Shao H. Yang et al.

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SUPPLEMENTAL FIGURES

Severe hepatocellular disease in mice lacking one or both protein prenyltransferases in the liver

Shao H. Yang et al.

Fig. I. Phenotypes of Fntbmice. (A) Body weight curves of male Fntb, Fntb, and Fntbmice (n = 12/genotype). (B) Body weight curves of female Fntb (n = 12), Fntb(n = 12), and Fntbmice (n = 14). Body weights in Fntbmice were lower than those of Fntbor Fntb mice (P < 0.0001 for both sexes by repeated-measures ANOVA). (C) Liver/body weight ratio of male and female Fntb and Fntbmice (n = 6 males and 6 females/genotype). Error bars indicate SEM.

Fig. II. Reduced frequency of binuclear hepatocytes in Fntb mice.(A, B) Primary hepatocytes from 3-month-old Fntb and Fntb mice, revealing fewer binuclear cells and increased numbers of cells with an enlarged nucleus in Fntb hepatocytes (black arrows). (C, D) Primary hepatocytes stained with DAPI, revealing an increased frequency of enlarged nuclei and misshapen nuclei in Fntb hepatocytes (red arrowheads). (E) Percentage of binuclear hepatocytes in 3-month-old Fntb and Fntb mice (n = 4 mice/group).Error bars indicate SEM.

Fig. III.Flow cytometry studies of primary hepatocytes from Fntb and Fntb mice. (A–C) Cell cycle acquisition analysis with FL2-W and FL2-A dot plots, revealing two populations(I and II) in Fntb hepatocytes (A, B) but only one population in Fntb hepatocytes (C). (D, E) DNA histogram of the two cell populations of Fntb hepatocytes (I and II). (F) DNA histogram of the main population of Fntb hepatocytes. The percentages of cells in G1, G2, and S phases are indicated.

Fig. IV. Phenotypes of Pggt1bmice. (A, B) Body weight curves for male (A) and female (B) Pggt1bandPggt1bmice (n = 8/genotype for males and 10/genotype for females). Body weights in Pggt1b and Pggt1bmice were not significantly different (repeated-measures ANOVA). (C) Enlarged liver and jaundice in a 3-month-old Pggt1bmouse and a littermate control mouse. (D) Liver/body weight ratio in Pggt1bandPggt1bmice (P < 0.0001; n = 6 males and 6 females mice/genotype). Error bars indicate SEM. (E) Kaplan-Meier survival plots for wild-type mice (n = 12), Fntb(n = 16),Pggt1b(n = 20), andFntbPggt1bmice (n = 8).

Fig.V. Morphological abnormalities in Pggt1bhepatocytes. (A, B) Primary hepatocytes isolated from 3-month-old Pggt1b (A) andPggt1b (B) mice, revealing that Pggt1b hepatocytes were smaller and rounder in shape (red arrows). (C, D) DAPI-stained hepatocyte nuclei, showing that nuclei of Pggt1bhepatocytes tended to be smaller. (E, F) Histochemical staining of primary hepatocytes from Pggt1bandPggt1bmice with palloidin (green), revealing loss of actin stress fibers in Pggt1b hepatocytes. (G, H) Immunohistochemical staining of primary hepatocytes for -tubulin (green), revealing an abnormal distribution of microtubules in Pggt1b hepatocytes. DNA was visualized with DAPI (blue).

Fig. VI. Body weight curves of wild-type and FntbPggt1bmice (n = 8 mice/genotype). Body weights in FntbPggt1bmice were lower than in wild-type mice (P < 0.0001 by repeated-measures ANOVA). Error bars indicate SEM.

Fig. VII. Reduced numbers of binuclear hepatocytes in FntbPggt1b mice.(A, B) Primary hepatocytes from 3-month-old wild-type and FntbPggt1b mice, revealing a reduced frequency of binuclearFntbPggt1b hepatocytes. (C, D) DAPI-stained primary hepatocytes, revealing that FntbPggt1b hepatocytes have more single enlarged nuclei and fewer cells containing two nuclei. (E) Percentage of binuclear hepatocytes in freshly isolated hepatocytes from 3-month-old wild-type and FntbPggt1b mice (n = 3 mice/group). Error bars indicate SEM.