Cancer chromosomal instability (CIN) results from dynamic changes to chromosome number and structure. The resulting diversity in somatic copy number alterations (SCNA) may provide the variation necessary for cancer evolution. Multi-sample phasing and SCNA analysis of 1421 samples from 394 tumours across 24 cancer types revealed ongoing CIN resulting in pervasive SCNA heterogeneity. Parallel evolutionary events, causing disruption to the same genes, such as BCL9, ARNT/HIF1B, TERT and MYC, within separate subclones were present in 35% of tumours. Most recurrent losses occurred prior to whole genome doubling (WGD), a clonal event in 48% of tumours. However, loss of heterozygosity at the human leukocyte antigen locus and loss of 8p to a single haploid copy recurred at significant subclonal frequencies, even in WGD tumours, likely reflecting ongoing karyotype remodeling. Focal amplifications affecting 1q21 (BCL9, ARNT), 5p15.33 (TERT), 11q13.3 (CCND1), 19q12 (CCNE1) and 8q24.1 (MYC) were frequently subclonal and exhibited an illusion of clonality within single samples. Analysis of an independent series of 1024 metastatic samples revealed enrichment for 14 focal SCNAs in metastatic samples, including late gains of 8q24.1 (MYC) in clear cell renal carcinoma and 11q13.3 (CCND1) in HER2-positive breast cancer. CIN may enable ongoing selection of SCNAs, manifested as ordered events, often occurring in parallel, throughout tumour evolution.
Microtubule-based vesicular transport is well documented in epithelial cells, but the specific motors involved and their regulation during polarization are largely unknown. We demonstrate that KIF5B mediates post-Golgi transport of an apical protein in epithelial cells, but only after polarity has developed. Time-lapse imaging of EB1-GFP in polarized MDCK cells showed microtubule plus ends growing toward the apical membrane, implying that plus end-directed N-kinesins might be used to transport apical proteins. Indeed, time-lapse microscopy revealed that expression of a KIF5B dominant negative or microinjection of function-blocking KIF5 antibodies inhibited selectively post-Golgi transport of the apical marker, p75-GFP, after polarization of MDCK cells. Expression of other KIF dominant negatives did not alter p75-GFP trafficking. Immunoprecipitation experiments demonstrated an interaction between KIF5B and p75-GFP in polarized, but not in subconfluent, MDCK cells. Our results demonstrate that apical protein transport depends on selective microtubule motors and that epithelial cells switch kinesins for post-Golgi transport during acquisition of polarity.
Metastases account for 90% of cancer-related deaths; thus, it is vital to understand the biology of tumour dissemination. Here, we collected and monitored >50 patient specimens ex vivo to investigate the cell biology of colorectal cancer (CRC) metastatic spread to the peritoneum. This reveals an unpredicted mode of dissemination. Large clusters of cancer epithelial cells displaying a robust outward apical pole, which we termed tumour spheres with inverted polarity (TSIPs), were observed throughout the process of dissemination. TSIPs form and propagate through the collective apical budding of hypermethylated CRCs downstream of canonical and non-canonical transforming growth factor-β signalling. TSIPs maintain their apical-out topology and use actomyosin contractility to collectively invade three-dimensional extracellular matrices. TSIPs invade paired patient peritoneum explants, initiate metastases in mice xenograft models and correlate with adverse patient prognosis. Thus, despite their epithelial architecture and inverted topology TSIPs seem to drive the metastatic spread of hypermethylated CRCs.
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