Subject: Re: Help FlyBase please about ey-Dominant
Hi Michael,
I'll see what I can do to help clarify things.
the info on these breakpoints probabaly comes from a response sheet:
>
>T(2;3)AlpXTl, 24A1,2;80
>T(2;3)AlpDTD5, 24A1; 68F1,2
>Dp(2;4)AlpeyeD,23DE;24A1;102D
>T(2;4)AlpXt2, 24A1;101D
>
>We believe there are no problems with 'T(2;3)AlpXTl', and that it is
>the same as T(2;3)XT1 mentioned in the Alp entry in Lindsley and Zimm
>(p24).  It does not have its own entry in the Aberrations section of
>L&Z; in FlyBase it is now T(2;3)AlpXT1 (<up></up> enclose superscripts).
I agree
>
>We believe (slightly less confidently) that 'T(2;3)AlpDTD5' is the
>aberration now known as T(2;3)DTD57, which was listed in L&Z under its
>provisional name (in a Gelbart stock list) of T(2;3)434.128.
correct
>
>The real 'fun' concerns the other two.  The L&Z entry for Alp links
>Alp2 with an aberration T(2;4)X2; is this the 'T(2;4)AlpXt2' listed
>above?
yes
T(2;4) Alp X2 and Dp (2;4) eyD are independently isolated chromosomes. the
designations above are ok.
As far as we can tell eyD is in fact two gain of functions -it shows an
eyeless phenotype and it shows an Alp dominant phenoytpe. We believe that
the Alp dominant is explainable on the basis of the transposition of
material from the 24A1,2 breakpoint (as for the other Alp dominants). We
have assumed, but not tested that the eyD phenotype is due to the site of
insertion, since no other Alp dominant shows an eyeless phenotype.
>To cap it all, we also have a listing for an allele AlpXT2, which is
>linked to a T(2;3) with breaks identical to those of T(2;3)AlpXT1.
>
this is wrong
>In summary, here are my questions.  Any help you can give, particularly
>if you can cite published references, will help us a lot.
All of this is unpublished, so you'll have to cite my unpublished data (I
know we should publish it!)
>(1) Is Alp sequenced?  I have a suggestive note but no accession number.
yes, but....
We have done a lot of work on this. We have found lack of function mutations
and gain of function mutations. There are 6 independent gain of function
alleles that break within 15 -20 kb in a region between 2 transcription
units. The dominant phenotypes segregate with the distal arms of the
breakpoints when we can test it (three unambiguous cases and eyD).
Paradoxically, the lack of function mutant is associated with the
transcription unit on the proximal side of the breakpoints. We have sequence
the genes on both sides of the breakpoint(s). Both are homologous to odd
skipped (the proximal one is the same as Doug Coulter's odd-related-gene).
Niether is responsible for the Alp dominant phenoytpe. (we sequenced the
coding region of the transcription unit distal to the breakpioint from a
chromosome carrying an ems induced revertant of the dominant phenotpye-
there were no changes in the aa seqeunce- therfore that can't be the gene).
These data suggest that the gain of function is due to a regulatory element
for leg expression that we know lies distal to the breakpoints of all
rearrangements and that it has nothing, as such, to do with the genes on
either side that were mutated to cause lack of function phenoytpes.
I think that this is genetically interesting and that we should write it
down, but I haven't really thought abouyt how to do it, in view of other
time pressures. Any thoughts from you on whether this should be written down
(and if so where to send it) would be welcome.
>(2) What is the evidence that Alp is transposed onto cme4 in eyD?
The evidence that the duplicated material in eyD is of 2nd chrosomsome
origin (from the Alp region) comes from cytology by Gerd Jurgens. Gerd
looked at a cross between ey D and T(2;4) Alp X2, and saw pairing which
revealed the 24A banding pattern (that we knew was associated with the
T(2;4) Alp X2 breakpoint). This suggested that the transposed material in
eyD is of second chromosome origin. Also, the eyD chrosomome has an Alp
dominant leg phenotype. This is not published anywhere as far as I can
recall, and you have obviously checked. It can be cited as Cohen and Jurgens
unpublished. We have also mapped the eye breakpoint moelecularly to our walk.
>(3) The cytological information we have indicates that the insertion is a
>    reverse duplication of the material (Adelaide reports that cme4 looks
>    Y-shaped on account of the two inserted copies synapsing).  Have you
>    any data on this point?
no
>(4) Is there evidence that Alp (or any other inserted gene) is mutant in
>    eyD, as opposed to the other option that the entire phenotype is due
>    to disruption of cme4 genes (possibly ey, possibly others) and/or to
>    expression of the extra wildtype copy/ies of an inserted gene, eg Alp?
The evidence that the Alp phenotype derives from the insertion of 2nd
chromosome material is outlined above
>(5) Is T(2;3)DTD57 an Alp mutation, specifically the one suggested above?
yes. this is DTD57. It has an Alp dominant leg phenotype and the break is
molecularly mapped.
>(6) Is Alp2 associated with a T(2;3), a T(2;4) or a Tp(2;4)?
T(2;4)Alp X2 is a reciprocal translocation stock. X2 cytology is 24A1,2
reciprocally translocated to 101D (cytology by Jurgens). It is distinct from
Dp (2;4) eyD.
>(7) Is AlpXT2 the same as Alp2?  If not, what is its associated aberr?
this is the same thing
>(8) Is the Alp allele on eyD (if any) either Alp2 or AlpXT2?
no, they are independent
eyD is the original designation, you could perhaps call it Dp (2;4) eyD,
or Dp 2;4) AlpeyD. the former is more conservative, but the latter reflect
that it is apparently two independent gain of function mutants.
Hope this helps. I would welcome having the chance to discuss the data on
this with you in some detail to see if we have a publishable story. As you
see we've done a lot of work on it. Let me know what you think
Best regards,
Steve
..
Subject: Re: Alp & eyD
At the time we were convinced that the transcript distal to the
translocation bkpts had to be the Alp gene. As you see, subsequent analysis
made it clear that this transcription unit cannot be the Alp GOF. I'd prefer
if it is made clear that the transcript is the odd-like gene distal to
org-1, and not refer to it as Alp, since that could be confusing. We would
like to reserve the right to name it properly. We have mutants and are
looking at some phenotypic stuff with Jackle, so it may get a 'proper' name
soon.
Steve
..
Subject: Re: Alp & eyD
Thanks Steve. I will contact Coulter about the org genes. We
have org1 and also an org2, with no reference, said to be in
24C ! I guess this might be yr next distal transcript.
This was curated from yr Mech Dev paper: is it wrong ?
\*p Alp is expressed in the embryo in a series of segmentally repeated
\*p stripes that lie posteriorly adjacent to the en stripes in each segment.
\*p In germ band retracted embryos Alp is also expressed in a small cluster
\*p of cells in the thoracic segments, and may be related to the appearance
\*p of dorsal disc primordia.
or should it be made clear that this refers to the pattern of the
(unnamed) distal transcript ?
Michael
..
Subject: Help FlyBase please about ey-Dominant
Dear Steve
Steve -Steve Henikoff pointed out to us that there
are problems with the FlyBase Alp/eyD records. After
some detective work we think we now know just what the
problems are. This is probably all a bit much, but any help
you can give would be great.
The start of the problem
is that, in the distant pre-FlyBase past, I seem to have implicated you
as the source of the belief that the 4th-cme insertion is of material
including Alp.  We have searched quite hard for published confirmation
of this, by you or anyone else, without success.  The best we have
found is that I quoted you (personal comm.) in my Laboratory Handbook
(p770) and that both John Merriam and I have notes to the effect that
you announced this at the ADRC in Chicago in 1991 (though I see it is
not mentioned in the abstract).
The confusion is a good deal wider than that, unfortunately.  Merriam's
notes indicate that your ADRC poster (or possibly your reply to one of
his request forms) mentioned the following aberrations; the frequency
of typos in his files was vast, so I will err on the side of caution
and quote verbatim.
T(2;3)AlpXTl, 24A1,2;80
T(2;3)AlpDTD5, 24A1; 68F1,2
Dp(2;4)AlpeyeD,23DE;24A1;102D
T(2;4)AlpXt2, 24A1;101D
We believe there are no problems with 'T(2;3)AlpXTl', and that it is
the same as T(2;3)XT1 mentioned in the Alp entry in Lindsley and Zimm
(p24).  It does not have its own entry in the Aberrations section of
L&Z; in FlyBase it is now T(2;3)AlpXT1 (<up></up> enclose superscripts).
We believe (slightly less confidently) that 'T(2;3)AlpDTD5' is the
aberration now known as T(2;3)DTD57, which was listed in L&Z under its
provisional name (in a Gelbart stock list) of T(2;3)434.128.
The real 'fun' concerns the other two.  The L&Z entry for Alp links
Alp2 with an aberration T(2;4)X2; is this the 'T(2;4)AlpXt2' listed
above?  The problem is that not only is T(2;4)X2 absent from the
Aberrations section of L&Z, but there is instead a mention of Tp(2;4)X2
on p1058, which horrifyingly is stated as 'Mutant for Alp2 and eyD'.
You may recall that L&Z instituted a change in the definition of what
was a translocation and what a transposition.  This was incompletely
implemented, and there are many cases where an aberration is called a
Tp in its own entry but is still called a T when referred to in free
text elsewhere.  We have so far presumed that this is a case of that.
However, the case is of course very weak, not least because the
deficiency segregant from eyD is long lost (if it was ever recovered
at all).  We now suspect that two independent mutations have mistakenly
been merged, both in L&Z and in FlyBase.
To cap it all, we also have a listing for an allele AlpXT2, which is
linked to a T(2;3) with breaks identical to those of T(2;3)AlpXT1.
In summary, here are my questions.  Any help you can give, particularly
if you can cite published references, will help us a lot.
(1) Is Alp sequenced?  I have a suggestive note but no accession number.
(2) What is the evidence that Alp is transposed onto cme4 in eyD?
(3) The cytological information we have indicates that the insertion is a
    reverse duplication of the material (Adelaide reports that cme4 looks
    Y-shaped on account of the two inserted copies synapsing).  Have you
    any data on this point?
(4) Is there evidence that Alp (or any other inserted gene) is mutant in
    eyD, as opposed to the other option that the entire phenotype is due
    to disruption of cme4 genes (possibly ey, possibly others) and/or to
    expression of the extra wildtype copy/ies of an inserted gene, eg Alp?
(5) Is T(2;3)DTD57 an Alp mutation, specifically the one suggested above?
(6) Is Alp2 associated with a T(2;3), a T(2;4) or a Tp(2;4)?
(7) Is AlpXT2 the same as Alp2?  If not, what is its associated aberr?
(8) Is the Alp allele on eyD (if any) either Alp2 or AlpXT2?  If not,
    does it have a published designation?  (We currently call it AlpeD.)
Best wishes
Michael