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Figure S1. hFPCL. a b c. -Tet. 804 Myc Merge. d e f. +Tet. 804 Myc Merge.
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Figure S1 hFPCL a b c -Tet 804MycMerge d e f +Tet 804 MycMerge Figure S1. Characterization of full-length human FPC construct stably expressed in LLC-PK1 (hFPCL) cells under tetracycline induced conditions. Both 804 and myc tag (d-f) antibodies detect the overexpression of FPC in tetracycline induced LLC-PK1 cells (hFPCL) but not in the non-induced controls (a-c).
Figure S2 A B Differentiated hFPCL cells + Tet Nondifferentiated hFPCL cells a b c a b c 3-day post-conf. Mitotic - Tet Myc 4883Merge 4883 Acetyl--tub Merge Myc 4883Merge d e f d e f Interphase Myc 4883Merge 4883 Myc Merge g h i g h i 7-day post-conf. Mitotic + Tet 4883 Acetyl--tub Merge Myc DAPI Merge j k l j k l Interphase Myc 4883Merge 4883 Myc Merge Figure S2. Recombinant FPC-myc is targeted to the primary cilia but not to the mitotic spindles in LLC-PK1 cells stably expressing full-length FPC. (A) The signal of overexpressed FPC-myc was detected in cilia in cells 3-day post-confluence (a-c, d-f). The signal is stronger in cells cultured 7-day post-confluence (g-i, j-l), when double-stained with 4883 and acetylated--tubulin or 4883 and myc tag antibodies. (B) 4883 but not c-myc antibody stained endogenous FPC on the mitotic spindle (a-c) in hFPCL cells without induction (a-f). Anti-myc tag antibody also did not detect a specific signal on the mitotic spindle (g) in tetracycline induced hFPCL cells.
Figure S3 Telophase Anaphase Metaphase Prophase mIMCD3 a b c d e f g h i j k l 4883 Acetyl--tub Merge Figure S3. FPC localization during mitosis in mIMCD3 cells. FPC (labeled by FPC polyclonal antibody 4883) is found on the MTOC at prophase, the mitotic spindle at metaphase, anaphase and the midbody at telophase. Acetylated -tubulin serves as the MTOC, spindle and midbody marker.
Figure S4 WT4 -tub (DAPI) S26 -tub (DAPI) Figure S4. Multipolarity in FPC knockdown mIMCD3 cells.Lower magnification figure shows that, unlike in WT4 control cells where most dividing cells undergo bipolar division (a, arrow), many cells undergo aberrant cell division in FPC knocked down cells (b, arrow and arrow head).
Figure S5 Non-cystic kidney (4/98A) Cystic kidney (25/01) * Merge Merge -tubulin Pericentrin DAPI -tubulin Pericentrin DAPI Figure S5. Centrosome amplification in kidneys from human ARPKD patients. The centrosome was stained with two centrosomal markers, -tubulin and pericentrin. Centrosome amplification was rarely observed in the cystic epithelial cells (right panel, ID. 25/01), but never seen in the control non-cystic kidneys (left panel. ID. 4/98A).
Figure S6 LLC-PK1 4883 Figure S6. FPC is upregulated in mitotic LLC-PK1 cells. Endogenous FPC expression was detected by 4883 antibody in asynchronized LLC-PK1 cells. FPC is obviously increased in the cells at metaphase (white arrow), compared to the interphase cells (yellow arrow). DAPI Merge
Figure S7 a b c Merge Acetyl--tub 804 d e f Merge Acetyl--tub 804 g h i Merge Acetyl--tub 804 j k l -tub Merge 804 Figure S7. 804 recognizes endogenous FPC on the cilia and basal bodies but not that on the centrosomes in LLCPK-1 cells. FPC N-terminal antibody 804 stains cilia (marked by acetyl--tub) and basal bodies in LLCPK-1 cells under different differentiation status (a-i and j-l, bottom insets). 804 does not stain endogenous FPC on the centrosomes (labeled by -tub) in an undifferentiated cell (j-l, top insets) but does label next to one of the two dots, probably the basal body of a short cilium or the mother centriol in a more differentiated cell (j-l, bottom insets).
