RESF1 Is a Metastasis Susceptibility Gene for Estrogen Receptor Negative Breast Cancer

Abstract

Breast cancer is the most frequently diagnosed and the second most fatal cancer for women in America. Most breast cancer-related mortality is due to metastatic disease. However, despite the importance of metastasis for breast cancer outcome, little is understood about the process. To investigate the effect of inherited genetic variation on metastasis, we used a mouse genetics strategy comparing strains with high and low metastasis susceptibility to identify 2810474O19Rik, now Resf1, a gene of unknown function, as a new potential metastasis enhancer. Reducing global Resf1 expression using a gene-trap in genetically engineered mouse models of metastatic breast cancer significantly increased pulmonary metastases and metastatic incidence, confirming Resf1 as a metastasis modifier gene. This data also matches with human breast cancer metastasis and RESF1 expression levels in tumor samples. However, paradoxically, orthotopic implantation of Resf1 knockdown cells reduced metastasis in mice, indicating a potential tissue culture in vitro artifact. Resf1 is a poorly characterized protein located in the nucleus. It has been shown to increase protein production when knocked down suggesting a potential role with mRNA processing or translation. Ethinyl uridine (EU) incorporation assays demonstrated an increase in nucleolar RNA upon Resf1 knockdown (KD). RNA-seq and GSEA analysis also revealed a highly significant increase in the ribosomal biosynthesis pathways in the Resf1 KD cells. KD of Resf1 was also accompanied by an increase of protein synthesis, as measured by o-propargyl-puromycin (OPP) labelling of nascent polypeptides, but paradoxically reduced rRNAs in both cell lines and in tumors from the gene-trap model. Taken together, these data suggest that Resf1 may function as a negative regulator of global ribosomal biosynthesis and alteration of global protein synthesis may play an important role in metastatic progression. Currently, we are investigating how Resf1 has a putative role in the nucleolar stress response. The nucleolus, due to the high metabolic demands of ribosome production, is a major component of cellular stress response. Cells frequently reduce ribosome production to divert resources to resolve stresses before resuming normal function. This response is due, in part, to stress-induced long non-coding RNAs (lncRNAs) encoded from the intergenic spacer regions of ribosomes. Intriguingly, preliminary data shows knockdown of Resf1 alters the transcription of at least one of these stress-induced lncRNAs, consistent with a role in the induction of nucleolar stress response. Given this connection, investigations into Resf1’s role in nucleolar stress response and metastasis are directions for future studies.