Figure 6.

MRTF silencing promotes ciliogenesis even in the presence of serum, partially via a transcription-independent mechanism

(A and B) LLC-PK1 cells were transfected with siNR or siMRTF and continuously kept in serum-containing medium for 48 hr before processing for immunofluorescence. Cells were stained with primary cilia markers Acetylated tubulin (A) or Arl13b (B). Scale bar: 20 μm.

(C) The percentage of ciliated cells was determined using both cilium markers (mean ± SD, n = 6, ≈200 cells/condition). ∗∗∗p < 0.0005, ∗∗∗∗p < 0.0001 using t test.

(D) LLC-PK1 cells were first transfected with the porcine-specific MRTF (psMRTF) siRNA, then re-transfected with siRNA-resistant (murine) HA-tagged wild-type (WT) or transcriptionally inactive (TL) truncation mutant of MRTF. Cells were then co-stained with anti-HA (to visualize MRTF-retransfection) and acetylated tubulin (to visualize the PC). Scale bar 20 μm.

(E) Cells possessing a long (>5μm) PC and expressing either HA WT or HA TL MRTF were counted and compared to the percentage of cells with a long PC without expressing re-transfected MRTF construct on the same coverslip (i.e. percentage of the neighboring control cells); mean ± SD, n = 3, ≈100 cells/condition in each experiment).∗∗∗∗p < 0.0001, t test.