Enterocyte Fatty Acid Binding Proteins (FABPs): Protein interaction studies in a Caco-2 cell model.

BOTTASSO ARIAS NATALIA; FALOMIR LOCKHART LISANDRO JORGE; CORSICO, BETINA; FRANCHINI GISELA; STORCH JUDITH; RODRIGUEZ SAWICKI LUCIANA; SCAGLIA NATALIA

Mont Blanc

Conferencia; 57th International Conference on the Bioscience of Lipids; 2016

Two isoforms of Fatty Acid Binding Proteins (FABPs) are abundantlyexpressed within intestinal epithelial cells: liver FABP (LFABP) and intestinalFABP (IFABP). Together they represent 1-2% of total proteins present within theenterocytes (Ockner 1972). In this cells LFABP is expressed over 40 times theconcentration of IFABP. They are associated with intracellular dietary lipidtransport and trafficking towards diverse cell fates. But still their functionsare not well understood. By means of Caco-2 intestinal cell model, we startedto address the matter following protein interaction studies.Firstly, screening studies were carried out toevaluate LFABP interaction with other proteins present in Caco-2 cell lysate. UsingFar western blot technique followed by MS we found two candidate proteins:HSP60 and calreticulin. Secondly, Co-immunoprecipitation (CoIP) was applied tostudy IFABP-protein interactions. Based on results reported in the literaturefor other FABPs (Hostetler 2009, Hughes 2015) we tested and confirmed theinteraction between IFABP and a member of the family of the PPARs (PeroxisomeProliferator Activated Receptors) transcription factors, PPARɣ. In this study wefound an apparent decrease in PPARɣ molecular weight in the CoIP eluatecompared to the band present in the Caco-2 lysate. We are now studying thepossible causes for this observation.Thirdly, PPARɣ expression levels were studied in aLFABP ablated Caco-2 cell line, generated in our lab by an antisense strategy(Rodriguez Sawicki, under preparation). Related to the diminished LFABPexpression we found a concomitant decrease in PPARɣ levels by western blot analysisin heterogeneous and clonal cell lines compared to controls. Currently, our efforts are focused on recombinant PPARɣpurification to analyze the interaction with IFABP in vitro and studyingchanges in the expression of PPARɣ downstream genes in cultivo, usinginhibitors and activators of FABPs and PPARɣ, alternatively.In summary, our work shows for the first time theinteraction of FABPs with other proteins expressed in Caco-2 cells, enforcingthe idea of differential roles for I and LFABP in this cell model.