Pi 103

Recombinant human insulin was purchased from Novo Nordisk (Bagsvaerd, Denmark). Akti1/2 was from Merck Millipore (Darmstadt, Germany), and PI-103 from Enzo Life Sciences (Farmingdale, NY, USA). Protein G-Sepharose was from GE Healthcare (Buckinghamshire, UK). High-fat diet (60 kcal% fat, Cat. No. 12492) was purchased from Research Diets (USA). All other chemicals were from Sigma-Aldrich (Shanghai, China) or Sangon Biotech (Shanghai, China). The commercial antibodies and resins used are listed in Supplementary Table 1. For immunoblotting, the commercial antibodies were used at 1:1000 dilution. The antibody recognising pSer¹⁰⁴³-TRIM24 was generated at ABclonal (Wuhan, China) and used at 1 μg/ml.

FR900359 was isolated as described previously (Schrage et al., 2015 (link)). Salts and drugs have been purchased from Sigma-Aldrich (Germany) with the exception of PTX and TGX 221, which were obtained from Merck Millipore (Calbiochem) (Germany) and PI-103 from Enzo Life Sciences. The PI3K inhibitors TGX-221, AS-252424, and PI-103 were administered at submaximal concentrations to achieve biological effects (Ali et al., 2008 (link); Tsvetkov et al., 2016a (link)). All drugs were freshly dissolved on the day of the experiment according to the material sheet. The relaxin peptides have been purchased from Sigma-Aldrich (Germany). Relaxins were dissolved in water. The following concentrations were used: relaxin-1 at 10 pM or at 100 pM, relaxin-2 at 10 pM or at 100 pM or from 1 pM to 10 nM, relaxin-3 at 10 pM or at 100 pM.

Recombinant human insulin was purchased from Novo Nordisk (Bagsvaerd, Denmark). High fat diet (60 kcal% fat, Cat. No. 12492) was from Research Diets (USA). Protein G-Sepharose was bought from GE-Healthcare (Little Chalfont, Buckinghamshire, UK). Precast NuPAGE^® Bis-Tris gels were from Thermo Fisher Scientific (Waltham, MA, USA). Akti1/2 was from Merck Millipore (Darmstadt, Germany), and PI-103 was from Enzo Life Sciences (Farmingdale, NY, USA). All other chemicals were from Sigma-Aldrich (Shanghai, China) or Sangon Biotech (Shanghai, China). The commercial antibodies and resins are listed in Supplementary Data 3. The antibodies recognizing RalGAPα1 and RalGAPα2 were described previously^{31 (link)}, and the pThr⁴⁸⁴-SERCA2a antibody was as previously reported^{16 (link)}.