The DNA Helicase Recql4 Is Required for Normal Osteoblast Expansion and Osteosarcoma Formation

Abstract

<div><p><i>RECQL4</i> mutations are associated with Rothmund Thomson Syndrome (RTS), RAPADILINO Syndrome and Baller-Gerold Syndrome. These patients display a range of benign skeletal abnormalities such as low bone mass. In addition, RTS patients have a highly increased incidence of osteosarcoma (OS). The role of RECQL4 in normal adult bone development and homeostasis is largely uncharacterized and how mutation of <i>RECQL4</i> contributes to OS susceptibility is not known. We hypothesised that Recql4 was required for normal skeletal development and both benign and malignant osteoblast function, which we have tested in the mouse. <i>Recql4</i> deletion <i>in vivo</i> at the osteoblastic progenitor stage of differentiation resulted in mice with shorter bones and reduced bone volume, assessed at 9 weeks of age. This was associated with an osteoblast intrinsic decrease in mineral apposition rate and bone formation rate in the <i>Recql4</i>-deficient cohorts. Deletion of <i>Recql4</i> in mature osteoblasts/osteocytes <i>in vivo</i>, however, did not cause a detectable phenotype. Acute deletion of <i>Recql4</i> in primary osteoblasts or shRNA knockdown in an osteoblastic cell line caused failed proliferation, accompanied by cell cycle arrest, induction of apoptosis and impaired differentiation. When cohorts of animals were aged long term, the loss of <i>Recql4</i> alone was not sufficient to initiate OS. We then crossed the Recql4^fl/fl allele to a fully penetrant OS model (<i>Osx</i>-Cre <i>p53^fl/fl</i>). Unexpectedly, the <i>Osx</i>-Cre <i>p53^fl/flRecql4^fl/fl</i> (dKO) animals had a significantly increased OS-free survival compared to <i>Osx</i>-Cre <i>p53^fl/fl</i> or <i>Osx</i>-Cre <i>p53^fl/flRecql4^fl/+</i> (het) animals. The extended survival was explained when the Recql4 status in the tumors that arose was assessed, and in no case was there complete deletion of <i>Recql4</i> in the dKO OS. These data provide a mechanism for the benign skeletal phenotypes of <i>RECQL4</i> mutation syndromes. We propose that tumor suppression and osteosarcoma susceptibility are most likely a function of mutant, not null, alleles of <i>RECQL4</i>.</p></div