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Tumor protein D52 (TPD52) is amplified and/or overexpressed in cancers of diverse cellular origins. Altered cellular metabolism (including lipogenesis) is a hallmark of cancer development, and protein–protein associations between TPD52 and known regulators of lipid storage, and differential TPD52 expression in obese versus non-obese adipose tissue, suggest that TPD52 might regulate cellular lipid metabolism. We found increased lipid droplet numbers in BALB/c 3T3 cell lines stably expressing TPD52, compared with control and TPD52L1-expressing cell lines. TPD52-expressing 3T3 cells showed increased fatty acid storage in triglyceride (from both de novo synthesis and uptake) and formed greater numbers of lipid droplets upon oleic acid supplementation than control cells. TPD52 colocalised with Golgi, but not endoplasmic reticulum (ER), markers and also showed partial colocalisation with lipid droplets coated with ADRP (also known as PLIN2), with a proportion of TPD52 being detected in the lipid droplet fraction. Direct interactions between ADRP and TPD52, but not TPD52L1, were demonstrated using the yeast two-hybrid system, with ADRP–TPD52 interactions confirmed using GST pulldown assays. Our findings uncover a new isoform-specific role for TPD52 in promoting intracellular lipid storage, which might be relevant to TPD52 overexpression in cancer.

Background: Tumor protein D52 (D52) overexpression is characteristic of both adult and childhood cancers and has been associated with reduced overall survival in breast cancer patients and an increase in proliferation and metastatic capacity of transfected cell lines. However, little is known about the mechanism(s) by which D52 overexpression contributes to cellular transformation or tumour progression. Aims: We aimed to identify genes which are differentially expressed in response to D52 expression in 3T3 fibroblasts, and to use these gene expression changes to predict cellular phenotypes which are altered by D52 expression. Methods: Gene expression profiling was performed using Affymetrix expression microarray analysis. Up- and down-regulated gene lists were analysed using GOSTAT, Metacore and literature searches. Differential expression of candidate proteins were validated using Western blot analysis of intracellular protein extracts or precipitates from conditioned media. Migration and invasion assays were performed using Boyden chambers. Results: D52 expression was associated with over 300 genes being differentially expressed at least two-fold relative to vector controls. A number of overexpressed genes were found to be normally involved in the inflammatory response. We focused on validating candidates such as COX-2, MMP-2 and Osteopontin as being overexpressed in D52-transfected cells relative to vector controls. We also confirmed an upregulation of phospho-AKT in D52 expressing cells, as reported by others. GOSTAT analysis revealed that gene ontology terms related to the extracellular matrix (ECM) were most over-represented amongst differentially expressed genes. The implied resultant ECM re-modelling was examined through migration and invasion assays, in which D52 expressing cells showed a significantly increased migratory and invasive capacity relative to vector control lines. Conclusions: These results implicate D52 as a pro-inflammatory intermediate, whose expression may act to drive cancer progression via an increase in phospho-AKT, followed by the production of inflammatory molecules such as Osteopontin. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2338.