PolyA RNA-seq of Paramecium tetraurelia after MKS2/TMEM216 depletion

Ciliogenesis is a widely used process in eukaryotic cells whose different steps start to be well understood. However, the molecular mechanisms leading to decilation or ciliary shedding are yet poorly known. This process observed from unicellular organisms such as Chlamydomonas or Paramecium to multiciliated cells from trachea or fallopian tube of vertebrates, seems to be a general process since recent observations on living cells recently demonstrate its requirement during the cell cycle or neurogenesis. Interestingly, in all cellular models, ciliary shedding occurs distally of the transition zone, essentially known to act as a diffusion barrier between the intracellular space and the cilium, suggesting conserved molecular mechanism. To determine if MKS and NPHP modules, known to cooperate to establish the transition zone formation and function, could control the ciliary shedding process, we studied the function of five proteins of these complexes in Paramecium: TMEM216/MKS2 and TMEM107 (two members of the MKS module), NPHP4 (the only member of the NPHP module in this organism), CEP290/NPHP6 and RPGRIP1L/MKS5. We show that all these proteins are recruited to the TZ as soon as cilia start to grow and localise in 9-fold symmetry at the level of the axonemal plate. Interestingly, we demonstrate that the depletion of the two MKS module proteins induces spontaneous cilia, while the depletion of either NPHP4, CEP290 and RPGRIP1L inhibits the process. Our results constitute the first evidence for a role of conserved TZ proteins in deciliation and open new directions for understanding motile cilia physiology.