The accumulation of misfolded proteins in intracellular amyloid inclusions, characteristic of neurodegenerative disorders like Huntington's and prion diseases, is thought to result from the failure of cellular protein quality control mechanisms. This study examines the formation of misfolded protein inclusions in yeast and mammalian cell cultures, identifying two distinct intracellular compartments for the sequestration of misfolded cytosolic proteins. The partitioning of quality control substrates into these compartments depends on their ubiquitination status and aggregation state. Soluble ubiquitinated misfolded proteins accumulate in a juxtanuclear compartment where proteasomes are concentrated, while terminally aggregated proteins are sequestered in a perivacuolar inclusion. Notably, disease-associated Huntington and prion proteins are preferentially directed to the perivacuolar compartment. Enhancing ubiquitination of a prion protein promotes its delivery to the juxtanuclear inclusion. These findings provide a framework for understanding the preferential accumulation of amyloidogenic proteins in inclusions linked to human diseases.The accumulation of misfolded proteins in intracellular amyloid inclusions, characteristic of neurodegenerative disorders like Huntington's and prion diseases, is thought to result from the failure of cellular protein quality control mechanisms. This study examines the formation of misfolded protein inclusions in yeast and mammalian cell cultures, identifying two distinct intracellular compartments for the sequestration of misfolded cytosolic proteins. The partitioning of quality control substrates into these compartments depends on their ubiquitination status and aggregation state. Soluble ubiquitinated misfolded proteins accumulate in a juxtanuclear compartment where proteasomes are concentrated, while terminally aggregated proteins are sequestered in a perivacuolar inclusion. Notably, disease-associated Huntington and prion proteins are preferentially directed to the perivacuolar compartment. Enhancing ubiquitination of a prion protein promotes its delivery to the juxtanuclear inclusion. These findings provide a framework for understanding the preferential accumulation of amyloidogenic proteins in inclusions linked to human diseases.