Abstract

Karyopherin-{beta}2 (Kap{beta}2) is a nuclear-import receptor that recognizes proline-tyrosine nuclear localization signals (PY-NLSs) of diverse cytoplasmic cargo for transport to the nucleus. Kap{beta}2 cargo include several disease-linked RNA-binding proteins (RBPs) with prion-like domains (PrLDs), such as FUS, TAF15, EWSR1, hnRNPA1, and hnRNPA2. These RBPs with PrLDs are linked via pathology and genetics to debilitating degenerative disorders, including amyotrophic lateral sclerosis (ALS), frontotemporal dementia (FTD), and multisystem proteinopathy (MSP). Remarkably, Kap{beta}2 prevents and reverses aberrant phase transitions of these cargo, which is cytoprotective. However, the molecular determinants of Kap{beta}2 that enable these activities remain poorly understood, particularly from the standpoint of nuclear-import receptor architecture. Kap{beta}2 is a superhelical protein comprised of 20 HEAT repeats. Here, we design truncated variants of Kap{beta}2 and assess their ability to antagonize FUS aggregation and toxicity in yeast and FUS condensation at the pure protein level and in human cells. We find that HEAT repeats 8-20 of Kap{beta}2 recapitulate all salient features of Kap{beta}2 activity. By contrast, Kap{beta}2 truncations lacking even a single cargo-binding HEAT repeat display reduced activity. Thus, we define a minimal Kap{beta}2 construct for delivery in adeno-associated viruses as a potential therapeutic for ALS/FTD, MSP, and related disorders.