Intracellular localization of GASP/ECOP/VOPP1

Abstract

Vesicular Over-expressed in cancer Prosurvival Protein 1 (VOPP1), also known as Glioblastoma Amplified and Secreted Protein and EGFR-Coamplified and Over-expressed Protein has been previously shown to be over-expressed in human glioblastoma multiforme and squamous cell carcinoma. Additionally, previous experimental work suggests that it confers a prosurvival cellular phenotype. A query of a public database of gene expression profiling data (Oncomine) shows that the VOPP1 transcript is also highly expressed in several other common human cancers, including breast carcinoma, pancreatic carcinoma, and lymphoma. Analysis of VOPP1 sequence structure shows both a signal sequence and a transmembrane domain, and examination of a public microarray dataset for endoplasmic reticulum (ER)-bound mRNA transcripts is consistent with the VOPP1 protein product being synthesized into the ER. Immunoblot analysis of cell culture and conditioned media confirms that the protein product is not secreted and is retained intracellularly. VOPP1 protein tagged with a fluorescence reporter, as well as antibody-mediated visualization of recombinant and native forms of the protein reveals an intracellular vesicular pattern of localization. Co-localization experiments reveal that VOPP1 vesicles do not co-localize with mitochondria or peroxisomes, but show partial co-localization with perinuclear lysosomes. Additionally, markers of endocytosis and autophagy show partial perinuclear co-localization, suggesting that VOPP1-containing vesicles enter final common pathways of the lysosomal system. These findings throw into doubt the hypothesis that VOPP1 interacts directly with cytoplasmic mediators of the NF kappa B pathway, and suggest that the prosurvival phenotype conferred by this gene product is mediated by other mechanisms.