B cell and CD8- dendritic cell survival require CD74 proteolysis by the intramembrane endopeptidase SPPL2A
Date
2016
Authors
Bergmann, Hannes
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Abstract
Unknown mechanisms that cause failures in B cell development can
lead to humoral immunodeficiencies, autoimmune disorders or B
cell malignancies. This thesis demonstrates that the endosomal
intramembrane cleaving protease signal-peptide-peptidase-like 2A
(SPPL2A) is critical for survival of developing B cells and
reveals SPPL2A-deficiency as a previously unknown mechanism of B
cell loss. Mice with an ENU-mutagenesis-induced inactivating
mutation in Sppl2a exhibited profound humoral immunodeficiency
and specifically lacked mature B cells, phenotypically mirroring
BAFF-deficient mice. Surviving B cells were characterised by
abnormal endosomal membrane expansions and low surface BAFFR and
BCR expression. Mature B cells, but not BAFFR and BCR surface
levels, could be rescued by over-expression of the pro-survival
protein BCL2, indicating that SPPL2A-deficient B cells failed to
mature due to a survival defect rather than developmental
arrest.
On a molecular level SPPL2A-deficiency blocked the ultimate
intramembrane proteolytic processing step of CD74 (MHCII
invariant chain), causing a dramatic build-up of residual 8kD
CD74 (p8-CD74) membrane stubs leading to endosomal enlargement.
The accumulation of p8-CD74 was responsible for the loss of
mature B cells, as inhibition of p8-CD74 build-up by deletion of
CD74 in SPPL2A-deficient animals reconstituted mature B cell
numbers to levels seen in mice solely deficient in CD74, and
restored surface BCR and BAFFR expression on B cells.
Interestingly, endosomal enlargement has been similarly observed
in B cells from Cathepsin S-deficient mice that also accumulate a
CD74 degradation product, p10-CD74. Despite more severe endosomal
expansion in these B cells, they did not exhibit survival
abnormalities. This finding illustrates the distinct capabilities
of p10-CD74 and p8-CD74 to regulate endosomal biology and B cell
survival.
Despite its role in CD74 metabolism, SPPL2A appeared disconnected
from the classical sequence of endosomal CD74 degradation steps,
as p8-CD74 production required neither penultimate processing by
Cathepsin S nor MHCII-chaperoning. Additionally, it was found
that p8-CD74 already accumulated in immature B cells from the
bone marrow of SPPL2A-deficient mice, which contrasted with
Cathepsin S-deficient mice where CD74 accumulation first occurred
in transitional B cells. This indicates that Cathepsin S
processes CD74 later than SPPL2A during B cell development.
p8-CD74 processing by SPPL2A was logically expected to play a
role in other CD74-expressing immune cells. However, only CD8-
dendritic cell (DC) numbers were reduced in SPPL2A-deficient
mice, whilst CD8+ DC and pDC subsets, as well as macrophages and
monocytes, appeared to survive. CD8- DCs showed elevated surface
MHCII levels, whereas CD8+ DCs were characterised by an
MHCII-chaperoning-dependent 10-fold increase in CD74 surface
expression, highlighting differences in the wiring of MHCII and
CD74 trafficking responses in these DC subsets. Functionally,
SPPL2A-inactivation interfered with the ability of spleen DCs to
present antigen on MHCII to CD4+ T cells, but not with MHCI
associated cross-presentation.
In summary, these findings reveal that SPPL2A mediated CD74
intramembrane proteolysis is a previously unknown mechanism
required for humoral immunity, and survival of developing B cells
and CD8- DCs. Targeting SPPL2A or CD74 may provide new
therapeutic avenues for the control of endosome and B cell
related diseases in the future.
Description
Keywords
B cell development, dendritic cell development, SPPL2A, CD74, invariant chain, MHCII, Cathepsin S, antigen presentation, MIIC, endosome, intramembrane proteolysis, RIP, i-CLiPs, membrane protein degradation, storage disease, multilamellar bodies
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Thesis (PhD)
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DOI
10.25911/5d7634cebddd6