Metastatic melanoma is the most aggressive skin cancer. Recently, phenotypically distinct subpopulations of tumor cells were identified. Among them, ABCB5-expressing cells were proposed to display an enhanced tumorigenicity with stem cell-like properties. In addition, ABCB5+ cells are thought to participate to chemoresistance through a potential efflux function of ABCB5. Nevertheless, the fate of these cells upon drugs that are used in melanoma chemotherapy remains to be clarified. Here we explored the effect of anti-melanoma treatments on the ABCB5-expressing cells. Using a melanoma xenograft model (WM266-4), we observed in vivo that ABCB5-expressing cells are enriched after a temozolomide treatment that induces a significant tumor regression. These results were further confirmed in a preliminary study conducted on clinical samples from patients that received dacarbazine. In vitro, we showed that ABCB5-expressing cells selectively survive when exposed to dacarbazine, the reference treatment of metastatic melanoma, but also to vemurafenib, a new inhibitor of the mutated kinase V600E BRAF and other various chemotherapeutic drugs. Our results show that anti-melanoma chemotherapy might participate to the chemoresistance acquisition by selecting tumor cell subpopulations expressing ABCB5. This is of particular importance in understanding the relapses observed after anti-melanoma treatments and reinforces the interest of ABCB5 and ABCB5-expressing cells as potential therapeutic targets in melanoma.
Edited by Velia FowlerMicrotubules are nucleated from multiprotein complexes containing ␥-tubulin and associated ␥-tubulin complex proteins (GCPs). Small complexes (␥TuSCs) comprise two molecules of ␥-tubulin bound to the C-terminal domains of GCP2 and GCP3. ␥TuSCs associate laterally into helical structures, providing a structural template for microtubule nucleation. In most eukaryotes ␥TuSCs associate with additional GCPs (4, 5, and 6) to form the core of the so-called ␥-tubulin ring complex (␥TuRC). GCPs 2-6 constitute a family of homologous proteins. Previous structural analysis and modeling of GCPs suggest that all family members can potentially integrate into the helical structure. Here we provide experimental evidence for this model. Using chimeric proteins in which the N-and C-terminal domains of different GCPs are swapped, we show that the N-terminal domains define the functional identity of GCPs, whereas the C-terminal domains are exchangeable. FLIM-FRET experiments indicate that GCP4 and GCP5 associate laterally within the complex, and their interaction is mediated by their N-terminal domains as previously shown for ␥TuSCs. Our results suggest that all GCPs are incorporated into the helix via lateral interactions between their N-terminal domains, whereas the C-terminal domains mediate longitudinal interactions with ␥-tubulin. Moreover, we show that binding to ␥-tubulin is not essential for integrating into the helical complex.In all eukaryotes, microtubules are nucleated from specialized multiprotein complexes containing ␥-tubulin and associated proteins (1-3). These complexes resemble small rings by electron microscopy and are thus called ␥-tubulin ring complexes (␥TuRCs) 4 (4 -7). Closer inspection revealed that these ␥TuRCs are helices of one turn, with the two ends overlapping. They are ubiquitous and essential for viability: the growth of new microtubules is crucial to drive mitotic spindle formation and cell division. ␥TuRCs are mainly composed of ␥-tubulin and of proteins of the GCP (␥-tubulin complex protein) family. GCPs are characterized by sequence homology in two specific regions, also referred to as the grip1 and grip2 motifs (8). Five members of this family are known: GCPs 2, 3, 4, 5, and 6. GCPs 2 and 3 associate with ␥-tubulin to form a V-shaped subcomplex, called ␥-tubulin small complex (␥TuSC). GCPs 2 and 3 constitute the arms of the V, interacting laterally via their N-terminal domains (Fig. 1, A and B). The C-terminal domains are located at the two tips of the V, each binding one molecule of ␥-tubulin (9). In the budding yeast Saccharomyces cerevisiae, ␥TuSCs are directly recruited to the spindle pole body (yeast centrosome equivalent) by the protein Spc110. Oligomers of Spc110 interact with the basis of the V-shaped ␥TuSCs and stabilize their lateral association (10 -13). Likely, seven ␥TuSCs assemble stepwise into a helix of one turn plus a small overlap (14). In this helical array, the ␥-tubulin molecules are exposed to form a platform from which ␣/-tubulin dimers assemble into...
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