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.
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. Interestingly, GO term analysis revealed terms associated with vesicle formation and amino acid metabolism to be among the most enriched biological processes (Fig. 1).
In parallel to our work in cell culture and C. elegans we focus on characterizing novel mutations found in affected families. In this context, human genetics analysis of a family affected by BHD syndrome revealed not mutation in the coding region. However, we found a sequence alteration in a splice site and could confirm its impact on splicing of the folliculin transcript (Fig. 2). We then went on to analyse the impact of this splicing defect on protein biology and could show that the mutant protein is strongly destabilized and shows a localization defect (Bartram et al. 2017).
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. 3). 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, 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.
M.P. Bartram, T. Mishra, N. Reintjes, F. Fabretti, H. Gharbi, A.C. Adam, H. Göbel, M. Franke, B. Schermer, S. Haneder, T. Benzing, B.B. Beck and R.-U. Müller (2017), Characterization of a splice-site mutation in the tumor suppressor gene FLCN associated with renal cancer, BMC Med. Genet.
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
Braun, F., Suarez, V., Dinter, J., Haneder, S., Quaas, A., Benzing, T., Nierhoff, D., and Muller, R.U. (2019). Successful use of TNFalpha blockade in a severe case of idiopathic non-granulomatous ulcerative jejunoileitis associated with thrombotic thrombocytopenic purpura. BMJ Open Gastroenterol 6, e000252.
Ignarski, M., Islam, R., and Muller, R.U. (2019). Long Non-Coding RNAs in Kidney Disease. Int J Mol Sci 20.
Ignarski, M., Rill, C., Kaiser, R.W.J., Kaldirim, M., Neuhaus, R., Esmaillie, R., Li, X., Klein, C., Bohl, K., Petersen, M., Frese, C.K., Hohne, M., Atanassov, I., Rinschen, M.M., Hopker, K., Schermer, B., Benzing, T., Dieterich, C., Fabretti, F., and Muller, R.U. (2019). The RNA-Protein Interactome of Differentiated Kidney Tubular Epithelial Cells. J Am Soc Nephrol 30, 564-76.
Jain, M., Kaiser, R.W.J., Bohl, K., Hoehne, M., Gobel, H., Bartram, M.P., Habbig, S., Muller, R.U., Fogo, A.B., Benzing, T., Schermer, B., Hopker, K., and Slaats, G.G. (2019). Inactivation of Apoptosis Antagonizing Transcription Factor in tubular epithelial cells induces accumulation of DNA damage and nephronophthisis. Kidney Int 95, 846-58.
Kaiser, R.W.J., Ignarski, M., Van Nostrand, E.L., Frese, C.K., Jain, M., Cukoski, S., Heinen, H., Schaechter, M., Seufert, L., Bunte, K., Frommolt, P., Keller, P., Helm, M., Bohl, K., Hohne, M., Schermer, B., Benzing, T., Hopker, K., Dieterich, C., Yeo, G.W., Muller, R.U., and Fabretti, F. (2019). A protein-RNA interaction atlas of the ribosome biogenesis factor AATF. Sci Rep 9, 11071.
Muller, R.U., and Schermer, B. (2019). Hippo signaling-a central player in cystic kidney disease? Pediatr Nephrol10.1007/s00467-019-04299-3.
Rinschen, M.M., and Muller, R.U. (2019). A knowledge-guided kidney cell census-reconciling bulk omics with cellular heterogeneity? Kidney Int 95, 733-5.
Spath, M.R., Bartram, M.P., Palacio-Escat, N., Hoyer, K.J.R., Debes, C., Demir, F., Schroeter, C.B., Mandel, A.M., Grundmann, F., Ciarimboli, G., Beyer, A., Kizhakkedathu, J.N., Brodesser, S., Gobel, H., Becker, J.U., Benzing, T., Schermer, B., Hohne, M., Burst, V., Saez-Rodriguez, J., Huesgen, P.F., Muller, R.U., and Rinschen, M.M. (2019). The proteome microenvironment determines the protective effect of preconditioning in cisplatin-induced acute kidney injury. Kidney Int 95, 333-49.
Volker, L.A., Burkert, K., Scholten, N., Grundmann, F., Kurschat, C., Benzing, T., Hampl, J., Becker, J.U., and Muller, R.U. (2019). A case report of recurrent membranoproliferative glomerulonephritis after kidney transplantation due to ventriculoatrial shunt infection. BMC Nephrol 20, 296.
Dafinger C, Rinschen MM, Borgal L, Ehrenberg C, Basten SG, Franke M, Hohne M, Rauh M, Gobel H, Bloch W, Wunderlich FT, Peters DJM, Tasche D, Mishra T, Habbig S, Dotsch J, Muller RU, Bruning JC, Persigehl T, Giles RH, Benzing T, Schermer B, and Liebau MC (2018). Targeted deletion of the AAA-ATPase Ruvbl1 in mice disrupts ciliary integrity and causes renal disease and hydrocephalus. Exp Mol Med 50, 75.
Dal Magro C, Keller P, Kotter A, Werner S, Duarte V, Marchand V, Ignarski M, Freiwald A, Muller RU, Dieterich C, Motorin Y, Butter F, Atta M, and Helm M (2018). A Vastly Increased Chemical Variety of RNA Modifications Containing a Thioacetal Structure. Angew Chem Int Ed Engl 57, 7893-7897.
Grundmann F, Muller RU, Reppenhorst A, Hulswitt L, Spath MR, Kubacki T, Scherner M, Faust M, Becker I, Wahlers T, Schermer B, Benzing T, and Burst V (2018). Preoperative Short-Term Calorie Restriction for Prevention of Acute Kidney Injury After Cardiac Surgery: A Randomized, Controlled, Open-Label, Pilot Trial. J Am Heart Assoc 7.
Karaiskos N, Rahmatollahi M, Boltengagen A, Liu H, Hoehne M, Rinschen M, Schermer B, Benzing T, Rajewsky N, Kocks C, Kann M, and Muller RU (2018). A Single-Cell Transcriptome Atlas of the Mouse Glomerulus. J Am Soc Nephrol10.1681/ASN.2018030238.
Muller RU, Haas CS, and Sayer JA (2018). Practical approaches to the management of autosomal dominant polycystic kidney disease patients in the era of tolvaptan. Clin Kidney J 11, 62-69.
Spath MR, Bartram MP, Palacio-Escat N, Hoyer KJR, Debes C, Demir F, Schroeter CB, Mandel AM, Grundmann F, Ciarimboli G, Beyer A, Kizhakkedathu JN, Brodesser S, Gobel H, Becker JU, Benzing T, Schermer B, Hohne M, Burst V, Saez-Rodriguez J, Huesgen PF, Muller RU, and Rinschen MM (2018). The proteome microenvironment determines the protective effect of preconditioning in cisplatin-induced acute kidney injury. Kidney Int10.1016/j.kint.2018.08.037.
Muller RU, and Benzing T (2018). Cystic Kidney Diseases From the Adult Nephrologist's Point of View. Front Pediatr 6, 65.
Muller RU, and Benzing T (2018). Management of autosomal-dominant polycystic kidney disease-state-of-the-art. Clin Kidney J 11, i2-i13.
Bartram MP, Mishra T, Reintjes N, Fabretti F, Gharbi H, Adam AC, Gobel H, Franke M, Schermer B, Haneder S, Benzing T, Beck BB, and Muller RU (2017). Characterization of a splice-site mutation in the tumor suppressor gene FLCN associated with renal cancer. BMC Med Genet 18, 53.
Tiku V, Jain C, Raz Y, Nakamura S, Heestand B, Liu W, Spath M, Suchiman HED, Muller RU, Slagboom PE, Partridge L, and Antebi A (2017). Small nucleoli are a cellular hallmark of longevity. Nat Commun 8, 16083.
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Hypoxia-signaling at the crossroads of cellular stress resistance and tumorigenesis
Clinic II of Internal Medicine
CMMC - PI - assoc. RG 13
roman-ulrich.mueller[at]uk-koeln.de
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+49 221 478 89041
Clinic II of Internal Medicine
Kerpener Str. 62
50937 Cologne
Michael Ignarski (Postdoc)
Reza Esmaillie (PhD student)
Tripti Mishra (PhD student)
Lisa Seufert (PhD student)
Katrin Bohl (bioinformatician)
Serena Greco-Torres (technician)
Copyright ©
Prof. Dr. med. Thomas Benzing
Chair of the CMMC
