CDKL5 and neuronal morphological defects: novel perspectives of microtubule related drugs

Mutations in the CDKL5 gene have been found in individuals with a rare neurodevelopmental disorder. CDKL5 is a serine/threonine kinase whose functions are still not fully understood. An aberrant neuronal morphology linked to Cdkl5 has been observed both in Cdkl5--‐silenced primary neurons as well as in Cdkl5--‐null brains strongly suggesting that these defects may underlie the cognitive impairment characterizing both patients and mice devoid of the kinase. The molecular basis of these defects is still far from understood but altered cytoskeletal dynamics are likely to be involved. The identification of IQGAP1 as novel CDKL5 interactor may provide a key to understand such neuronal defects. IQGAP1 promotes microtubule dynamics through its association with Rac1 and CLIP--‐170, whose activity on MTs might play a significant role in the regulation of cytoskeletal dynamics in cycling cells and neurons. Considering how the loss of CDKL5 negatively impacts on cellular and neuronal morphology, we envisaged that this kinase could regulate cytoskeletal dynamics acting directly or indirectly on CLIP--‐170. Indeed, CLIP--‐170 is known to regulate several aspects of neuronal morphology such as axon outgrowth, dendritic arborization and growth cone organization. By bridging the actin cytoskeleton to MTs, CLIP--‐170 coordinates proper cytoskeletal dynamics. In the current study, we analysed the role of CDKL5 in the regulation of CLIP--‐170 activity. Through a FRET analysis on COS7 cells we demonstrated that the loss of CDKL5 causes CLIP--‐170 to be mainly in its closed inactive conformation, thus reducing its interaction with MTs. Furthermore, using Cdkl5--‐ KO primary hippocampal neurons we demonstrated that the loss of CDKL5 is detrimental for the correct progression of several steps of neuronal maturation. Moreover, we demonstrated that such defects can be restored upon treatment with pregnenolone (a neurosteroid which stabilizes the extended conformation of CLIP--‐170, increasing its affinity for MTs and promoting their polymerization) and Pregnenolone--‐Methyl--‐Ether, a synthetic non--‐metabolizable P5 derivative. We speculate that the positive effects of the two compounds are likely to be due to an activation of CLIP--‐ 170 functionality and a concomitant increase in MT dynamics.