Posted 15 February 1998

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Participants: Deborah Watson, Ken Kosik, Bart De Strooper, Kieran Breen, Steven Barger, June Kinoshita, Ben Wolozin, Wilma Wasco, Carmela Abrahams, Dmitry Goldgaber, Weiming Xia, Taisuke, Rong, Walker McGraw, Minami, Jean Floring, Chris Weihl.

Note: Transcript has been edited for clarity and accuracy.

Deborah Watson: I'd like to start by asking Dr. DeStrroper about the effects of one copy of PS on APP processing. Is there an intermediate effect between +/+ and -/-?

Bart De Strooper: We did not do extensive experiments on the +/- but there might be a decrease in amyloid peptide production in the +/- cultures as well.

Ken Kosik: But your paper shows a suggestive change in Fig. 1 g and it could be important for the mechanism.

Steven Barger: Steve Younkin's group presented data at Neuroscience from a PS antisense model. Their results were essentially the opposite of this knockout model -- any speculation on the resolution?

Ken Kosik: Responding to Steve Barger: could you tell us how the effect was the opposite?

Bart De Strooper: Hello everybody (with a lot of delay). First again on the PS1+/- .There is as I told, a decrease in amyloid peptide production in the PS1 +/- cutltures. We focussed however until now mostly on the PS1-/-

Steven Barger: They showed that antisense of PS1 INCREASED Aβ 42(43). One caveat is that it was in HEK cells.

Ken Kosik: Did they prove that they completely suppressed the expression of PS1? That's for S.Barger.

Ken Kosik: Weiming, do you have any comments about the heterozygotes?

Weiming Xia: In what aspect of heterozygotes?

Ken Kosik: Regarding Aβ production and CTF production.

Weiming Xia: We have not tried Aβ detection on hetorozygotes.

Steven Barger: No, I don't think there was complete annihilation of PS1; antisense is almost never that complete. So, you're right, there's a quantitative issue to deal with. But this might be a mechanistic hint, as well (?).

Deborah Watson: Do you have any more ideas, Bart, about the residual Aβ production in PS 1 null mice?

Weiming Xia: Is there any linear relationship regarding the decrease of PS1 expression and changes of Aβ, particularly Aβ 42?

Ken Kosik: Bart, could you reiterate why you believe PS1 mutants are gain of function?

Bart De Strooper: to Deborah Watson: either it is PS2 or it is residual activity of γ-secretase.

Rong: Can you explain why the decrease of intracellular Aβ is not so significant Taisuke: Did you see the expression of PS2 in PS1 -/- mice?

Ken Kosik: Bart, do you agree that the decrease applies to both intracellular and secreted a-β?

Weiming Xia: to Bart, any comments on p3?

Ken Kosik: Rong, do you think there are specific secretases for the other fragments?

Taisuke: Did you see the expression of PS2 in PS1 -/- mice?

Bart De Strooper: To everybody, It is impossible for me to really participate in the discussion. I get disconnected every few minutes. To Wxia: p3 behaves like β A4

Rong: To Kosik: At this moment, I don't think so. I may argue that same γ-secretase is responsible for generating Aβ42,40,38,37,..

Ken Kosik: To Rong: does the same secretase create all these fragments? Bart, sorry for the problems you are having, but we do appreciate your responses.

Steven Barger: The gain-of-function slant is satifying since it is consistent with the ability of PS mutants to restore development of somites, spine, etc. in the PS KO's.

Ken Kosik: Steve, are your referring to the Sisodia data about rescue? Wilma, do you have any thoughts about whether decreasing amounts of presenilin increase or decrease Aβ levels?

Steven Barger: Yes, to my knowledge, Sisodia's are only publicly presented data on this issue.

Rong: To Kosik: I don't have definite answers for that, but it is possible.

Ken Kosik: Steve and others: isn't it unusual that so many different mutations in PS1 all lead to gain of function?

Carmela Abrahams: Do Bart's results mean that γ secretase cleavage occurs in ER or Golgi, thus Aβ is completely formed there?

Ken Kosik: I'd like to revisit a previous question -- Bart, why is there the discrepant responses of Aβ in the supernatant where you see a nice effect vs the Aβ in cell extracts where the effect looks minimal?

Steven Barger: Yes, I certainly agree that the number of mutations doesn't fit Occam's razor. One thing that initially occurred to me is that the function we talk about gaining or losing may be two different functions: 1) a developmental role and 2) APP processing.

Ken Kosik: Bart, are you still with us?

Bart De Strooper: Ken, would it be possible to pool the questions and to send them to me by email. I get only part of the discussion on my computer.

June Kinoshita: Apologies to all. Apparently Bart's server is having problems. I will go through the questions for Bart and email them to him. The questions and his responses will be posted on the newsgroup.

Ken Kosik: Thank you June! Steve, even applying gain of function just to APP processing we have a problem with regard to Occam's razor.

June Kinoshita: We can use this time for discussion of these issues among ourselves. I'm sure many interesting questions will grow out of the discussion.

Ken Kosik: Rong, do you then believe the γ secretase is not highly specific for a particular amino acid and are any of these fragments seen in vivo (37, 38...)?

Weiming Xia: Rong, do you think there is one γ-secretase producing Aβ ending at different positions?

Jean Floring: Steve, would you repeat the paradox (vs Younkin's results). My computer crashed.

Bart De Strooper: To all, I am leaving and will do as June suggested.

June Kinoshita: Thank you so much, Bart. Sorry this Brave New Cyberworld isn't always what it's cracked up to be!

Weiming Xia: Thank you, Bart.

Ken Kosik: Thanks Bart.

Deborah Watson: Carmela, do you believe there are other sites besides ER/Golgi where Aβ is produced--if you say endosomes could you also be more specific? (From Kosik)

Carmela Abrahams: Nice work. Thank you Bart.

Steven Barger: Younkin used antisense to decrease PS1 levels in HEK293 cells and showed an increase in Aβ 42(43) (in contrast to the decrease that occurred in these KO cultures).

Carmela Abrahams: Debra, I don't think at this moment we can tell whether Aβ is formed also in endosomes or gets there from the ER. Does anyone who is plugged in know more?

Jean Floring: Thanks, Steve..I'll reserve my opinion about antisense. Anyone: Is there any reason to imagine that the overexpression of APP may have led to the PS-1 effects?

That is, what's going on with normal APP processing? Is the double knockout plus transgene on anyone's list?

Rong: Yes, all of these Aβ fragments have been detected in vivo.

Ken Kosik: Rong, does the concentration of the fragments differ greatly?

Rong: The are more or less associated with Aβ40 and Aβ42.

Steven Barger: To Ken: Actually, having two functions for PS might explain why AD would involve so many mutations: the developmental role would select (in terms of lethality) for mutations that were mild enough to retain the developmental role. But ...

Taisuke: To Jfloring: I transfected only PS2 mutant cDNA into Neuro2a cells, and endogenous Aβ42 was increased.

Steven Barger: ... But once this developmental selection is satisfied, there might be many routes to loss-of-function in the APP processing role.

Ken Kosik: Steve, are you suggesting that the PS1 mutations are loss of function with regard to APP processing?

Rong: I have to leave. It is very nice to discuss with everyone here. Thanks.

Steven Barger: Yes, Ken, I think that's a possibility.

Ken Kosik: I'd like to thank everyone for participating. Please make any last comments now that can be posted to Bart.

June Kinoshita: Thank you Ken for moderating and for putting together the thoughtful discussion.

Steven Barger: Very intriguing work, congratulations!

June Kinoshita: Again, please check the newsgroup for questions and responses from Bart.

Ken Kosik: Thanks everyone. Good-bye.

June Kinoshita: Bye Ken! Thanks to all for participating!

Jean Floring: Steve: what's your email address?