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Summary; Previous
achievements of this research group.
Different groups have been involved in elucidating the
pathways of intracellular routing, the sorting machineries,
and the mechanisms behind peptide loading and blocking the
peptide-groove until the newly synthesized MHC molecules reach
the compartment for peptide loading. We have participated in
various ways towards this goal and here we summarize the
results we have obtained particularly concerning the invariant
chain, MHC-II and CD1d, and interactions with specific
adapters.
- Back in 1990 we found that the cytosolic tail of invariant
chain contains endosomal sorting signals (Bakke and
Dobberstein, 1990) and these signals direct MHC class II to
endosomes (Simonsen et al., 1993). We were also the first to
show that the sorting information consist of two independent
signals; an isoleusine-leusine and a methionine-leucine signal
(Pieters, Bakke and Dobberstein, 1993, B. Bremnes et al.,
1994).
- Ii is able to abrogate peptide binding until it is removed (Roche
et al., 1992; Dodi et al., 1994) and Ii is essential for
efficient transport of the trimeric complex out of the ER (Nijenhuis
et al., 1994)
- As has been confirmed by other groups MHC-II is found in
both early and late endosomes and also lysosomes (Pieters et
al., 1991), making practically all of the endosomal pathway
potential antigen loading compartments (Stang et al., 1998)
- The general belief was that the MHC-II/Ii complex is sorted
directly to endosomes from the trans Golgi network. In the
group of Eric Long (NIH) and in our group we found that
transport via the plasma membrane and endocytosis is just as
important. As we used the same cell lines but different
methods we published this together (Roche et. al., 1993). The
internalization process is rapid and the half-life of the
complex on the plasma membrane is less than 1 minute (Roche
et. al., 1993, B. Bremnes et. al., 1994). Our later studies of
adaptor interactions supports this sorting pathway ((Bremnes
et al., 1998a; Rodionov and Bakke, 1998a; Hofmann et al.,
1999)
- Ii may change the cellular phenotype when transfected into
cells and expressed at relatively high level. This was
observed by Oddmund back in 1989, but it was only later
that we understood the possible significance of this
observation. The mechanisms behind this process are, however,
unknown. We have published this observation using light and
electron-microscopic studies in cell lines that we transfected
both transiently and stably (Romagnoli et al, 1993, Pieters,
Bakke and Dobberstein, 1993, B.Bremnes et al., 1994, Stang and
Bakke, 1997).
- Ii-induced change of phenotype to enlarged endosomes is
correlated with trimerization of Ii, and this is dependent on
a specific domain in the luminal part of Ii, the trimerization
domain (Gedde-Dahl et al., 1997)
- Ii delays endosomal transport from early to late endosomes (Romagnoli
et. al, 1993, Gorvel et. al., 1994, Stang and Bakke, 1997).
- Using antibodies specific the complex of a degradation
product of Ii (CLIP) and MHC class II we found that this
complex was present late in the endosomal pathway. Mature
class II with a foreign peptide is also present in this
compartment and we believe that this is a major peptide
loading compartment (Stang et. al, 1998).
- NMR studies shows that the sorting signal is within an
alfa-helix-strukture and beta turn and the cytoplasmic Ii tail
is a trimer in solution (Motta et al., 1995; Motta et al.,
1997).
- In polarized cells Ii is itself sorted to endosomes via the
basolateral membrane. The two leucine based sorting signals
are also polarized sorting signals and there is one additional
basolateral signal in the cytoplasmic tail. MHC-II alone is
also sorted basolaterally, and Ii contributes to the polarized
transport of these molecules (Simonsen et al., 1997; Simonsen
et al., 1998; Simonsen et al., 1999).
- MHC class II beta chain contains a sorting signal that can
re-internalize MHC-II and present special antigens (Pinet et
al., Nature 1995). This signal is a FL based motif, which is
also a signal for basolateral sorting (Simonsen et al., 1999).
- Using random phage display library we found that the LI
sorting signal within the Ii tail may react with a specific
domain of the adaptor mu chain (Bremnes et al., 1998b). That
the signal binds this chain of the adaptor complexes was
further demonstrated by biochemical means (Rodionov and Bakke,
1998a) and surface plasmon resonance (Hofmann et al., 1999).
- Tyrosine and leucine based sorting signals compete, possibly
using parts of the same sorting machinery (Nordeng and Bakke,
1994)
- The formation of Ii enlarged endosomes is dependent on a
particular charged residue in the cytoplasmic tail (Gregers et
al., submitted) and this sequence alteration changes the
trimerization of the tail as detected by NMR (Nordeng et al.,
2002).
- CD1d harbors sorting information for endosomal compartments
(Rodionov et al., 1999a) and the basolateral sorting signal
consists of two overlapping sorting signals (Rodionov et al.,
2000)
- Invariant chain promotes endosome fusion using both in vivo
and in vitro assays. This process is independent of wortmannin
seems that invariant chain may act as a “docking” protein for
endosome-endosome fusion (Nordeng et al., 2002).
- As shown in a study of live cells and confocal microscopy,
during cytokinesis the endosomes are partitioned using an
ordered, yet imprecise, mechanism, and they are stable
throughout the process (Bergeland et al., 2001).
- We have defined the aminoacid recidues determiningbinding of
leucine signals to AP1 and AP2 (Kongsvik et al., 2002) and AP3
(Rodionov et al., 2002)-
To be continued...
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MDCK cells overexpressing invariant chain. In green is the
early endosomal marker EEA1-GFP, in red lysotracker red.
Micrograph by Frode M. Skjeldal.
Immunogold labelling on ultra-thin cryosection in electron
microscopy. The left picture shows multilaminar MIICs in
immature dendritic cells labelled for MHC class II molecules.
The right
picture shows a MIIC in maturing dendritic cells also labelled
for MHC class II molecules. In addition of class II
redistribution at the cell surface, maturation of dendritic
cells induces morphological changes of MIICs: packaging of
internal membranes and formation of tubules from the limiting
membrane.The picture was awarded at the SCANDEM 2003 meeting. |
