Mapping genetic variations to three- dimensional protein structures to enhance variant interpretation: a proposed framework

The translation of personal genomics to precision medicine depends on the accurate interpretation of the multitude of genetic variants observed for each individual. However, even when genetic variants are predicted to modify a protein, their functional implications may be unclear. Many diseases are caused by genetic variants affecting important protein features, such as enzyme active sites or interaction interfaces. The scientific community has catalogued millions of genetic variants in genomic databases and thousands of protein structures in the Protein Data Bank. Mapping mutations onto three-dimensional (3D) structures enables atomic-level analyses of protein positions that may be important for the stability or formation of interactions; these may explain the effect of mutations and in some cases even open a path for targeted drug development. To accelerate progress in the integration of these data types, we held a two-day Gene Variation to 3D (GVto3D) workshop to report on the latest advances and to discuss unmet needs. The overarching goal of the workshop was to address the question: what can be done together as a community to advance the integration of genetic variants and 3D protein structures that could not be done by a single investigator or laboratory? Here we describe the workshop outcomes, review the state of the field, and propose the development of a framework with which to promote progress in this arena. The framework will include a set of standard formats, common ontologies, a common application programming interface to enable interoperation of the resources, and a Tool Registry to make it easy to find and apply the tools to specific analysis problems. Interoperability will enable integration of diverse data sources and tools and collaborative development of variant effect prediction methods

Calcium–axonemal microtubuli interactions underlie mechanism(s) of primary cilia morphological changes

We have used cell culture of astrocytes aligned within microchannels to investigate calcium effects on primary cilia morphology. In the absence of calcium and in the presence of flow of media (10 µL.s-1) the majority (90%) of primary cilia showed reversible bending with an average curvature of 2.1 ± 0.9 × 10-4 nm-1. When 1.0 mM calcium was present, 90% of cilia underwent bending. Forty percent of these cilia demonstrated strong irreversible bending, resulting in a final average curvature of 3.9 ± 1 × 10-4 nm-1, while 50% of cilia underwent bending similar to that observed during calcium-free flow. The average length of cilia was shifted toward shorter values (3.67 ± 0.34 µm) when exposed to excess calcium (1.0 mM), compared to media devoid of calcium (3.96 ± 0.26 µm). The number of primary cilia that became curved after calcium application was reduced when the cell culture was pre-incubated with 15 µM of the microtubule stabilizer, taxol, for 60 min prior to calcium application. Calcium caused single microtubules to curve at a concentration ˜1.0 mM in vitro, but at higher concentration (˜1.5 mM) multiple microtubule curving occurred. Additionally, calcium causes microtubule-associated protein-2 conformational changes and its dislocation from the microtubule wall at the location of microtubule curvature. A very small amount of calcium, that is 1.45 × 1011 times lower than the maximal capacity of TRPPs calcium channels, may cause gross morphological changes (curving) of primary cilia, while global cytosol calcium levels are expected to remain unchanged. These findings reflect the non-linear manner in which primary cilia may respond to calcium signaling, which in turn may influence the course of development of ciliopathies and cancer