Summary Background Tks5 regulates invadopodium formation, but the precise timing during invadopodium lifetime (initiation, stabilization, maturation) when Tks5 plays a role is not known. Results We report new findings based on high-resolution spatiotemporal live-cell imaging of invadopodium precursor assembly. Cortactin, N-WASP, cofilin, and actin arrive together to form the invadopodium precursor, followed by Tks5 recruitment. Tks5 is not required for precursor initiation but is needed for precursor stabilization, which requires the interaction of the phox homology (PX) domain of Tks5 with PI(3,4)P2. During precursor formation, PI(3,4)P2 is uniformly distributed but subsequently starts accumulating at the precursor core 3–4 min after core initiation, and conversely, PI(3,4,5)P3 gets enriched in a ring around the precursor core. SHIP2, a 5′-inositol phosphatase, localizes at the invadopodium core and regulates PI(3,4)P2 levels locally at the invadopodium. The timing of SHIP2 arrival at the invadopodium precursor coincides with the onset of PI(3,4)P2 accumulation. Consistent with its late arrival, we found that SHIP2 inhibition does not affect precursor formation but does cause decreases in mature invadopodia and matrix degradation, whereas SHIP2 overexpression increases matrix degradation. Conclusions Together, these findings lead us to propose a new sequential model that provides novel insights into molecular mechanisms underlying invadopodium precursor initiation, stabilization, and maturation into a functional invadopodium.

A large imprinted gene cluster in human chromosome 11p15.5 has been implicated in Beckwith-Wiedemann syndrome and Wilms' tumor. We have identified a paternally expressed imprinted gene, PEG8/IGF2AS, in this locus. It is transcribed in the opposite direction to the IGF2 transcripts and some genomic regions are shared with the IGF2 gene, as in the case of the mouse imprinted Igf2as gene reported previously by T. Moore et al. As to the relationship between these genomic regions, the human and mouse genes are very similar but there is no homology in their middle parts. Interestingly, PEG8/IGF2AS and IGF2 were found to be overexpressed in Wilms' tumor samples, at levels over ten and a hundred times higher than that in normal kidney tissues neighboring the tumors, respectively. These findings indicate that PEG8/IGF2AS is a good marker of Wilms' tumor and also suggest the possibility of PEG8/IGF2AS being one of the candidate Wilms' tumor genes.

Previous reports have revealed that estrogen agonists or anti-androgenic chemicals induce abnormal spermiogenesis in rodents. In the seminiferous epithelium, the apical ectoplasmic specialization (ES) is an actin-based (cell-cell) junctional structure developing between the Sertoli cells and spermatids as is the basal ES also--although it is located between adjoining Sertoli cells. In the apical and basal ES are several adhesion complex proteins that control the spermatid developing process. Cortactin, an actin-binding protein, is one of the ES adhesion proteins, combining with several cell-cell adhesions associating proteins. In the present study, 17beta-estradiol (E2, 1.2 microg/kg), bisphenol A (BPA, 2.4 microg/kg), and diethylstilbestrol (DES, 2.5 microg/kg) were subcutaneously injected in ICR 12-week-old male mice. Mice testes were observed for the expression of cortactin protein after E2, BPA, and DES treatments by Western blot analysis, immunohistochemical analysis, and immunoelectron microscopic analysis. Observations showed that the immunoreactivity of the treated testes was significantly decreased. The immunohistochemical reactivity of cortactin in the apical ES was decreased in the treated testis. In immunoelectron microscopic observations, ultrastructural immunolocalizations of cortactin protein in the apical ES by both E2 and BPA were decreased, and the immuno-gold particles of apical and basal ES by DES were much less than the control. In the toxicological field, cortactin may be considered to be one of the indicator proteins of abnormal spermiogenesis which is affected by exogenous chemicals, such as endocrine disrupting chemicals. In summary, this study helps toward understanding the cortactin protein expression underlying the histological abnormalities of spermatogenesis induced by exogenous hormonal chemical treatment.