Overexpression of a novel human lysophosphatidylcholine acyltransferase in colorectal cancer and its role in lipid metabolism Free

369

Colorectal cancer is the third most frequent form of cancer worldwide. Finding new molecules that provide earlier and more accurate diagnosis, together with a better understanding of the underlying molecular biology of the disease are the aims of our research efforts.

Previous microarray analyses identified the transcript of an expressed sequence tag (EST) to be differentially expressed in pools of distal sporadic colon cancers when compared to a pool of normal mucosas. This EST was identified as a gene encoding for a hypothetical acyltransferase (HAT).

Microarray profiling on 168 colorectal samples using U133plus2.0 arrays showed that the gene transcript was significantly upregulated in 158 out of 168 adenocarcinomas, when compared to normal mucosa (p <10^-12). HAT expression level was significantly (p <10^-7) higher in microsatellite instable (MSI) than in microsatellite stable (MSS) tumors. In contrast, there was no significant difference due to gender, age, Dukes stage, grade, outcome or metastasis.

The coding sequence of the acyltransferase gene was PCR amplified from a colon adenocarcinoma cDNA library and cloned into mammalian expression vectors.

Using a monospecific polyclonal peptide raised antibody, against HAT, immunofluorescence microscopy of transiently transfected COS7 cells, showed that the HAT protein was located at the ER and mitochondria and may be modified in the Golgi apparatus.

Proliferation studies performed on transfected COS7 and SW480 cells showed a significant faster growth rate for the colon cancer line SW480.

Immunohistochemical analysis of formalin fixed, paraffin embedded colon adenocarcinoma specimens, showed very strong protein expression in malignant cells but only weakly positive staining in matching normal mucosas.

Functional studies showed that the coding protein is a lysophosphatidylcholine acyltransferase with preference for palmitoyl CoA increasing phosphatidylcholine (PtdCho) remodeling/recycling.

Interestingly, PtdCho content in colon cancer is significantly increased. Alterations of membrane lipid levels influence cell proliferation and viability. In addition, a change in the superficial membrane potential is associated with malignancy and higher amounts of membrane phospholipids are required for growth of the tumour.

We conclude that HAT is overexpressed in colorectal cancer and this upregulation may contribute to the increase in PtdCho via a remodeling pathway, and thus be one of the mechanisms that crucially change the phospholipid membrane composition in colorectal cancer cells.