Intranuclear ataxin1 inclusions contain both fast- and slow-exchanging components

Abstract

A hallmark of neurodegenerative diseases caused by polyglutamine expansion is the abnormal accumulation of mutant proteins into ubiquitin-positive inclusions1. The local build-up of these ubiquitinated proteins suggests that the proteasome machinery inadequately clears misfolded proteins, resulting in their increase to potentially toxic levels2. Inclusions may disrupt normal cell homeostasis by sequestering vital cellular factors, such as chaperones, proteasomes and transcription components3, 4, 6, 7. Here, we used fluorescence recovery after photobleaching (FRAP) to examine the intranuclear dynamics of polyglutamine-expanded ataxin1 and inclusion-associated proteins. These experiments demonstrated that at least two types of ataxin1 inclusions exist; those that undergo rapid and complete exchange with a nucleoplasmic pool and those that contain varying levels of slow-exchanging ataxin1. Slow-exchanging inclusions contain high ubiquitin levels, but surprisingly low proteasome levels, suggesting an impairment in the ability of proteasomes to recognize ubiquitinated substrates. Proteasomes and CBP remained highly dynamic components of inclusions, indicating that although enriched with ataxin1, they are not irreversibly trapped. These results redefine our perception of polyglutamine inclusions and demonstrate the usefulness of FRAP and live cell imaging to study factors that modulate their behaviour.