Fig. 3. Mff recruits DLP1 to peroxisomes.

(A) Control fibroblasts were treated for 72 h with control dsRNA (upper panel) or MFF #1 dsRNA (lower panel) and then stained with antibodies to Mff (a,e), Pex14p (b,f), and DLP1 (c,g); the merged view of Pex14p and DLP1 is shown (d,h). Scale bar: 10 µm. Insets, higher magnification images of the boxed regions, scale bar: 2 µm. (B) Control fibroblasts were treated for 72 h with control dsRNA or MFF #1 dsRNA. Mff levels were assessed by immunoblotting with anti-Mff antibody. Actin was used as a loading control. (C) In HeLa cells, HA₂-DLP1 was expressed (a–d) or co-expressed with FLAG-MFF (e–h), FLAG-MFFΔTMD (i–l), or FLAG-MFFΔN (m–p). After 24 h, cells were stained with antibodies to Pex14p (a,e,i,m), FLAG (b,f,j,n) and HA (c,g,k,o); the merged view of the three proteins is shown (d,h,l,p). Scale bar: 10 µm. Insets, higher magnification images of the boxed regions, scale bar: 2 µm. (D) HEK293 cells were treated with 0.5 mM DSP and subjected to immunoprecipitation using Mff antiserum (α-Mff, lane 3) or a preimmune serum (preimmune, lane 2). Immunoprecipitates were analyzed by SDS-PAGE and immunoblotting with antibodies to DLP1, Mff, and Fis1. Input (5%) was loaded in lane 1.