FIGURE 5.

Follow-up analysis of the identified proteins. A, immunostaining of elution fractions (before protease digestion) prepared from PC12 cells expressing either HttQ74-GFP or HttQ23-GFP. Anti-TDP-43 or anti-FUS antibodies were used. B, SDD-AGE analysis of 74D-694 [RNQ+] and [rnq-] strains that expressed the GAL1-controlled RNQ1-GFP for 5 h. As a positive control, 74D-694 [RNQ+] with overproduction of Sis1 was used. Immunostaining was performed with anti-Sis1 antibodies (Ab, left) or anti-Rnq1 antibodies (right). HMW, high molecular weight aggregates. C and D, analysis of amyloid-forming potential of the identified proteins. Eight common proteins (HA-tagged, from the yeast ORF library) were expressed in BY4741. Lysates were treated with 1% SDS and loaded on an acrylamide gel for SDS-PAGE (C) or directly on PVDF membrane (D, to estimate the expression level) followed by immunostaining with the anti-HA antibodies. In C, only a portion of the gel just below the loading wells is shown.