paper - craniofacial disease

Question 1

How do proteins in secretory pathway exit ER?

Answer 1

Are packaged into ER exit sites and into COPII coated vesicles

Question 2

what is CLSD?

Answer 2

cranio-lenticulo sutural dysplasia

Question 3

what mutation causes CLSD?

Answer 3

missence substitution mutation in sec23A (single aa substtution)

• disturbs ER to golgi traffiking
• sec23a mutant poorly recruits sec13-sec31 complex

Question 4

what is sec23A?

Answer 4

• a gene coding for copII protein

- so when mutated no membrane copII coat can form so cargo accumulates in exit sites

Question 5

how is copII coat formation initiated?

Answer 5

by sar1 (small GTPase)
- sar1 recruited to SER membrane by sec12 (\gef?)

Question 6

what is sec13-sec31 required for?

Answer 6

required for vesicle formation

- causes coat polymerisation and membrane curvature and membrane fission

Question 7

what does GTP sar1 recruit?

Answer 7

recruits sec23-sec24 (prebudding complex)(adaptor protein)- engages with TMP cytosolic tails
- sec23-sec24 has GAP activity (stimulated by sec13-31) which can hydrolyse GTPSAR1 to GDP SAR1)

Question 8

What happens when SAR1GTP is hydrolysed (by sec23-sec24 GAP activity)?

Answer 8

had reduced affinity to sec23-sec24

-coat dissociates after vesicle budding

Question 9

what do mutations in sar1b and sec23 cause?

Answer 9

sar1b= CMRD (lack of lipoprotein secretion into blood)
sex23a= CLSD

Question 10

why are osteoblasts effected by mutation in sec23A the most?

Answer 10

calvarial osteoblasts have low levels of sec23B

- so when sec23A is mutated there is little/not enough sec23B to compensate.

Question 11

what happens in CLSD fibroblasts?

Answer 11

accumulate tubular profiles lacking an obserable coat (tubular pertrusions from a swollen ER)

Question 12

does sec23a mutant effect sec31?

Answer 12

yes - sec23a mutant binds less sec31a- less coat polymerisation at the membrane???? (although sec23-24 could still bind to membrane)

• especially aapparent when with sar1b
• sec13-31 increases GAP activity of sec23-24 so if this cannot be recruited sar1a cannot by hydrolyed to GDP so sec23-24 does not dissociate from the membrane?????????? - accumulation of cargo?)-also sec13-31 required for copII coat polymerisation?

Question 13

what were differences in sec23-sec24 GAP activity between sar1a and sar1b?

Answer 13

• sar1a (with sec13-31) stimulated GAP activity of WT and mutant sec23-24 (rescued?)
• sar1a has higher affinity for coat (sec13-31)

-sar1b (with sec13-31) did not stimulate GAP activity of ,mutant sec23-24

Question 14

is sorting of copII cargo into ER exit sites affected by sec23a mutant?

Answer 14

no - COPII cargo is recruited at a steady state to ER exit sites (but still no coat )