Atypical CF Flashcards
What are the characteristics of atypical CF?
Mild symptoms
1 or 0 CFTR mutations
carriers shouldnt have symptoms
modifier genes involved?
What was the hypothesis with atypical CF?
Other genes are mutated in these patients
one that stood out as having potential in mimicking the CF airway systems= ENac
These patients have gain of function ENac mutations- enhanced absorption of sodium- deplete PCL- mucus thick= patients have CF like symptoms
What did they find when they screened 31 atypical CF patients?
11 mutations in the alpha subunit
7 mutations in the beta subunit
8 mutations in the gamma subunit
How did they find the function of the mutated ENac channels?
They looked at the amiloride-sensitive current and expressed it as a % of the WT for each mutation
3,4,6 and 7- enhanced sodium currents- greater function of sodium channels
8,9 and 10- didnt show anything at all- maybe these mutations arent the problem
2 and 5- reduced sodium currents- against the hypothesis
What did they think was going on in the gof mutations in the Enac channels?
Levels of the ENac were essentially the same as WT
thought it was like to be a change in the open probability
What study did they do in Atypical patients to validate the results of the previous study on GoF Enac?
Used atypical patients
measured their nasal Vte- wanted to validate that there were gain of function mutations in ENac and that chloride handling was the same
What was the results of the study of atypical CF patients?
People with w493r mutation- resting vte is -25mv- inbetween normal and a cf patient
squirt low cl solution- shift in vte of 15mv- similar to normal
added amiloride- 20mv shift in potential- much bigger than normal shift expected- similar to a cf patient
What does the alphaW493R mutation in Enac channels cause?
Gain of function mutation in the extracellular loop of the alpha subunit
overexpressed the mutant- huge increase in amiloride sensitive current
What is Na feedback inhibition? How was it different in the W493R?
Process by which Enac channels are endocytosed from the apical membrane
Increase in IC Na stimulated endocytosis- ENac removed from the membrane- then recycled back- cyclical process
Drop in WT current in a high NA concentration is almost the same as in the mutant
No change in sodium feedback inhibition in the mutant
What happens when ENac is cleaved?
Open probability increases
uncleaved Enac= near silent, very low Po, very small currents
How do mutant channels respond to chymotrypsin?
Mutant channels dont respond to c.trypsin
when looking at single channel recordings- Po of the channel is high before c.tyrpsin and doesn’t change when c.trypsin is added
The mutant gating is different to the WT irrespective of cleavage
What is sodium self inhibition?
The amount of sodium going into the cell regulates the Po of the channels
High IC Na reduces the channels open probability
Dont see this is mutant channels
Summary of the work on alpha W493R
Mutation increases ENac currents
no change in sodium feedback inhibition
high currents not due to increased cleavage
Loss of self-inhibition- high currents= greater water absorption and CF like symptoms
Whats another mutation that has been explored in atypical CF?
betaV348M
GOF ENac mutation- increased sodium current
How is the Po different in the beta subunit mutation, how did they measure this?
Use MTSET
MTSET stabilises the channels in an open state- open and stay open- Po of 1
Increased Po in the mutant
