Protein-Protein Interactions

The first evidence of protein-protein interaction by folliculin (FLCN) was provided by Baba et al (2006) when they identified FLCN-interacting protein 1 (or FNIP). Co-immunoprecipitation studies were used to identify FNIP1 in HEK293 cells, with an initial interrogation of the protein on blast revealing no known functional domains but showing some degree of homology between several species. This normally indicates that the protein carries out an important function. Further antibody studies and in vitro binding assays subsequently showed that interaction between FLCN and FNIP occurs through the C-terminal end of FLCN and requires full length FNIP protein for optimum binding. In order to elucidate the function of FNIP, similar co-immunoprecipitation assays were carried out on FNIP itself to directly identify other binding partners. Crucially, AMPK was identified as a binding partner and shown to bind independently of folliculin. Subsequent assays showed that FNIP can phosphorylate FLCN and that this activity is mTOR and AMPK activity dependent. Also, expression studies of showed that FNIP1 and FLCN phosphorylation was suppressed by AMPK inhibition, indicating a functional interaction between FLCN/FNIP1 and mTOR/AMPK signalling pathways.

We already know that mutations in genes leading to the disruption of mTOR signalling give rise to many of the hamartomatous syndromes we have previously examined.  AMPK has a functional role within this pathway. Cellular energy deficits triggers AMPK phosphorylation by LKB1 and calmodulin kinase kinase, which negatively regulates mTOR through TSC2 phosphorylation, inhibiting protein translation and cell growth. Crucially, this study suggested that both FNIP and FLCN may be downstream effectors of mTOR and AMPK and that they are able to effect these pathways by an unknown role. This study was significant in clarifying that FLCN functions as a tumour suppressor and further implicated the mTOR as the causative signalling pathway disrupted to give rise to BHD syndrome.

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