发信人: reer (), 信区: Biology
标 题: Re: MCBJC, cell division, Jan 20 2003, by Assasin
发信站: The unknown SPACE (Sun Jan 26 18:38:34 2003) WWW-POST

A good CSF extract should be arrested @ metaphase.
After the addition of calcium or fertilization which causes a transient
increase in cytoplasmic calcium conc., the CSF will be inactivated, following
by APC activation, cyclin B destruction and mitotic exit. And this process is
roughly 30 min. It is easy to collect phosphorylated separase before the
addition of calcium. To collect the unphosphorylated one, like you say, you
can throw sperm chromatin into extract and monitor the progresstion by DAPI or
some other fluorescent dye of DNA.
Actually, can you guys answer me why they use human proteins in this study?
~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~
hard for me to understand.

For Q6, another regulation of separase which is introduced by phosphorylation
can be so-well fit into the network of cell cycle regulation( phosphorylation
counts for much relationship between proteins, even proteolysis is dependent
on phosphorylation although there may be some exception.) In Marc's paper "
the timing of events in mitosis", they proposed two extreme models of
regulation of the timing of events in each phase of the cell cycle, which are
"regulator-controlled model" and "substrate-controlled model" Their conclution
is that "during early mitosis the timing of biochemical events and
morphological events is at least partly controlled by the responses of the
substrates themselves to a common set of signals." Now they may add separase
as another substrate which can controll the downstream events.

【 在 assasin (冷血---何时能被加热？) 的大作中提到: 】
: 【 在 reer () 的大作中提到: 】
: : If I am wrong, correct me.
: : For Q5, if they use CSF extract( a kind of Xenopus extract, which should
be
: : natually mitotic extract, once you add Calcium, a good CSF extract will
: start
: : to cycle.), they can avoid the possible artifact.
: ~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~
: The P-level reaches the highest just before metaphase-anaphase transition
: (MAT) and goes down very quickly at the onset of annaphase. MAT is very
quick.
: I am wondering when sampling the rection, how do you know exactly at which
: phase your CSF is. Oh, maybe you can sample every 5 minutes. Or we can add
: some fluorescent cellular makers (green chromatid?) into the extract and
: monitor the progression of cell cycle under microscope. .......?
:
: The problem is in their
: : paper they use all human proteins(separase, securin and cohesin.) If they
: use
: : CSF extract, the proteins they can analyze for MS are Xenopus proteins.
What
: I
: : don't understand is why they use human proteins.
: :
: : For Q6,
: : Nagao K, Yanagida M.
: : Regulating sister chromatid separation by separase phosphorylation.
: : Dev Cell. 2002 Jan;2(1):2-4. Review.
: : PMID: 11782307
: : I cannot get the full text paper. I believe this one is talking about
Marc's
: : cell paper.
: :
: : For me, this paper provides a new regulation point in mitosis. This
: mechanism
: : happens to be related to CDC2/cycB activity.
: ~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~
: good point, i agree.
:
:
: : 在 assasin (冷血---何时能被加热？) 的大作中提到: 】
: : : MCBJC, cell division, Jan 20 2003, by Assasin
: : :
: : : paper:
: : :
: : : Stemmann, O., Zou, H., Gerber, S.A., Gygi, S.P., and Kirschner, M.W.
: (2001)
: : : Dual inhibition of sister chromatid seperation at metaphase. Cell 107,
: : : 715-726.
: : :
: : : Questions:
: : :
: : : 1. Describe the differences between S. cerevisiae chromosome seperation
: : : pathway and that of metazoan cells.


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