FIG 1.

Generation of the PDKO mouse model. (A) Dematin is homologous to the villin-family of proteins. Dematin shares its headpiece domain with villin and supervillin, but its core domain is characteristically distinct. Unique gelsolin repeats are labeled as “G” in villin and supervillin. Dematin shares its core domain with abLIM and UNC115 but lacks the LIM domains labeled as “L.” (B) Mice homozygous for the DMTN (dematin) floxed allele containing loxP sites are crossed with Pf4-Cre expressing mice to recombine exons 5 through 8 of the DMTN gene. This yields a frame shift creating a premature stop codon in exon 10 generating an unstable dematin transcript that degrades and is not translated. (C) Genotyping strategy for PDKO mice. Pf4-Cre genotyping was performed using the Pf4-Cre forward primer: CCA AGT CCT ACT GTT TCT CAC TC and the Pf4-Cre reverse primer TGC ACA GTC AGC AGG TT resulting in an expected 400 bp amplicon. DMTN floxed/floxed genotyping was tested using DMTN floxed/floxed forward primer: GCC GGC TGA CTT AAG TGG GAT CC and DMTN floxed/floxed reverse primer: GTT TTC CAG GGT GAC AGC TGT TCA resulting in an expected 351 bp amplicon. Lane 1: DNA marker, Lane 2: PDKO mouse Pf4-Cre genotyping, Lane 3: WT mouse Pf4-Cre genotyping, Lane 4: DMTN floxed/floxed genotyping. (D) Dematin protein expression in WT and PDKO RBCs and platelets. The expected 48 kDa size band is marked by a red asterisk in platelet samples. (E) Wright stain of WT and PDKO blood smears.