Lyn associated with chaperone machinery is longer protected from proteasome degradation in ChAc red cells than in healthy erythrocytes. Cytosol from control or ChAc red cells were incubated in the presence (upper 4 panels) or absence of the 20S proteosome inhibitor bortezomib (20 µM; lower 4 panels) for 12 or 36 hours and subsequently subjected to the separation procedure described in Figure 2B. Steady-state patterns of high-molecular-weight complexes from healthy and chorea-acanthocytosis cells at time zero (0h) is shown in supplemental Figure 4. Eighteen gradients were collected from the top and analyzed by immunoblotting with antibodies to Lyn, HSP90, and HSP70. Arrows represent glycerol gradient molecular mass standards: glutamate dehydrogenase (62 kDa), alcohol dehydrogenase (150 kDa), apoferritin (443 kDa), and thyroglobulin (669 kDa). The experiment shown is representative of 8 such experiments, each from an individual healthy or ChAc subject. Densitometric analysis is shown in supplemental Figure 5.