Alterations in intranuclear localization of Runx2 affect biological activity

The transcription factor Runx2 controls osteoblast proliferation and differentiation. Runx2 organizes and assembles gene-regulatory complexes in nuclear microenvironments where target genes are activated or suppressed in a context-dependent manner. Intranuclear localization of Runx2 is mediated by the nuclear matrix-targeting signal (NMTS), an autonomous motif with a loop (L1)-turn-loop (L2) structure that forms predicted protein-protein interaction surfaces. Here we examined the functional consequences of introducing mutations in the L1 and L2 loops of the NMTS. These mutant proteins enter the nucleus, interact with the hetero-dimeric partner Cbfbeta, and bind to DNA in vitro and in vivo. In addition, these mutants retain interaction with the carboxy-terminus interacting co-regulatory proteins that include TLE, YAP, and Smads. However, two critical mutations in the L2 domain of the NMTS decrease association of Runx2 with the nuclear matrix. These subnuclear targeting defective (STD) mutants of Runx2 compromise target gene activation or repression. The biological significance of these findings is reflected by decreased osteogenic differentiation of mesenchymal progenitors, concomitant with major changes in gene expression profiles, upon expression of the STD Runx2 mutant. Our results demonstrate that fidelity of temporal and spatial localization of Runx2 within the nucleus is functionally linked with biological activity.