Fig. 7
POPDC1 and POPDC2 undergo heteromeric complex formation. a Adult mouse ventricular cardiomyocytes immunostained for POPDC1 (red) and POPDC2 (green). b PLA of POPDC1 and POPDC2 in transverse sections of right atrium and left ventricle from wild-type and Popdc1/Popdc2 knockout mice. Sections were counterstained with WGA (green) and DAPI. c and d Co-precipitation of c POPDC2-FLAG alone or together with POPDC1-MYC or POPDC3-MYC, respectively, or d POPDC1-HA alone or together with POPDC3-MYC. e Co-precipitation of ventricular tissue lysates of Popdc2 null mutant and wild-type mice. f Western blot of lysates from COS-7 cells expressing POPDC1-CFP and/or POPDC2-FLAG (left), or POPDC1-CFP and/or POPDC3-MYC (right). (x) monomer, (*) homodimer, (o) heterodimer (#) heterotetramer. g Quantitative Type-1 BRET saturation curves of POPDC1 and POPDC2 homo- and heteromeric complexes (N ≥ 2). h and i TREK-1 current in Xenopus oocytes expressing TREK-1 alone or together with POPDC1, POPDC2, or both. h Examples of TREK-1 current in response to a voltage jump from − 80 to 40 mV and i relative current amplitudes without or with theophylline (+ theo). Number of oocytes are given in each graph. Data are presented as mean ± SEM. *p < 0.05; ***p < 0.001. j and k Chimeric constructs containing j the N-terminal and transmembrane domains of POPDC1 and the cytoplasmic region of POPDC2 or k N-terminal and transmembrane domains of POPDC2 and the cytoplasmic region of POPDC1 were co-transfected with POPDC1 or POPDC3, respectively and subjected to co-precipitation analysis. l Truncations were introduced into POPDC1-MYC and subjected to co-precipitation analysis in COS-7 cells after co-transfection with POPDC2-FLAG. m A POPDC2-FLAG construct truncated to residue W188 was subjected to co-precipitation analysis after co-expression with POPDC1-MYC. n BiFC signal after co-expression of wild-type POPDC1 and POPDC2, or of POPDC1 p.V183F, p.Q153X, p.S201F and POPDC2 p.W188X mutants in HEK293 cells. POPDC1-VN155 + POPDC2-VN155 was used as a negative control. ***p < 0.001; ****p < 0.0001