Investigation of novel accessory proteins of neuronal endosomal CLC-3 and CLC-4 antiporters involved in neurodevelopmental disorders

More than a thousand different genes are involved in various forms of intellectual disability, neurodevelopmental delay, neurodegeneration and other neurological diseases affecting pediatric patients. These include the members of the CLC gene family, CLCN3 and CLCN4, which encode anion transporting proteins that are important for the functioning of the intracellular structures of neurons, such as endosomes and lysosomes.  CLCN3 and CLCN4 variants are identified in an increasing number of patients.

Rarely do proteins work alone but rather are part of larger complexes, in association with other proteins. No such accessory proteins have been known so far for CLCN3 and CLCN4. In preliminary experiments we discovered that two proteins of unknown function strongly interact with CLCN3 as well as CLCN4. These proteins are called TMEM9 and TMEM9B. We found that co-expression of these proteins reduces the function of CLCN3 and CLCN4 and also alters the properties of CLCN3. With advanced microscopy we found that TMEM9B physically interacts with CLCN3 and CLCN4. This discovery is potentially very important for understanding the mechanisms underlying CLCN3/-4 related conditions.In this project we aim to characterize the interaction of CLCN3/4 with TMEM9/9B in detail and verify its pathological relevance. In particular we will test whether the interaction occurs in neurons and whether CLCN3/4 variants alter the interaction with TMEM9(B) and thereby cause disease.

The identification of these new molecular players can expand the frontiers of research on the pathologies caused by CLCN3 and CLCN4 variants and provide new strategies for the care and therapy of patients.