The circadian system in cystic fibrosis mice is regulated by histone deacetylase 6

Abstract

Disordered sleep experienced by people with cystic fibrosis (CF) suggest a possible disruption in circadian regulation being associated with the loss of cystic fibrosis transmembrane conductance regulator (Cftr) function. To test this hypothesis, circadian regulation was assessed in a F508del/F508del CF mouse model. CF mice exhibited significant alterations in both timing of locomotor activity and in mean activity per hour in both light-dark (LD) and dark-dark (DD) photoperiods compared to wild type (WT) controls. It was also noted that in DD, periodicity increased in CF mice, while shortening in WT mice as is expected. CF mice also exhibited altered timing of circadian gene expression and a reduction of melatonin production at all time points. Mechanistically, the role of microtubules in regulating these outcomes was explored. Mice lacking expression of tubulin polymerization promoting protein (Tppp) effectively mimicked CF mouse phenotypes with each measured outcome. Depleting expression of the microtubule regulatory protein histone deacetylase 6 (Hdac6) from CF mice (CF/Hdac6) resulted in the reversal of each phenotype to WT profiles. These data demonstrate an innate disruption of circadian regulation in CF mice and identify a novel microtubule-related mechanism leading to this disruption that can be targeted for therapeutic intervention.

In vivo impact of tubulin polymerization promoting protein (Tppp) knockout to the airway inflammatory response

“…To determine if the Tppp/HDAC6 pathway could be responsible for CF-related airway inflammation, we examined Tppp − / − mice. We have previously shown that Tppp − / − MNE cells have WT levels of CFTR function so we can conclude that any CF-like phenotypes in these mice are not due to a secondary impact on CFTR activity 21 .…”

“…This hypothesis stems from previous observations that Hdac6 depletion from CF mice normalized responses to this challenge suggesting a microtubule-related mechanism 5 . We have also recently demonstrated that Tppp − / − mice effectively replicate another phenotype of CF mice, circadian timing dysregulation 21 , 28 . To determine if the Tppp/HDAC6 pathway could be responsible for CF-related airway inflammation, we examined Tppp − / − mice.…”

“…We have also demonstrated that knocking down expression of Tppp in an immortalized epithelial cell line reproduced these microtubule phenotoypes, consistent with the function of Tppp as an enhancer of microtubule elongation and as an HDAC6 inhibitor 12 , 20 . Inhibition of HDAC6 has been shown to have significant benefit in addressing multiple CF phenotypes 5 , 21 , 26 . Tppp has also been identified as a candidate modifier gene associating with CF airway disease severity in GWAS studies 27 .…”

“…The trend seems similar in male and female mice, so a gender impact seems unlikely. In our previous work showing circadian disruptions in CF (F508del) mice and Tppp − / − mice, we did not examine the impact of Tppp heterozygosity on circadian regulation 21 , 28 . It is well known that sleep disruption can increase inflammation, so it is possible that Tppp + / − mice have a level of sleep disruption that is sufficient to impact monocyte recruitment 29 .…”

“…Observing the phenotype of a mouse lacking expression of Tppp ( Tppp − / − model) investigates the role of microtubule dysregulation in inflammation by directly affecting a protein known to be responsible for microtubule stabilization independently of other pathology. Previous studies have shown that Tppp − / − cells have WT levels of CFTR activity 21 . The hypothesis of this study is that disruption of microtubule stability through knockout of the Tppp gene will result in a hyperinflammatory state.…”