Fig. 2: Slitrk5 is a negative regulator of hedgehog signaling in osteoblasts.

a Primary osteoblasts from WT and Slitrk5^−/− mice were treated with BSA control or 40 ng/ml BDNF and cultured in osteoblast differentiation medium. ALP activity was measured at day 6 of osteoblast differentiation. Data are presented as mean ± s.d. n = 5 biologically independent samples, two-tailed unpaired t test. b RT-PCR analysis of hedgehog signaling related gene expression at day 6 and day 12 of differentiation in WT and Slitrk5^−/− osteoblasts. n = 4 biologically independent samples, two-tailed unpaired t test. c C3H10T1/2 cells transfected with either control or Slitrk5 overexpression vectors were treated with BSA or 100 ng/ml SHH in serum-free medium for 48 h. Gli1, Ptch1, Hhip, and Slitrk5 mRNA levels were measured by RT-PCR. n = 4 biologically independent samples, two-tailed unpaired t test. d C3H10T1/2 cells transfected with GLI1-luc/Renilla together with either control vector or Slitrk5 overexpression vector were treated with BSA or 100 ng/ml SHH in serum-free medium for 36 h. Luciferase activity was measured to assess Hh signaling. n = 3 biologically independent samples, two-tailed unpaired t test. e Primary osteoblasts from WT and Slitrk5^−/− mice were treated with different doses of SHH and cultured in osteoblast differentiation medium. ALP activity was measured at day 8 of differentiation. Data are presented as mean ± s.d. n = 5 biologically independent samples. f Primary osteoblasts transduced with Slitrk5 or Gfp shRNAs were treated with the indicated doses of SHH and cultured in osteoblast differentiation medium. ALP activity was measured at day 8 of osteoblast differentiation. Data are presented as mean ± s.d. n = 2 biologically independent samples. g Knockdown efficiency of Slitrk5 was validated by qRT-PCR. n = 4 biologically independent samples, two-tailed unpaired t test.