In order to test whether the significant inhibition of TRPM8 channel activity by scPPX1 detected in the patch clamp and Ca2+ measurements is due to the alteration of the levels and/or localization of the TRPM8 protein, we performed
Western blot and immunocytochemical analyses on HEK-293 cells expressing TRPM8 alone, or with the enzyme (see Methods S1). In the immunocytochemistry experiments, TRPM8 showed both intracellular and plasma membrane localization, consistent with earlier studies [37]. Co-expression of scPPX1 did not alter the localization of TRPM8 (Fig. S1A) and did not decrease the amount of the protein as detected with Western blot (Fig. S1B).