Supplementary Fig. 7. Serotonin enhances Hsc70

B WT-LP

GAPDH

1.5 −

1.2 −

1.0 −

Ondansetron

0.8 −

m-CPBG

0.6 −

Serotonin

0.4 − 0.2 − 0.0 −

Hsc70 4 4 4 4 4

m-CPBG Serotonin Hsc70 Pan cadherin EEA1

m-CPBG Serotonin Hsc70 EEA1 Pan cadherin

NS

1.2 −

1.5 −

0.0 −

4 4 4 4 4 1 2 3 4 5

1.2 −

NS

1.0 − 0.8 − 0.6 − 0.4 − 0.2 − 0.0 −

4 4 4 4 4 1 2 3 4 5

(b) Membrane preparation *

* Ondansetron

1.0 −

m-CPBG Serotonin

0.5 − 0.0 −

(c) Cytosol preparation

Ondansetron

GAPDH

1 2 3 4 5

(b) Membrane preparation

Ondansetron

(a) Whole cell preparation Hsc70 protein expression (A.U.) (normalized to GAPDH)

Hsc70

NS

Hsc70 protein expression (A.U.) (normalized to pan cadherin)

Serotonin

Hsc70 protein expression (A.U.) (normalized to GAPDH)

m-CPBG

Hsc70 protein expression (A.U.) (normalized to pan cadherin)

Ondansetron

1.2 −

Hsc70 protein expression (A.U.) (normalized to EEA1)

(a) Whole cell preparation

Hsc70 protein expression (A.U.) (normalized to EEA1)

A WT-NP

Hsc70 4 4 4 4 4 1 2 3 4 5 NS

1.0 −

Serotonin

0.4 −

1.0 − 0.5 −

(c) Cytosol preparation

m-CPBG

0.6 −

0.0 −

EEA1

Ondansetron

0.8 −

0.2 −

Pan cadherin

NS

Hsc70 4 4 4 4 4

EEA1

1 2 3 4 5

Pan cadherin

1.0 − 0.8 − 0.6 − 0.4 − 0.2 − 0.0 −

4 4 4 4 4 1 2 3 4 5

Supplementary Fig. 7. Serotonin enhances Hsc70-mediated Kv4.3 membrane trafficking in WT-NP (A), but not in WT-LP mice (B). (a) No effect of serotonin or m-CPBG stimulation on Hsc70 protein expression in whole cell preparations for both WT-NP and WT-LP mice. (b) Plasma membrane preparations of ventricular myocytes showing greater Hsc70 plasma membrane expression, following serotonin and m-CPBG stimulation in WT-NP, but not in WT-LP mice. This effect was abolished by co-incubation with ondansetron. (c) No effect of serotonin or m-CPBG stimulation on Hsc70 levels in cytosolic preparations. Blots are representative of four experiments, with the input and pharmacological interventions outlined above each panel. Bar graphs represent the Kv4.3 expression level. *p