Runx2 deficiency and defective subnuclear targeting bypass senescence to promote immortalization and tumorigenic potential

Abstract

The osteogenic Runt-related (Runx2) transcription factor negatively regulates proliferation and ribosomal gene expression in normal diploid osteoblasts, but is up-regulated in metastatic breast and prostate cancer cells. Thus, Runx2 may function as a tumor suppressor or an oncogene depending on the cellular context. Here we show that Runx2-deficient primary osteoblasts fail to undergo senescence as indicated by the absence of ␤-gal activity and p16 INK4a tumor suppressor expression. Primary Runx2-null osteoblasts have a growth advantage and exhibit loss of p21 WAF1/CIP1 and p19 ARF expression. Reintroduction of WT Runx2, but not a subnuclear targeting-defective mutant, induces both p21 WAF/CIP1 and p19 ARF mRNA and protein resulting in cell-cycle inhibition. Accumulation of spontaneous phospho-H2A.X foci, loss of telomere integrity and the Mre11/Rad50/Nbs1 DNA repair complex, and a delayed DNA repair response all indicate that Runx2 deficiency leads to genomic instability. We propose that Runx2 functions as a tumor suppressor in primary diploid osteoblasts and that subnuclear targeting contributes to Runx2-mediated tumor suppression.cancer ͉ genomic instability ͉ growth control ͉ nuclear organization ͉ osteoblast W hen exposed to inappropriate growth signals, mammalian cells engage in elaborate mechanisms that provide alternative fates to prevent tumorigenesis. These defenses include apoptosis and senescence, which guard against unrestrained proliferation. Several tumor suppressors, including the retinoblastoma protein (pRb), p53, and the products of the INK4b-ARF-INK4a locus (p15-p19-p16), control telomere integrity, cellular senescence, and genomic stability (1-4). Tumors arise when cells override these intrinsic defenses. Cancer cells typically exhibit independence from extrinsic growth signals, lack of senescence or apoptosis, compromised telomeric integrity, and genomic instability, as well as changes in nuclear organization (2, 4-6). However, mice with genetic ablations of tumor suppressors are prone to tumorigenesis that is often lineage-restricted (7, 8). These observations suggest that lineage-specific mechanisms contribute to tumorigenesis.Runt-related (Runx) transcription factors determine cell fate and regulate lineage-specific proliferation and differentiation (9-11). Genetic studies reveal that Runx1 is essential for definitive hematopoiesis, Runx2 is required for osteogenesis, and Runx3 is involved in gut development as well as neurogenesis (12-16). Runx proteins support epigenetic regulation at mitosis and control ribosomal gene expression during the cell cycle (9)(10)(11)(17)(18)(19). Runx transcription factors are nuclear-scaffolding proteins that integrate signaling pathways by organizing macromolecular complexes in nuclear microenvironments and facilitating chromatin remodeling (20). Mice with gene replacements expressing subnuclear targeting defective mutants (mSTDs) of Runx1 or Runx2 from the native loci exhibit phenotypes identical to mice in which the gene has been ablated (21, 22), sugge...

TNF-α's effects on proliferation and apoptosis in human mesenchymal stem cells depend on RUNX2 expression

“…Work by Zaidi and colleagues (12) has demonstrated that RUNX2 is important for the expression of p21, p16, and p19, which mediated the antiproliferative mechanisms in osteoblasts by the blockade of cyclin action. (16,17) It was clearly noticeable that at 0.5% FCS, the rise in RUNX2 (Fig.…”

“…Since cyclin D1 is involved in the G 0 /G 1 transition, our results suggest that inhibition of RUNX2 activity accelerates exit from G 1 and entry into S phase but does not change cell exit from G 0 . Moreover, Zaidi and colleagues (12) have shown that primary RUNX2 À/À osteoblasts exhibited loss of p21, p19, and p16, which all regulate the activity of the cyclin/cyclin-dependent kinase complex. (12) In fact, p16 and p19 reportedly inhibit cyclin D (19,20) and p21 inhibits cyclins D, E, and A.…”

“…Moreover, Zaidi and colleagues (12) have shown that primary RUNX2 À/À osteoblasts exhibited loss of p21, p19, and p16, which all regulate the activity of the cyclin/cyclin-dependent kinase complex. (12) In fact, p16 and p19 reportedly inhibit cyclin D (19,20) and p21 inhibits cyclins D, E, and A. (21,22) Our results showed clearly that RUNX2 siRNA provoked strong inhibition of p21 in hMSCs (p16 and p19 were not expressed in these cells).…”

“…(7) The involvement of RUNX2 as a regulator of osteoblast proliferation also was consistent with data obtained in other biologic contexts. (8)(9)(10)(11) More recently, Zaidi and colleagues (12) proposed that RUNX2 functions as a tumor suppressor in primary diploid osteoblasts, whereas Eliseev and colleagues (13) suggested that RUNX2 acts as a proapoptotic factor.…”

Subnuclear domain proteins in cancer cells support transcription factor RUNX2 functions in DNA damage response

“…HDACs), co-regulators (e.g. TLE-1, a homolog of groucho), YAP and SMADs (Zaidi et al, 2002;Zaidi et al, 2004) Disruption of RUNX protein subnuclear targeting alters transcriptional programs and compromises cell growth and differentiation (Zaidi et al, 2006), reflecting its pathological linkage to acute myelogenous leukemia and breast and prostate cancers (Zaidi et al, 2007b).…”