Molecular genetics of congenital cataracts.

Congenital Cataract-Causing Mutation G129C in γC-Crystallin Promotes the Accumulation of Two Distinct Unfolding Intermediates That Form Highly Toxic Aggregates

β/γ-Crystallins are predominant structural proteins in the cytoplasm of lens fiber cells and share a similar fold composing of four Greek-key motifs divided into two domains. Numerous cataract-causing mutations have been identified in various β/γ-crystallins, but the mechanisms underlying cataract caused by most mutations remains uncharacterized. The S228P mutation in βB1-crystallin has been linked to autosomal dominant congenital nuclear cataract. Here we found that the S228P mutant was prone to aggregate and degrade in both of the human and E. coli cells. The intracellular S228P aggregates could be redissolved by lanosterol. The S228P mutation modified the refolding pathway of βB1-crystallin by affecting the formation of the dimeric intermediate but not the monomeric intermediate. Compared with native βB1-crystallin, the refolded S228P protein had less packed structures, unquenched Trp fluorophores and increased hydrophobic exposure. The refolded S228P protein was prone to aggregate at the physiological temperature and decreased the protective effect of βB1-crystallin on βA3-crystallin. Molecular dynamic simulation studies indicated that the mutation decreased the subunit binding energy and modified the distribution of surface electrostatic potentials. More importantly, the mutation separated two interacting loops in the C-terminal domain, which shielded the hydrophobic core from solvent in native βB1-crystallin. These two interacting loops are highly conserved in both of the N- and C-terminal domains of all β/γ-crystallins. We propose that these two interacting loops play an important role in the folding and structural stability of β/γ-crystallin domains by protecting the hydrophobic core from solvent access.

/pdf/cataract-causing-mutation-s228p-promotes-bb1-crystallin-1xk9eik339.pdf

Cataract-causing mutation S228P promotes βB1-crystallin aggregation and degradation by separating two interacting loops in C-terminal domain

Increasing susceptibility to oxidative stress by cataract-causing crystallin mutations.

The high solubility and lifelong stability of crystallins are crucial to the maintenance of lens transparency and optical properties Numerous crystallin mutations have been linked to congenital cataract, which is one of the leading causes of newborn blindness Besides cataract, several crystallin mutations have also been linked to syndromes such as congenital microcornea-cataract syndrome (CMCC) However, the molecular mechanism of CMCC caused by crystallin mutations remains elusive In the present study, we investigated the mechanism of CMCC caused by the X253R mutation in βB1-crystallin The exogenously expressed X253R proteins were prone to form p62-negative aggregates in HeLa cells, strongly inhibited cell proliferation and induced cell apoptosis The intracellular X253R aggregates could be successfully redissolved by lanosterol but not cholesterol The extra 26 residues at the C-terminus of βB1-crystallin introduced by the X253R mutation had little impact on βB1-crystallin structure and stability, but increased βB1-crystallin hydrophobicity and decreased its solubility Interestingly, the X253R mutant fully abolished the aggregatory propensity of βB1- and βA3/βB1-crystallins at high temperatures, suggesting that X253R was an aggregation-inhibition mutation of β-crystallin homomers and heteromers in dilute solutions Our results suggest that an increase in hydrophobicity and a decrease in solubility might be responsible for cataractogenesis induced by the X253R mutation, while the cytotoxic effect of X253R aggregates might contribute to the defects in ocular development Our results also highlight that, at least in some cases, the aggregatory propensity in dilute solutions could not fully mimic the behaviours of mutated proteins in the crowded cytoplasm of the cells

Jing Li

Papers

Cataract-causing mutation R233H affects the stabilities of βB1- and βA3/βB1-crystallins with different pH-dependence.