Characterisation and Validation of Insertions and Deletions in 173 Patient Exomes

Recent advances in genomics technologies have spurred unprecedented efforts in genome and exome re-sequencing aiming to unravel the genetic component of rare and complex disorders. While in rare disorders this allowed the identification of novel causal genes, the missing heritability paradox in complex diseases remains so far elusive. Despite rapid advances of next-generation sequencing, both the technology and the analysis of the data it produces are in its infancy. At present there is abundant knowledge pertaining to the role of rare single nucleotide variants (SNVs) in rare disorders and of common SNVs in common disorders. Although the 1,000 genome project has clearly highlighted the prevalence of rare variants and more complex variants (e.g. insertions, deletions), their role in disease is as yet far from elucidated.We set out to analyse the properties of sequence variants identified in a comprehensive collection of exome re-sequencing studies performed on samples from patients affected by a broad range of complex and rare diseases (N = 173). Given the known potential for Loss of Function (LoF) variants to be false positive, we performed an extensive validation of the common, rare and private LoF variants identified, which indicated that most of the private and rare variants identified were indeed true, while common novel variants had a significantly higher false positive rate. Our results indicated a strong enrichment of very low-frequency insertion/deletion variants, so far under-investigated, which might be difficult to capture with low coverage and imputation approaches and for which most of study designs would be under-powered. These insertions and deletions might play a significant role in disease genetics, contributing specifically to the underlining rare and private variation predicted to be discovered through next generation sequencing.     

Modeling Huddling Penguins

We present a systematic and quantitative model of huddling penguins. In this mathematical model, each individual penguin in the huddle seeks only to reduce its own heat loss. Consequently, penguins on the boundary of the huddle that are most exposed to the wind move downwind to more sheltered locations along the boundary. In contrast, penguins in the interior of the huddle neither have the space to move nor experience a significant heat loss, and they therefore remain stationary. Through these individual movements, the entire huddle experiences a robust cumulative effect that we identify, describe, and quantify. This mathematical model requires a calculation of the wind flowing around the huddle and of the resulting temperature distribution. Both of these must be recomputed each time an individual penguin moves since the huddle shape changes. Using our simulation results, we find that the key parameters affecting the huddle dynamics are the number of penguins in the huddle, the wind strength, and the amount of uncertainty in the movement of the penguins. Moreover, we find that the lone assumption of individual penguins minimizing their own heat loss results in all penguins having approximately equal access to the warmth of the huddle.     

Karrikins force a rethink of strigolactone mode of action

Strigolactones (SL) and karrikins (KAR) both contain essential butenolide moieties, and both require the F-box protein MAX2 to control seed germination and photomorphogenesis in Arabidopsis thaliana. A new discovery that SL and KAR also require related α/β-hydrolase proteins for such activity suggests that they operate through a similar molecular mechanism. Based on structural similarity, a previously proposed mode of action for SL was also considered for KAR, but recent structure-activity studies suggest that this mechanism may not apply. Here we rationalise these observations into a hypothesis whereby different α/β-hydrolases distinguish SL and KAR by virtue of their non-butenolide moieties and catalyze nucleophilic attack on the butenolide. The products would be different for SL and KAR, and in the case of SL they have no biological activity. The inference is that nucleophilic attack on SL and KAR by α/β-hydrolases is required for their bioactivity, but the hydrolysis products are not.     

Thermodynamic contributions of the reactions of DNA intramolecular structures with their complementary strands

One focus of our research is to further our understanding of the physico-chemical properties of unusual DNA structures and their interaction with complementary oligonucleotides. We have investigated three types of reactions involving the interaction of intramolecular DNA complexes with their complementary single strands of varied length. Specifically, we have used a combination of isothermal titration (ITC) and differential scanning (DSC) calorimetry and spectroscopy techniques to determine standard thermodynamic profiles for the reaction of an i-motif, G-quadruplex, and triplex with their complementary strands. The enthalpies for each reaction are measured directly in ITC titrations and compared with those obtained indirectly from Hess cycles using DSC unfolding data. All reactions investigated yielded favorable free energy contributions, indicating that each single strand is able to invade and disrupt the corresponding intramolecular DNA complex. These favorable free energy terms are enthalpy driven, which result from a compensation of exothermic contributions, due to the formation of additional base-pair stacks (or base-triplet stacks) in the duplex product (or triplex product), immobilization of electrostricted water by the base-pair and base-triplet stacks, and the removal of structural water from the reactant single strands; and endothermic contributions from the disruption of base-base stacking interactions of the reactant single strands. This investigation of nucleic acid reactions has provided new methodology, based on physico-chemical principles, to determine the molecular forces involved in the interactions between DNA nucleic acid structures. This methodology may be used in targeting reactions for the control of gene expression.     

Can general practitioners influence exercise habits? Controlled trial.

A health promotion campaign in the community organised by the general practitioner was evaluated by the use of a questionnaire before and after the campaign, both in the village where the campaign was based and in a control village. There was a general increase in the amount of exercise that people in both villages took. The results showed that having received a questionnaire, and been subject to the campaign, more people took exercise regularly.     

Increased amounts of hybrid (heavy/heavy) DNA in Bloom's syndrome fibroblasts

The nuclear DNA of fibroblasts from patients suffering with Bloom's syndrome, density labeled for less than one round of DNA replication to give heavy/light molecules, was examined for spontaneous amounts of heavy/heavy DNA (hybrid DNA). When compared to normal fibroblasts the Bloom's syndrome cells exhibited a sixfold increase in such DNA.