Analysis of the specific roles of the enterocyte fatty acid binding proteins (FABP): intestinal and liver- FABP.

LUCIANA RODRIGUEZ SAWICKI; NATALIA M. BOTTASSO ARIAS; NATALIA SCAGLIA; LISANDRO JORGE FALOMIR LOCKHART; GISELA RAQUEL FRANCHINI; JUDITH STORCH; BETINA CÓRSICO

Workshop; Cellular Imaging of Lipids; 2014

EMBO

Lipid hydrolysis in the intestinal lumen releases great quantities of long chain fatty acids (LCFA). Following uptake, they are reversibly bound to two homologous proteins that are abundantly expressed in the enterocytes: I- (1) and L-FABP (2). It is currently not known why a single cell type. However, a number of interesting differences related to tissue distribution and binding properties have been found between L- and IFABP, suggesting that they have different functions within the enterocyte, contributing to differential trafficking and compartmentation of lipids (3, 4). To study intestinal FABPs functions, a Caco-2 cell line with LFABP´s ablated expression has been characterized. No compensation by IFABP was observed. Knockdown cells showed a marked decrease in oleate assimilation, while palmitate assimilation was increased. Differences in oleate distribution were observed at short times. Cell proliferation and differentiation were slowed in the knockdown cells. These results indicate that LFABP is an important factor for the proper function of the enterocytes. A new technique to modify FABPs levels has been established in our group. Purified IFABP labeled with Alexa555 was incorporated into Caco-2 cell lines using Direct Protein Transfer (DPT). This methodology will be employed to analyse FA distribution into Caco-2 using BODIPY-FLC16. DPT of FABPs or its structural variants in modified cell lines will allow us to obtain information of the molecular transit of FA in the enterocyte. The interacting protein partners for intestinal FABPs are still unknown. A screening assay employing Far Western Blot analysis of 2D gels from Caco-2, revealed spots that indicate LFABP´s interaction with other proteins. The spots are being analyzed using mass spectrometry. The information obtained combining protein-protein interaction assays with cell culture studies will contribute to determine intestinal FABPs specific functions in the enterocyte. 1- Ockner et al, J. Clin. Invest. 1974. 2- Ockner et al, Science 1972. 3-Pelsers et al, Clin Biochem. 2003. 4-Richieri et al, J. Biol. Chem. 1994.