The nonreceptor isoform of tyrosine phosphatase epsilon (cyt-PTPe) supports osteoclast adhesion and activity in vivo, leading to increased bone mass in female mice lacking PTPe (EKO mice). The structure and organization of the podosomal adhesion structures of EKO osteoclasts are abnormal; the molecular mechanism behind this is unknown. We show here that EKO podosomes are disorganized, unusually stable, and reorganize poorly in response to physical contact. Phosphorylation and activities of Src, Pyk2, and Rac are decreased and Rho activity is increased in EKO osteoclasts, suggesting that integrin signaling is defective in these cells. Integrin activation regulates cyt-PTPe by inducing Src-dependent phosphorylation of cyt-PTPe at Y638. This phosphorylation event is crucial because wild-type-but not Y638F-cyt-PTPe binds and further activates Src and restores normal stability to podosomes in EKO osteoclasts. Increasing Src activity or inhibiting Rho or its downstream effector Rho kinase in EKO osteoclasts rescues their podosomal stability phenotype, indicating that cyt-PTPe affects podosome stability by functioning upstream of these molecules. We conclude that cyt-PTPe participates in a feedback loop that ensures proper Src activation downstream of integrins, thus linking integrin signaling with Src activation and accurate organization and stability of podosomes in osteoclasts. INTRODUCTIONOsteoclasts are large multinucleated cells of hematopoietic origin that degrade bone matrix. To perform this function osteoclasts must adhere firmly to bone using specialized adhesion structures called podosomes (Geiger et al., 2001;Gimona and Buccione, 2006;Linder, 2007;Teitelbaum, 2007). Podosomes are punctate structures that contain an actin-rich core surrounded by a ring of associated proteins, which convey the signal generated by contact with matrix to the actin core and the actin cytoskeleton. Proteins present in the podosomal ring include integrins and associated molecules such as vinculin, paxillin, Cbl, Cas, Src, Pyk2, and the small GTPases Rho, Rac, and CDC42 (Linder and Aepfelbacher, 2003;Gimona, 2008).In addition to osteoclasts podosomes are found in, among other cell types, macrophages, dendritic cells, and in endothelial and epithelial cells (Linder, 2007;Gimona et al., 2008). In osteoclasts, however, the organization of podosomes is linked to the ability of the cell to fulfill its physiological role.In cultured osteoclasts that are not actively resorbing bone, podosomes are scattered at random. Podosomes can assemble into clusters that grow and transform into dynamic rings, which further expand to form a large superstructure at the cell periphery that is characteristic of mature, boneresorbing cells. In osteoclasts grown on degradable matrix, this peripheral, belt-like superstructure, referred to as the sealing zone, contains densely packed podosomes that are usually not individually discernible. In osteoclasts grown on nondegradable surface, podosomes are arranged in a lesscrowded sealing zone-like structure (SZL)...

Background: Protein-tyrosine phosphatase ⑀ (cyt-PTPe) dephosphorylates and activates Src to promote osteoclast adhesion. Results: The adaptor protein GRB2 binds phosphorylated cyt-PTPe and recruits Src, thus promoting its activation. Conclusion: GRB2 links cyt-PTPe to Src downstream of activated integrins in osteoclasts.Significance: GRB2-mediated recruitment may constitute a general mechanism by which cyt-PTPe activates Src and related kinases.

The closely related tyrosine phosphatases PTPa and PTPe fulfill distinct roles in osteoclasts. The various effects of each PTP on podosome organization in osteoclasts are caused by their distinct N-termini. The function of PTPe in these cells requires the presence of its 12 N-terminal residues, in particular serine 2.

The integration of viral DNA into the host genome is mediated by viral integrase, resulting in the accumulation of double-strand breaks. Integrase-derived peptides (INS and INR) increase the number of integration events, leading to escalated genomic instability that induces apoptosis. CD24 is a surface protein expressed mostly in cancer cells and is very rarely found in normal cells. Here, we propose a novel targeted cancer therapeutic platform based on the lentiviral integrase, stimulated by integrase-derived peptides, that are specifically delivered to cancerous cells via CD24 antigen-antibody targeting. INS and INR were synthesized and humanized and anti-CD24 antibodies were fused to the lentivirus envelope. The activity, permeability, stability, solubility, and toxicity of these components were analyzed. Cell death was measured by fluorescent microscopy and enzymatic assays and potency were tested in vitro and in vivo. Lentivirus particles, containing non-functional DNA led to massive cell death (40–70%). Raltegravir, an antiretroviral drug, inhibited the induction of apoptosis. In vivo, single and repeated administrations of INS/INR were well tolerated without any adverse effects. Tumor development in nude mice was significantly inhibited (by 50%) as compared to the vehicle arm. In summary, a novel and generic therapeutic platform for selective cancer cell eradication with excellent efficacy and safety are presented.

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