A native chemical chaperone in the human eye lens.

Cataract is one of the most prevalent protein aggregation disorders and still the most common cause of vision loss worldwide. The metabolically quiescent core region of the human lens lacks cellular or protein turnover; it has therefore evolved remarkable mechanisms to resist light-scattering protein aggregation for a lifetime. We now report that one such mechanism involves an unusually abundant lens metabolite, myo-inositol, suppressing aggregation of lens crystallins. We quantified aggregation suppression using our previously well-characterized in vitro aggregation assays of oxidation-mimicking human γD-crystallin variants and investigated myo-inositol's molecular mechanism of action using solution NMR, negative-stain TEM, differential scanning fluorometry, thermal scanning Raman spectroscopy, turbidimetry in redox buffers, and free thiol quantitation. Unlike many known chemical chaperones, myo-inositol's primary target was not the native, unfolded, or final aggregated states of the protein; rather, we propose that it was the rate-limiting bimolecular step on the aggregation pathway. Given recent metabolomic evidence that it is severely depleted in human cataractous lenses compared to age-matched controls, we suggest that maintaining or restoring healthy levels of myo-inositol in the lens may be a simple, safe, and globally accessible strategy to prevent or delay lens opacification due to age-onset cataract.

Profoundly different prion diseases in knock-in mice carrying single PrP codon substitutions associated with human diseases.

In man, mutations in different regions of the prion protein (PrP) are associated with infectious neurodegenerative diseases that have remarkably different clinical signs and neuropathological lesions. To explore the roots of this phenomenon, we created a knock-in mouse model carrying the mutation associated with one of these diseases [Creutzfeldt-Jakob disease (CJD)] that was exactly analogous to a previous knock-in model of a different prion disease [fatal familial insomnia (FFI)]. Together with the WT parent, this created an allelic series of three lines, each expressing the same protein with a single amino acid difference, and with all native regulatory elements intact. The previously described FFI mice develop neuronal loss and intense reactive gliosis in the thalamus, as seen in humans with FFI. In contrast, CJD mice had the hallmark features of CJD, spongiosis and proteinase K-resistant PrP aggregates, initially developing in the hippocampus and cerebellum but absent from the thalamus. A molecular transmission barrier protected the mice from any infectious prion agents that might have been present in our mouse facility and allowed us to conclude that the diseases occurred spontaneously. Importantly, both models created agents that caused a transmissible neurodegenerative disease in WT mice. We conclude that single codon differences in a single gene in an otherwise normal genome can cause remarkably different neurodegenerative diseases and are sufficient to create distinct protein-based infectious elements.