‘Gain’ of supernumerary copies of the 8q24.21 chromosomal region has been shown to be common in many human cancers1–13 and is associated with poor prognosis7,10,14. The well-characterized myelocytomatosis (MYC) oncogene resides in the 8q24.21 region and is consistently co-gained with an adjacent ‘gene desert’ of approximately 2 megabases that contains the long non-coding RNA gene PVT1, the CCDC26 gene candidate and the GSDMC gene. Whether low copy-number gain of one or more of these genes drives neoplasia is not known. Here we use chromosome engineering in mice to show that a single extra copy of either the Myc gene or the region encompassing Pvt1, Ccdc26 and Gsdmc fails to advance cancer measurably, whereas a single supernumerary segment encompassing all four genes successfully promotes cancer. Gain of PVT1 long non-coding RNA expression was required for high MYC protein levels in 8q24-amplified human cancer cells. PVT1 RNA and MYC protein expression correlated in primary human tumours, and copy number of PVT1 was co-increased in more than 98% of MYC-copy-increase cancers. Ablation of PVT1 from MYC-driven colon cancer line HCT116 diminished its tumorigenic potency. As MYC protein has been refractory to small-molecule inhibition, the dependence of high MYC protein levels on PVT1 long non-coding RNA provides a much needed therapeutic target.

Limb skeletal elements originate from the limb progenitor cells, which undergo expansion and patterning to develop each skeletal element. Posterior-distal skeletal elements, such as the ulna/fibula and posterior digits develop in a Sonic hedgehog (Shh)-dependent manner. However, it is poorly understood how anterior-proximal elements, such as the humerus/femur, the radius/tibia and the anterior digits, are developed. Here we show that the zinc finger factors Sall4 and Gli3 cooperate for proper development of the anterior-proximal skeletal elements and also function upstream of Shh-dependent posterior skeletal element development. Conditional inactivation of Sall4 in the mesoderm before limb outgrowth caused severe defects in the anterior-proximal skeletal elements in the hindlimb. We found that Gli3 expression is reduced in Sall4 mutant hindlimbs, but not in forelimbs. This reduction caused posteriorization of nascent hindlimb buds, which is correlated with a loss of anterior digits. In proximal development, Sall4 integrates Gli3 and the Plzf-Hox system, in addition to proliferative expansion of cells in the mesenchymal core of nascent hindlimb buds. Whereas forelimbs developed normally in Sall4 mutants, further genetic analysis identified that the Sall4-Gli3 system is a common regulator of the early limb progenitor cells in both forelimbs and hindlimbs. The Sall4-Gli3 system also functions upstream of the Shh-expressing ZPA and the Fgf8-expressing AER in fore-and hindlimbs. Therefore, our study identified a critical role of the Sall4-Gli3 system at the early steps of limb development for proper development of the appendicular skeletal elements.Sall4 | Gli3 | limb progenitors | appendicular skeletal elements | Plzf-Hox H ow progenitor cells are spatially and temporarily organized to construct an organ is a central question in developmental biology. Limb skeletal elements develop from limb progenitors, which arise from the lateral plate mesoderm (LPM) that is originated from epithelial somatopleure (1). Limb progenitors initially form two paired protrusions, fore-and hindlimb buds, whose initiation occurs around embryonic day (E) 9.0 and E9.5, respectively, in mouse embryos. In the following steps, limb signaling centers, known as the zone of polarizing activity (ZPA) and apical ectodermal ridge (AER), are established. SHH (Sonic hedgehog) from the ZPA and FGF8 from the AER are major signal molecules that regulate proliferative expansion and patterning of early limb progenitor cells (reviewed in ref. 2). These processes lead to development of functional limbs with each skeletal element adopting a unique shape at a distinct location.Several studies suggest that limb progenitors consist of two distinct pools, an anterior progenitor pool and a posterior progenitor pool. The posterior progenitor pool consists of cells that once expressed Shh and cells that received paracrine effects of SHH, which contribute to digit 2 (d2), d3, d4, and d5 and the posterior zeugopod (ulna, fibula) (3-6). Contrary to this, the anterior ...

Vertebrate segments called somites are generated by periodic segmentation of the anterior extremity of the presomitic mesoderm (PSM). During somite segmentation in zebrafish, mesp-b determines a future somite boundary at position B-2 within the PSM. Heat-shock experiments, however, suggest that an earlier future somite boundary exists at B-5, but the molecular signature of this boundary remains unidentified. Here, we characterized fibroblast growth factor (FGF) signal activity within the PSM, and demonstrated that an anterior limit of downstream Erk activity corresponds to the future B-5 somite boundary. Moreover, the segmentation clock is required for a stepwise posterior shift of the Erk activity boundary during each segmentation. Our results provide the first molecular evidence of the future somite boundary at B-5, and we propose that clock-dependent cyclic inhibition of the FGF/Erk signal is a key mechanism in the generation of perfect repetitive structures in zebrafish development. KEY WORDS: FGF signalling, Clock, Segmentation, Somitogenesis INTRODUCTIONProper formation of a well-proportioned body in multicellular organisms requires spatiotemporal control of multiple biological processes during development. For instance, the regularity of repetitive structures in vertebrate bodies is derived from the vertebrate segments called somites, which are generated by periodic segmentation of the uniform presomitic mesoderm (PSM) (Pourquié, 2001). The periodicity of segmentation is regulated by oscillation of the segmentation clock genes, and the position of segmentation is determined by a gradient of fibroblast growth factor (FGF) (Dubrulle et al., 2001;Dubrulle and Pourquié, 2004;Holley, 2007;Pourquié, 2001;Sawada et al., 2001).The first detectable sign of a future segment boundary, as visualized by the expression of mesoderm posterior homolog-b (mesp-b; mespba -Zebrafish Information Network), appears at a distance of two somite lengths posterior to B0 (B-2, between S-I and S-II; see Fig. 1D for nomenclature) in the anterior PSM in zebrafish (Sawada et al., 2000), although this may not be the earliest event in boundary determination. A brief exposure of zebrafish embryos to heat shock disrupts somite segmentation after four cycles of normal segmentation (Roy et al., 1999), suggesting that the boundaries of at least five somites (from B-1 to B-5, between S-IV and S-V) are predetermined somewhere in the uniform PSM. However, no RESEARCH ARTICLEGene Regulation Research, Nara Institute Science and Technology, 8916-5 Takayama, Nara 630-0101, Japan. RESULTS The fgf8a gradient moves continuously towards the posterior during somitogenesisIn chick, an Fgf8 gradient, generated by restricted transcription and mRNA decay of Fgf8, is essential for setting up the position of the segmentation (Dubrulle and Pourquié, 2004). In zebrafish, segmentation defects can be seen in ace/ace embryos (fgf8a mutants) (supplementary material Fig. S1) (see also Reifers et al., 1998), suggesting that zebrafish fgf8a has a similar role a...

Highlights d Calcium wave propagates from extruding cells within the epithelium d Calcium wave promotes apical extrusion of transformed or apoptotic cells d IP 3 R, gap junction, and TRPC1 are involved in calcium wave d Calcium wave induces polarized movement of surrounding cells toward extruding cells