Center for Molecular Medicine Cologne

Müller, Roman-Ulrich / Fabretti, Francesca - A 4

Characterizing the molecular functions of the tumor suppressor protein Folliculin using the model organism Caenorhabditis elegans

Recent evidence suggests that the Birt-Hogg-Dubé syndrome is a ciliopathy with the tumor suppressor protein mutated in this disease – Folliculin (FLCN) - localizing to primary cilia and influencing their function. Previous work from our group could establish the nematode Caenorhabditis elegans as a model organism for studying the molecular biology underlying this disease. In this project we investigate the putative ciliary role of FLCN exploiting the power of this model for analyzing ciliopathy genes.


Birt-Hogg-Dubé syndrome (BHD) is an autosomal dominant multitumor syndrome that is characterized by the triad of benign skin tumors, recurrent pneumothoraces and renal cyst and tumor formation. Germline mutations in Folliculin (FLCN) are the underlying cause of this syndrome. However, its exact molecular functions have remained elusive.
In a recent study we have been able to introduce the nematode C. elegans as a novel model organism for studying the biology of the BHD syndrome by characterizing a knockout strain of flcn-1, the worm homologue of FLCN. We will now continue this project using both cell culture and particularly the nematode model to examine the signaling pathways FLCN is involved in more closely. Polycystic kidney diseases such as the one occurring in Birt-Hogg-Dubé syndrome are generally considered to be the consequences of dysfunction or dysgenesis of primary cilia – i.e. ciliopathies. Importantly, C. elegans is one of the key model organisms for studies addressing the ciliary function of cystopathy genes. The nematode paired with studies in cultured cells are exploited in this project to gain a better understanding of the molecular function of the tumor suppressor protein FLCN, in particular as to a putative role in ciliary biology.

Determination of the FLCN interactome

Only few interacting proteins of FLCN are known by now. To allow for IP-MS analysis of the FLCN interactome we generated a single-copy transgenic HEK293T cell line expressing an FLCN::GFP fusion protein using TALEN technology. Putative interaction partners that co-precipitated with FLCN were identified using nLC-MS/MS. Doing this we could confirm validity of the approach through the strong enrichment of the known interactor FNIP1. Interestingly, GO term analysis revealed terms associated with vesicle formation and amino acid metabolism to be among the most strongly enriched biological processes (Fig. 1).
This finding nicely fits to recently published data showing FLCN to be involved in the regulation of small vesicular RAG GTPases required for mTOR activation after amino acid starvation. Interestingly, recent cell biological analysis of a splice-site mutation we identified in a BHD-patient revealed that the mutant protein lost the partially nuclear localization of FLCN. Taking into account the enrichment of proteins in the interactome linked to DNA strand elongation and the role of FLCN as a tumor suppressor protein it will be intriguing to study potential consequences of FLCN mutations on this biological process.

C. elegans as a model for a putative ciliary function of FLCN

Recently, two publications have linked FLCN to ciliary functions and shown its ciliary localization in cultured cells. We set out to analyse FLCN mutant worms using established C. elegans phenotypes  So far no overt phenotype in ciliary morphology and function (osmotic avoidance, velocity of IFT, mechanosensation, male mating behaviour, dye filling) could be observed (Fig. 2).
However, commonly the loss of a ciliopathy gene in the worm only effects one specific phenotype. Nonetheless analysis of a strain expressing a FLCN::GFP fusion protein under the control of the FLCN promoter we generated clearly shows expression of the protein in sensory neurons – the only ciliated cell type of the nematode. Consequently, we are currently completing our analysis of ciliary readouts.


The identification of the FLCN interactome will now pave the way to elucidating the molecular function of this protein as well as the consequences of a loss of FLCN for cellular biology. In this context the C. elegans will greatly facilitate to gain a better understanding on whether and how FLCN and its interactors impact on ciliary functions. Elucidation of the functional consequences of actual mutations from BHD patients on binding to the interactors and the cellular functions of FLCN continue to be at the heart of our project.

Selected publications

Bartram, M.P., Amendola, E., Benzing, T., Schermer, B., de Vita, G., Müller, R.-U., (2016). Mice lacking microRNAs in Pax8-expressing cells develop hypothyroidism and end-stage renal failure. BMC Mol. Biol.

Gharbi H, Fabretti F, Bharill P, Rinschen MM, Brinkkötter S, Frommolt P, Burst V, Schermer B, Benzing T, Müller RU (2013). Loss of the Birt-Hogg-Dubé gene product folliculin induces longevity in a hypoxia-inducible factor-dependent manner. Aging Cell

Former Funding Period 01/2014 - 12/2016

Information from this funding period will not be updated anymore. New research related information is available here.

Prof. Dr. Roman-Ulrich Müller CMMC Cologne
Prof. Dr. Roman-Ulrich Müller

Clinic II of Internal Medicine

CMMC - PI - assoc. RG 13

Publications - Roman-Ulrich Müller

Link to PubMed

Dr. rer. nat. Francesca Fabretti CMMC Cologne
Dr. rer. nat. Francesca Fabretti

Dept. II of Internal Medicine

Co-Principal Investigator A 4

+49 221 478 89033

+49 221 478 89041

Dept. II of Internal Medicine

Kerpener Str. 62

50937 Cologne

CMMC Profile Page

Publications - Francesca Fabretti

Link to PubMed

Group Members

Michael Ignarski (Postdoc)
Reza Esmaillie (PhD student)
Tripti Mishra (PhD student)
Rainer Kaiser (MD student)
Serena Greco-Torres (technician)

Figure 1

Figure 2