GNL3 and SKA3 are novel prostate cancer metastasis susceptibility genes

Abstract

Prostate cancer (PC) is very common in developed countries. However, the molecular determinants of PC metastasis are unclear. Previously, we reported that germline variation influences metastasis in the C57BL/6-Tg(TRAMP)8247Ng/J (TRAMP) mouse model of PC. These mice develop prostate tumors similar to a subset of poor outcome, treatment-associated human PC tumors. Here, we used TRAMP mice to nominate candidate genes and validate their role in aggressive human PC in PC datasets and cell lines. Candidate metastasis susceptibility genes were identified through quantitative trait locus (QTL) mapping in 201 (TRAMP × PWK/PhJ) F2 males. Two metastasis-associated QTLs were identified; one on chromosome 12 (LOD = 5.86), and one on chromosome 14 (LOD = 4.41). Correlation analysis using microarray data from (TRAMP × PWK/PhJ) F2 prostate tumors identified 35 metastasis-associated transcripts within the two loci. The role of these genes in susceptibility to aggressive human PC was determined through in silico analysis using multiple datasets. First, analysis of candidate gene expression in two human PC datasets demonstrated that five candidate genes were associated with an increased risk of aggressive disease and lower disease-free survival. Second, four of these genes (GNL3, MAT1A, SKA3, and ZMYM5) harbored SNPs associated with aggressive tumorigenesis in the PLCO/CGEMS GWAS of 1172 PC patients. Finally, over-expression of GNL3 and SKA3 in the PC-3 human PC cell line decreased in vitro cell migration and invasion. This novel approach demonstrates how mouse models can be used to identify metastasis susceptibility genes, and gives new insight into the molecular mechanisms of fatal PC.

Mapping Complex Traits in a Diversity Outbred F1 Mouse Population Identifies Germline Modifiers of Metastasis in Human Prostate Cancer

“…Microarray gene expression analysis was performed to analyze patterns of gene expression in PC-3 and LNCaP cell lines overexpressing candidate genes, as described (Lee et al 2015). Briefly, 200 ng of total RNA was used for labelling in conjunction with the recommended protocol for the Affymetrix Human GeneChip 2.0 ST chips (Santa Clara, CA).…”

“…Hybridization and subsequent processing of the chips were performed as previously described (Williams et al 2014). Quantitative real-time PCR was performed as previously described (Lee et al 2015). Reverse transcription was performed using cDNA Synthesis Kit (Bio-Rad, Hercules, CA).…”

“…Cell proliferation assays were performed as previously described (Lee et al 2014a; Lee et al 2015). Soft agar assays were performed as previously described (Lee et al 2014a; Lee et al 2015), using 2 × 10 3 cells per 24-well in 0.33% bacto-agar and incubated for 14 days.…”

“…Soft agar assays were performed as previously described (Lee et al 2014a; Lee et al 2015), using 2 × 10 3 cells per 24-well in 0.33% bacto-agar and incubated for 14 days. For in vitro migration and invasion assays, PC-3 or LNCaP cells were starved in serum-free media for 12 hours.…”

“…Since tumorigenesis was initiated by the same somatic event (i.e., expression of the SV40-T transgene) in each mouse, and transgene expression was equal in each F1 strain we concluded that germline variation was impacting disease aggressiveness. Subsequent studies have since demonstrated that it is possible to identify modifiers of aggressive prostate cancer through quantitative trait locus (QTL) mapping (Lee et al 2015; Williams et al 2014). However, these earlier QTL studies were limited by the small amount of genetic variation encompassed in mouse mapping populations.…”