Cataract is the leading cause of blindness worldwide (World Health Organisation, 2010). The exact mechanism behind cataract formation remains elusive, although previous studies have implicated an enigmatic zone known as the nucleocortical barrier (Sweeney and Truscott, 1998; Friedrich and Truscott, 2009; Friedrich and Truscott, 2010). It has been proposed that the presence of this barrier, which first appears around middle-age, prevents the diffusion of free radical scavengers such as glutathione into the nucleus of aged lenses, resulting in a cellular environment conducive to the accumulation of oxidants capable of protein modification and damage (Sweeney and Truscott, 1998; Truscott, 2005). Phosphorylation of αB-crystallin (αB) has been shown to increase the affinity of this small heat shock protein for destabilised substrate in vitro (Aquilina et al., 2004). In this study we have asked the question: does αB phosphorylation contribute to the formation of the barrier due to this heightened activity?

To investigate this, human lenses were either sectioned manually to isolate the cortical, barrier and nuclear regions, or sectioned on a cryostat for immunolabelling. Western blots of tissue protein extracts have provided a tentative order in which phosphorylation sites in αB are phosphorylated, suggesting that Ser59 is phosphorylated preferentially over Ser19 and Ser45. A strong pattern not seen in younger lenses has also been identified in relation to the number of phosphorylated residues between the different regions of an aged lens, with a marked increase in prevalence of singly phosphorylated αB, coupled with a decrease of doubly and triply phosphorylated αB towards the nucleus when compared to the cortex and barrier regions. In conjunction, membrane association of phosphorylated αB has been directly observed using immunoconfocal microscopy, with a specific zone between .5 and 1mm in from the cortex being identified as the point at which phosphorylated αB begins associating with membranes.

These results indicate that phosphorylation of αB initiates membrane association in a region that coincides with the location of the nucleo-cortical barrier. This association is potentially regulated by a single phosphorylation event, although the residue responsible is yet to be identified.