Journal Club: Structural Biology, Physiology, and Virology.

Wednesday, January 8, 2014

STRUCTURAL BIOLOGY: Structure of the TRPV1 ion channel determined by electron cryo-microscopy. Liao, M., Cao, E., Julius, D., Cheng, Y. Nature. 504(7478):107-112. & TRPV1 structures in distinct conformations reveal activation mechanisms. Cao, E., Liao, M., Cheng, Y., Julius, D. Nature 504(7478):113-118.

Your ability to enjoy the zing of a jalapeño depends on the TRPV1 channel, which is activated by capsaicin, the substance responsible for peppers tasting hot, and high temperatures. Not knowing in detail the structure of this receptor has made it difficult to determinine how it responds to these stimuli.

Protein structures are usually determined by X-ray crystallography, but crystallizing some proteins is a titanic challenge. An alternative technology, called single-particle electron cryomicroscopy (cryo-EM), uses electron microscopy of a solution of the protein that was frozen extremely rapidly. By combining images of many individual protein molecules, each seen from a different angle, a three-dimensional structure can be reconstructed.

In these papers, the researchers used cryo-EM to determine the structure of TRPV1 at a resolution of 3.4 angstroms, comparable to that of X-ray crystallography. They developed a new direct electron detector and new motion-correcting algorithms and created the most high-resolution structure of a membrane protein yet achieved using cryo-EM.

The researchers determined the structure of the unbound receptor and how the receptor changed when activated. In comparison to the related voltage-gated ion channels, TRPV1 shared the same basic structure but displayed a different mechanism of opening when activated.  

PHYSIOLOGY: Targeting of αv integrin identifies a core molecular pathway that regulates fibrosis in several organs. Henderson, N.C., et al. (Sheppard). Nat Med. 19(12):1617-1624.

The formation of excessive fibrous connective tissue, or fibrosis, is characteristic of many diseases, including cirrhosis, pulmonary fibrosis and chronic kidney disease. Currently, treatment options for fibrosis are very limited, and the diseases often progress until the organs fail.  

Myofibroblasts are a known major contributor to fibrosis. Previous research has suggested that the cell adhesion molecule αv integrin promotes this process by activating a fibrosis-promoting factor, TGF-β. However, a lack of methods for specifically genetically targeting these cells has hindered investigation.

In this paper, researchers demonstrated that Pdgrfrb promoter-driven Cre can be used to selectively ablate αv integrin in myofibroblasts in multiple organs. Such loss of αv integrin reduced fibrosis in models in which it was experimentally induced in the liver, kidney and lung. They further showed that an αv integrin inhibitor could decrease experimentally induced liver and lung fibrosis, suggesting that it may be possible to develop drugs that target αv integrin to treat fibrosis.

VIROLOGY: Mutational and fitness landscapes of an RNA virus revealed through population sequencing. Acevedo, A., Brodsky, L., Andino, R. Nature. doi:10.1038/nature12861.

RNA viruses mutate frequently, resulting in a genetically diverse population even within one infected individual. This increases the likelihood that the virus can adapt to new challenges and can be essential to causing disease.

Using next-generation sequencing would seem to be the natural way to assess such viral genetic diversity, but the very low frequency of each mutation makes it difficult to distinguish a mutation from errors in the sequencing process itself. To get around this problem, the researchers generated tandem repeats of genomic RNA fragments before sequencing. The chance of having a sequencing error at a given base is already low, and the chance of the same error occurring in all three tandem repeats is many orders of magnitude lower.  

In this paper, the researchers comprehensively documented changes in serially passaged poliovirus. They determined the rate at which each type of point mutation occurs — such as adenine to guanine or guanine to uracil. They also calculated the effect of thousands of different mutations on viral fitness, which may lead to a better understanding of the function of the virus’s proteins.