Figure 6. ERdj3–client protein complexes are co-secreted when BiP activity is limiting.

1. A Representative immunoblot of M2 anti-FLAG immunopurification of media conditioned for 24 h on HEK293^DAX cells co-overexpressing ^FTTTR^A25T and ERdj3^WT or ERdj3^H53Q. Media contained vehicle or trimethoprim (TMP, 10 μM, 16 h; activates ATF6), as indicated. Media inputs (1:400) are shown as a control. Lysates from these cells are also shown to confirm the increase of the ATF6 target proteins BiP and GRP94 in cells following TMP-dependent activation of ATF6.
2. B Bar graph depicting quantification of the change in the amount of ERdj3 co-immunoprecipitating with TTR^A25T following activation of ATF6 as in (A). The ERdj3/TTR ratio is normalized to the ratio in the absence of ATF6 activation. *P-value < 0.05; n = 3, mean ± SEM.
3. C Representative immunoblot of M2 anti-FLAG immunopurification of media conditioned for 24 h on HEK293T cells co-overexpressing ^FTTTR^A25T, ERdj3^WT, ERdj3^H53Q, and/or BiP as indicated. Media inputs (1:400) are shown as a control. Lysates from these cells are also shown to confirm the increase of BiP afforded by overexpression.
4. D Quantification of immunoblots of M2 anti-FLAG immunopurification of media collected from cells co-overexpressing either ERdj3^WT or ERdj3^H53Q, ^FTTTR^A25T, and BiP as in (C). The ERdj3/TTR ratio is normalized to the ratio in the absence of BiP overexpression. **P-value < 0.01; n = 5, mean ± SEM.
5. E Immunoblot of 6E10 anti-APP immunopurifications from the conditioned media of CHO-derived APP₇₅₁-expressing (7WD10) cells overexpressing ERdj3^WT, ERdj3^H53Q, and BiP as indicated. Media inputs (1:100) are shown as a control. Lysates from these cells are also shown to confirm the increase of BiP afforded by overexpression.

Source data are available online for this figure.