Study of intestinal and liver fatty acid binding proteins (IFABP & LFABP) specific functions

RODRIGUEZ SAWICKI, L; BOTASSO ARIAS, N; FALOMIR LOCKHART, LJ; FRANCHINI, GR; STORCH, J; CÓRSICO, B

La Plata

Congreso; 8th International Conference on Lipid Binding Proteins; 2013

INIBIOLP

Fatty acids (FA) are relevant for the cell as a source of energy and substrates for membrane biogenesis, and participate in many cellular processes such as signals for metabolic regulation and gene expression. Intestinal enterocytes express two homologous proteins that reversibly bound FA: IFABP and LFABP. It has long been hypothesized that FABPs are essential to the intracellular transport and processing of the FA absorbed by the intestine. Nevertheless, their precise functions are not fully understood. The present work evaluates FABP´s specific roles employing 2 approaches. A Caco-2 cell line with LFABP´s ablated expression showed marked variations in FA assimilation and distribution as well as cell proliferation and differentiation. Recently, a Caco-2 cell line with partially diminished expression of LFABP has been obtained. Both cell lines phenotypes will be used to obtain information about LFABP specific functions. A new technique in order to modify intestinal FABPs has been established in our group. Purified IFABP labeled with Alexa555 was incorporated into Cos-7 and Caco-2 cell lines using Direct Protein Transfer (DPT). DPT was performed using JBS-Proteoducin Cocktail Peptide (Jena Bioscience), and protein incorporation was checked by fluorescence microscopy. This methodology will be employed to analyze FA distribution into Caco-2 using BODIPY-FLC16. DPT of IFABP or its structural variants in modified cell lines will allow the study of the effect of these specific proteins in FA distribution in the enterocyte. The interacting partners for intestinal FABPs are still unknown. As a first approach, a screening by Far Western Blot analysis of 2D electrophoresis from Caco-2 homogenates revealed spots that indicated putative 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.