Slitrk1, an Integral Membrane Protein, Controls Osteoblast Differentiation

Objectives: Slit and Trk-like protein 1 (Slitrk1) is a member of the SLITRK family of integral membrane proteins known to regulate neurodendrite elongation in the nervous system. Slitrk1 is associated with Tourette syndrome. Patients exhibit delayed bone formation and increased bone fracture tendency. However, the role of Slitrk1 in bone homeostasis is completely unknown. Here, we examined the role of Slitrk1 in osteoblastogenesis.
Methods: Murine femoral bone was immunostained with Slitrk1 antibody. Osteoblast differentiation was induced in osteoblast lineage cell cultures such as MC3T3-E1 cells, primary murine bone marrow stromal cells (BMSCs), and primary murine calvarial osteoblasts by treatment with ascorbic acid and β-glycerophosphate, or recombinant human (rh) bone morphogenetic protein 2 (BMP2). For formation of ectopic bone, collagen pellets soaked with 2 mg of rhBMP2 were implanted beneath the fascia of 8 weeks-old male mice. 10T1/2 cells were transfected with runt-related transcription factor 2 (Runx2) along with or without Slitrk1 expression vectors using lipofection methods. Endogenous Slitrk1 in MC3T3-E1 cells was knocked down by siRNA. Femoral bone volume of 6 month-old Slitrk1 null mice was assessed by micro CT.
Results: Slitrk1 was strongly expressed by osteoblasts in murine femoral bone. The expression levels of Slitrk1 increased with osteoblast differentiation in osteoblast lineage cells. Alkaline phosphatase (ALP) activity was decreased in Slitrk1 knock-down cells or BMSCs isolated from Slitrk1 null mice. In contrast, overexpression of Slitrk1 in C3H10T1/2 cells, which do not express Slitrk1 endogenously, led to increased mRNA levels of ALP and osteocalcin induced by Runx2. BMP2-induced ectopic bone formed in Slitrk1 null mice was smaller than in wild-type littermates. Femoral bone volume of Slitrk1 null mice showed a decreased tendency compared to control mice.
Conclusions: Taken together, these data suggest that Slitrk1 is expressed in osteoblast cells where it enhances osteoblast differentiation.