improve this understanding. It is a useful tool in (i) identifying gaps in knowledge and informing future research, and (ii) providing the basis for an in silico model of parturition in which novel drugs to prevent or treat preterm labour can be “tested”. Despite their merits, computational models are rarely used to study the molecular events initiating labour. Here, we present the first attempt to generate a dynamic kinetic model that has relevance to the molecular mechanisms of preterm labour. Using published data, we model an important candidate signalling pathway in infection-induced preterm labour: that of lipopolysaccharide (LPS) -induced activation of Nuclear Factor kappa B. This is the first model of this pathway to explicitly include molecular interactions upstream of Nuclear Factor kappa B activation. We produced a formalised graphical depiction of the pathway and built a kinetic model based on ordinary differential equations. The kinetic model accurately reproduced published in vitro time course plots of Lipopolysaccharide-induced Nuclear Factor kappa B activation in mouse embryo fibroblasts. In this preliminary work we have provided proof of concept that it is possible to build computational models of signalling pathways that are relevant to the regulation of labour, and suggest that models that are validated with wet-lab experiments have the potential to greatly benefit the field.     

Flyway‐scale analysis reveals that the timing of migration in wading birds is becoming later

1. Understanding the implications of climate change for migratory animals is paramount for establishing how best to conserve them. A large body of evidence suggests that birds are migrating earlier in response to rising temperatures, but many studies focus on single populations of model species.
2. Migratory patterns at large spatial scales may differ from those occurring in single populations, for example because of individuals dispersing outside of study areas. Furthermore, understanding phenological trends across species is vital because we need a holistic understanding of how climate change affects wildlife, especially as rates of temperature change vary globally.
3. The life cycles of migratory wading birds cover vast latitudinal gradients, making them particularly susceptible to climate change and, therefore, ideal model organisms for understanding its effects. Here, we implement a novel application of changepoint detection analysis to investigate changes in the timing of migration in waders at a flyway scale using a thirteen‐year citizen science dataset (eBird) and determine the influence of changes in weather conditions on large‐scale migratory patterns.
4. In contrast to most previous research, our results suggest that migration is getting later in both spring and autumn. We show that rates of change were faster in spring than autumn in both the Afro‐Palearctic and Nearctic flyways, but that weather conditions in autumn, not in spring, predicted temporal changes in the corresponding season. Birds migrated earlier in autumn when temperatures increased rapidly, and later with increasing headwinds.
5. One possible explanation for our results is that migration is becoming later due to northward range shifts, which means that a higher proportion of birds travel greater distances and therefore take longer to reach their destinations. Our findings underline the importance of considering spatial scale when investigating changes in the phenology of migratory bird species.

     

Drosophila katanin-60 depolymerizes and severs at microtubule defects

Microtubule (MT) length and location is tightly controlled in cells. One novel family of MT-associated proteins that regulates MT dynamics is the MT-severing enzymes. In this work, we investigate how katanin (p60), believed to be the first discovered severing enzyme, binds and severs MTs via single molecule total internal reflection fluorescence microscopy. We find that severing activity depends on katanin concentration. We also find that katanin can remove tubulin dimers from the ends of MTs, appearing to depolymerize MTs. Strikingly, katanin localizes and severs at the interface of GMPCPP-tubulin and GDP-tubulin suggesting that it targets to protofilament-shift defects. Finally, we observe that binding duration, mobility, and oligomerization are ATP dependent.     

A minimal spliceosomal complex A recognizes the branch site and polypyrimidine tract.

The association of U2 snRNP with the pre-mRNA branch region is a critical step in the assembly of spliceosomal complexes. We describe an assembly process that reveals both minimal requirements for formation of a U2 snRNP-substrate RNA complex, here designated the Amin complex, and specific interactions with the branch site adenosine. The substrate is a minimal RNA oligonucleotide, containing only a branch sequence and polypyrimidine tract. Interactions at the branch site adenosine and requirements for polypyrimidine tract-binding proteins for the Amin complex are the same as those of authentic prespliceosome complex A. Surprisingly, Amin complex formation does not require U1 snRNP or ATP, suggesting that these factors are not necessary for stable binding of U2 snRNP per se, but rather are necessary for accessibility of components on longer RNA substrates. Furthermore, there is an ATP-dependent activity that releases or destabilizes U2 snRNP from branch sequences. The simplicity of the Amin complex will facilitate a detailed understanding of the assembly of prespliceosomes.     

MinCD-dependent regulation of the polarity of SpoIIIE assembly and DNA transfer

During Bacillus subtilis sporulation, the SpoIIIE DNA translocase moves a trapped chromosome across the sporulation septum into the forespore. The direction of DNA translocation is controlled by the specific assembly of SpoIIIE in the mother cell and subsequent export of DNA into the forespore. We present evidence that the MinCD heterodimer, which spatially regulates cell division during vegetative growth, serves as a forespore-specific inhibitor of SpoIIIE assembly. The deletion of minCD increases the ability of forespore-expressed SpoIIIE to assemble and translocate DNA, and causes otherwise wild-type cells to reverse the direction of DNA transfer, producing anucleate forespores. We propose that two distinct mechanisms ensure the specific assembly of SpoIIIE in the mother cell, the partitioning of more SpoIIIE molecules into the larger mother cell by asymmetric cell division and the MinCD-dependent repression of SpoIIIE assembly in the forespore. Our results suggest that the ability of MinCD to sense positional information is utilized during sporulation to regulate protein assembly differentially on the two faces of the sporulation septum.     

GABAergic modulation of primary gustatory afferent synaptic efficacy

Modulation of synaptic transmission at the primary sensory afferent synapse is well documented for the somatosensory and olfactory systems. The present study was undertaken to test whether GABA impacts on transmission of gustatory information at the primary afferent synapse. In goldfish, the vagal gustatory input terminates in a laminated structure, the vagal lobes, whose sensory layers are homologous to the mammalian nucleus of the solitary tract. We relied on immunoreactivity for the GABA-transporter, GAT-1, to determine the distribution of GABAergic synapses in the vagal lobe. Immunocytochemistry showed dense, punctate GAT-1 immunoreactivity coincident with the layers of termination of primary afferent fibers. The laminar nature and polarized dendritic structure of the vagal lobe make it amenable to an in vitro slice preparation to study early synaptic events in the transmission of gustatory input. Electrical stimulation of the gustatory nerves in vitro produces synaptic field potentials (fEPSPs) predominantly mediated by ionotropic glutamate receptors. Bath application of either the GABA(A) receptor agonist muscimol or the GABA(B) receptor agonist baclofen caused a nearly complete suppression of the primary fEPSP. Coapplication of the appropriate GABA(A) or GABA(B) receptor antagonist bicuculline or CGP-55845 significantly reversed the effects of the agonists. These data indicate that GABAergic terminals situated in proximity to primary gustatory afferent terminals can modulate primary afferent input via both GABA(A) and GABA(B) receptors. The mechanism of action of GABA(B) receptors suggests a presynaptic locus of action for that receptor.     

RFLP-based genetic maps of wheat homoeologous group 7 chromosomes

Summary Restriction fragment length polymorphism (RFLP) mapping was attempted using 18 cDNA clones, 14 anonymous and 4 of known function, which had been shown to have homologous DNA sequences on the group 7 chromosomes of wheat. The loci identified by these probes have been mapped on one or more chromosomes in this homoeologous group using linkage data derived from various F₂, random inbred, doubled haploid and single chromosome recombinant populations. The maps also include three isozyme loci, five disease resistance loci, two anthocyanin pigment loci and a vernalisation response locus. The mapping data have been used to determine the extent of map co-linearity over the A, B and D genomes, the degree of RFLP variability in the three genomes and the relative efficiency of various restriction enzymes in detecting RFLPs in wheat. The strategy for future mapping in wheat, particularly the use of alien genomes or segments, such as that from Aegilops ventricosa used here, is discussed.