PORTFOLIO

Autosomal Dominant Polycystic Kidney Disease (ADPKD) is a chronic disorder marked by progressive growth of numerous fluid-filled cysts in kidneys, and sometimes in liver or other organs. Patients suffer from declining renal function, and by the age of 60, up to 50% need kidney transplants. Other complications include elevated blood pressure, and increased risk of kidney stones or infections. Among rare diseases, PKD is the most common hereditary disease of the kidney, affecting more than 1 in 1000 individuals worldwide. A cure for ADPKD does not exist.

Future therapies will have to halt cystic growth that is induced by mutations in polycystin-1 or -2. Recent progress in PKD research has identified only few so-called «targets» of polycystins. Among these, the signaling molecule cAMP has attracted the most attention, because it accumulates at abnormally high levels in ADPKD kidneys and thereby causes at least some of the alterations seen in cystic cells. Recently, our lab observed that the levels of cAMP also dramatically rise in cystic kidneys of mice with mutations in a different gene called Bicaudal-C. In fruitflies, the corresponding protein Bicaudal-C was predicted to bind specific RNA molecules to control the rate of their translation into proteins, but the relevant target RNAs had not been identified. We found that mammalian Bicaudal-C associates with the RNA that encodes the cAMP-synthesizing enzyme AC6. The presence of Bicaudal-C marks this RNA for silencing by much shorter complementary «microRNA» molecules and by associated repressive factors.

The aim of this project is to determine how the binding of Bicaudal-C to this and other specific RNA molecules is regulated. We expect that insights into the underlying molecular mechanisms will identify new targets for future drugs that restore the balance between the synthesis and the degradation of cAMP.

Research by multiple labs established a causal role for polycystin mutations and for altered cAMP levels in the etiology of PKD. Additional metabolic changes e.g. in the mode of glucose consumption further enhance cyst formation in a manner akin to many cancers. However, effective and well-tolerated therapies to restore normal kidney functions in patients are still unavailable.

One major difficulty is that cAMP likely affects a multitude of cellular processes, and that interfering with any one of them alone will likely have only minimal benefits. On the other hand, a recently developed small molecule inhibitor of cAMP production has adverse side effects because it was designed to block the action of the hormone that controls the body's water balance. Our discovery that Bicaudal-C acts as a biochemical rheostat to directly inhibit cAMP synthesis suggests that improved therapeutic strategies should seek to augment its RNA silencing function.

Apart from this new approach, the project is unique in terms of the proposed strategies how the activity of Bicaudal-C might possibly be stimulated in cystic kidneys. Finally, there is good reason to hope that this study also will provide new non-invasive diagnostic tools to assess the efficacy of future treatments.

In a collaboration with Prof. Olivier Bonny (Univ. Lausanne) and other clinicians, we found that Fetuin-A levels in urine of ADPKD patients are indeed a novel potential marker of disease progression (Piazzon et al., 2015).

One of the first Bicaudal-C mutations found in patients with renal cysts resides in a protein domain called SAM that can associate with itself to form large head-to-tail polymers. Protein structure modeling indicated that such polymers consist of large helices with 6 SAM subunits per turn that display the mRNA-binding domains of Bicaudal-C at the periphery in regular arrays.

No known disease-associated human Bicaudal-C mutations affected SAM-SAM self-interactions. However, self-polymerization was blocked by a spontaneous SAM domain mutation that associated with polycystic kidneys in mice, highlighting the importance of this process in vivo. To distinguish how the SAM domain facilitates translational repression of Bicaudal-C targets, we imaged its effect on the distribution of a reporter mRNA. We found that self-polymerization concentrates Bicaudal-C together with associated mRNA in macroscopic cytoplasmic foci (Rothé et al., 2015).

Polymeric Bicaudal-C foci may act as giant RNA traps to impose a specific folding on bound mRNAs. In search for novel factors that may regulate this process, we found two other SAM domain proteins, ANKS3 and ANKS6, which previously have been shown to associate with situs abnormalities and/or nephronophthisis, another rare disease with polycystic kidneys. Functional of analysis of these interactions suggest that they regulate the assembly or disassembly of homo- versus heterooligomeric Bicaudal-C complexes in giant macromolecular scaffolds but, surprisingly, appeared to be dispensable for Bicaudal-C-mediated mRNA silencing (Rothé et al., submitted).

In molecular screens for novel targets, we found that Bicaudal-C also binds several factors implicated in the regulation of cell metabolism (Leal et al., submitted). Future work will test whether targeting of such interactions or their regulation by Bicaudal-C polymerization or by Ca2+ signaling can slow down or halt cyst formation.

C. Maisonneuve, I. Guilleret, P. Vick, T. Weber, P. Andre, T. Beyer, M. Blum, and D.B. Constam. 2009. Bicaudal C, a novel regulator of Dvl signaling abutting RNA-processing bodies, controls cilia orientation and leftward flow. Development. 136:3019-3030;
M.R. Kraus, S. Clauin, Y. Pfister, M. Di Maio, T. Ulinski, D. Constam, C. Bellanne-Chantelot, and A. Grapin-Botton. 2012. Two mutations in human BICC1 resulting in WNT pathway hyperactivity associate with cystic renal dysplasia. Hum. Mutat. 33:86-90;
N. Piazzon, C. Maisonneuve, I. Guilleret, S. Rotman, and D.B. Constam. 2012. Bicc1 links the regulation of cAMP signaling in polycystic kidneys to microRNA-induced gene silencing. Journal of molecular cell biology. 4:398-408;
Piazzon, N., Bernet, F., Guihard, L., Leonhard, W.N., Urfer, S., Firsov, D., Chehade, H., Vogt, B., Piergiovanni, S., Peters, D.J., Bonny, O., D.B. Constam. 2015. Urine Fetuin-A is a biomarker of autosomal dominant polycystic kidney disease progression. Journal of translational medicine 13, 103;
Rothe, B., Leal-Esteban, L., Bernet, F., Urfer, S., Doerr, N., Weimbs, T., Iwaszkiewicz, J., and D.B. Constam. 2015. Bicc1 polymerization regulates the localization and silencing of bound mRNA. Mol. Cell. Biol. 35, 3339-3353.

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