Department of Molecular Biology and Pharmacology, Washington University School of Medicine, St Louis, MO 63110, USA
(e-mail: kopan{at}molecool.wustl.edu )
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Introduction |
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In the poster, panel A summarizes the proteins in the Notch signal transduction pathway for which biochemical functions have been proposed. Panel B outlines the events that occur in response to ligand binding and during the regulation of transcription and return to the basal state. In panel C a speculative pathway, independent of core components, is presented.
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Panel A |
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Panel B |
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Despite being associated with the presenilin complex, Notch cannot be
cleaved since its extracellular domain somehow blocks presenilin activity.
After ligand binding, neuralized adds a ubiquitin to the intracellular domain
of delta and triggers its endocytosis (2)
(Kramer, 2001). Ligand
endocytosis triggers a conformational change in Notch that permits
metalloproteases to cleave near the membrane at a second site (S2), then the
extracellular domain is trans-endocytosed to the ligand-expressing cell (3)
(Parks et al., 2000
). This
event permits presenilin to cleave Notch at a third site (S3) located within
its transmembrane domain (4) (Mumm and
Kopan, 2000
). NICD is now free to translocate to the nucleus. In
cells expressing both Notch and ligand, ligand interferes with this process by
an unknown mechanism; this interference may be relieved by the neuralized
protein, which targets Delta for degradation
(Kramer, 2001
).
Once in the nucleus, NICD converts CSL from a transcriptional repressor to
a transcriptional activator (5-8). Our current understanding proposes that
this conversion occurs by direct protein-protein interactions between the
Notch intracellular domain, SKIP and CSL, which leads to SMRT/HDAC
dissociation (reviewed by Mumm and Kopan,
2000). This alone is sufficient to express some target genes.
Notch/CSL can recruit histone acetylases
(HATs) to assist in chromatin remodeling, and Mastermind/Lag-3 to activate
additional targets. The metabolism of NICD in the nucleus is controlled by
phosphorylation and ubiquitination by the E3 ubiquitin ligase Sel-10
(Gupta-Rossi et al., 2001
;
Oberg et al., 2001
;
Wu et al., 2001
) and Su(Dx)
(Cornell et al., 1999
). NICD
degradation resets the cell and prepares it for the next round of Notch
signaling.
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Panel C |
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Two specific regions of Notch are required for this activity: the
`Abruptex' region of Notch (EGF repeats 17-29, dark blue)
(Brennan et al., 1997;
Brennan et al., 1999b
;
Ramain et al., 2001
) and the
region C-terminal to the ANK repeats that contains the PEST domain
(Brennan et al., 1997
;
Ramain et al., 2001
). This
region was previously shown to bind Dishevelled (Dsh). Dsh is a mediator of
Wnt signals downstream of the Frizzled (Frz) receptors
(Huelsken and Birchmeier,
2001
). During Drosophila neurogenesis, CSL-dependent
Notch signals restrict neural competence to a small number of cells within an
equivalence group (Kopan and Turner,
1996
). In this newly recognized role, Notch acts to prevent cells
from acquiring neural or myogenic competence earlier in development. Although
it is unclear how Notch mediates this effect, or how many additional proteins
are involved, this activity requires Deltex, a cytoplasmic ring finger protein
(Ramain et al., 2001
) and the
kinase GSK3ß (Sgg) (Brennan et al.,
1999b
; Ramain et al.,
2001
). Deltex and CSL appear to be antagonistic, possibly because
both compete for the ANK domain of Notch. This cytoplasmic Notch activity may
not require proteolysis (1,1'), however, it has not been determined yet
whether presenilin or other proteases are required for the Deltex-dependent
Notch activity. Wnt acts to block Notch either directly, via the Abruptex
domain (Wesley and Saez, 2000
)
(1-2) or, more likely, indirectly by stimulating Dsh to block GSK3 or to
circumvent Deltex/Notch interaction (or both; 1',2'). Interference
with Notch/Deltex activity by Dsh requires the Dsh-binding region of Notch;
deletions of this region and some Abruptex mutations render Notch a
constitutive repressor of neural competence (2')
(Ramain et al., 2001
). Removal
of both Notch and wingless restores competence, suggesting that the only role
of Wnt in acquisition of neural or myogenic competence is to antagonize
Notch.
It is clear that not all the proteins facilitating inhibition of competence have been described. There is at this time no biochemical mechanism proposed for this process. Similar Wnt/Notch interactions in other organisms are yet to be discovered, casting doubt on the generality of these observations. The importance of the Abruptex mutations was recognized early the name was given even before it was realized that locus is identical to Notch but a biochemical explanation for the complex genetic behavior of Abruptex mutations is still lacking. These recent observations may begin to provide an answer.
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References |
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