Cardiac Fpn knockouts.
The strategy for the generation of cardiac Fpn knockout mice is outlined in Fig. S4. Briefly, C57BL/6N mice harboring a knockout first mutation at the Fpn (Slc40a1) locus were obtained from the European Mouse Mutant Cell Consortium and were crossed with a C57BL/6 Flp recombinase deleter mouse, Tg(ACTB-FlpE)9205Dym/J, to remove the selection cassette and generate a Fpn allele with floxed exons 4 and 5, which encode the transmembrane domain. As a first line of validation, we crossed floxed mice (Fpn fl/fl) with mice transgenic for the ubiquitously expressed Pgk1-Cre recombinase, Tg(Pgk1-cre) (32), and confirmed embryonic lethality in ubiquitous homozygous knockouts (Fpn−/−). Cardiac Fpn knockouts then were generated by crossing homozygous Fpn fl/fl animals with mice transgenic for Myh6-Cre recombinase B6.FVB-Tg(Myh6-cre)2182Mds/J (33), which is under the control of cardiomyocyte-specific myosin alpha heavy-chain 6 promoter. The subsequent breeding strategy was designed to produce cardiac Fpn knockouts and homozygous floxed controls (hereafter referred to as “Fpn fl/fl Myh6.Cre+” and “Fpn fl/fl,” respectively) in the same litter.