Composition and activity of the proteasome in human iPSC-derived neuronal model of early-stage sporadic Alzheimer's disease

Abstract

IntroductionThe proteasome is a critical cellular degradative machinery impaired in late-stage Alzheimers disease (AD). However, the status and activity of the proteasome in early-stage sporadic AD (sAD) is unknown. MethodsA cellular model of human early-stage sAD was generated from sAD patient iPSC-derived cortical neurons by dual-SMAD inhibition. The iPSCs, neuroprogenitors, and cortical neurons were validated by the expressions of key markers. The level of total intraneuronal A{beta} was measured by ELISA. Composition and native proteolytic activities of the proteasome in control and sAD cortical neurons were measured using complementary fluorogenic probes. ResultsControl and sAD patients iPSCs expressed pluripotent markers OCT4, NANOG, and SSEA4 which induced into neuroprogenitors expressing NESTIN and PAX6. The neuroprogenitors terminally differentiated into cortical neurons expressing neuronal markers MAP2 and TUJ1, and cortical layer marker TBR1. The level of intraneuronal A{beta} in the sAD cortical neurons was significantly higher compared to control. Control and sAD cortical neurons expressed native 30S, 26S, and 20S proteasome assemblies with the sAD cortical neurons displaying higher 20S assemblies. Increased active 20S assemblies was associated with higher {beta}1, {beta}2, and {beta}5 proteolytic sites activities. DiscussionThe significant elevation in the proteolytic activities of the {beta}1, {beta}2, and {beta}5 subunits of 20S proteasome in sAD cortical neurons suggests that this may be a possible compensatory response to elevated intraneuronal A{beta}. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=120 SRC="FIGDIR/small/734021v1_ufig1.gif" ALT="Figure 1"> View larger version (30K): org.highwire.dtl.DTLVardef@1d8c382org.highwire.dtl.DTLVardef@b92e8org.highwire.dtl.DTLVardef@1d9c699org.highwire.dtl.DTLVardef@7d826d_HPS_FORMAT_FIGEXP M_FIG C_FIG