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● ● ● IMMUNOBIOLOGY
Comment on Risueño et al, page 601
Antibody catches T-cell receptor
in
the act
---------------------------------------------------------------------------------------------------------------Michael L. Dustin
NEW YORK UNIVERSITY SCHOOL OF MEDICINE
Risueño and colleagues report that APA1/1 monoclonal antibody can be used to
detect TCRs that have interacted with antigenic MHC-peptide complexes in vivo.
long-standing controversy in T-cell activation is whether the T-cell receptor
(TCR) undergoes conformational changes
upon engaging agonist major histocompatibility complex (MHC)–peptide complexes. In
2002, Gil et al1 demonstrated that TCR ligation by agonist MHC-peptide complexes or
activating antibodies induced binding of the
cytoplasmic adapter protein Nck to a polyproline motif in the cytoplasmic domain of the
CD3⑀ chain. This unmasking provided the
best evidence for a ligand-induced conformational change in the TCR. While the functional necessity of this change for TCR signaling has not been proved, there is no question
that this change is biochemically useful for
identification of TCR complexes that have
been engaged by agonist MHC-peptide
complexes.
The utility of Nck recruitment to this site
in situ is limited because Nck is also recruited
to a downstream signaling complex including
Src homology 2 domain-containing leukocyte
protein of 76 kDa (SLP-76) and Vav, which,
due perhaps to amplification, appear to account for most of the Nck recruited to the immunologic synapse.2 Therefore, another way
to detect this conformational change would
greatly increase its accessibility in intact cells
or in vivo.
In this issue of Blood, Risueño and colleagues show that the commercially available
anti-CD3⑀ antibody APA1/1 detects the same
conformational change in the TCR that is detected by Nck and can do so in fixed tissues. As
with Nck binding, the APA1/1 epitope exposure did not require metabolic activity or signaling through the TCR, so this appears to be
a direct response of the TCR machinery to
agonist MHC-peptide complexes without
intervening events. The APA1/1 epitope thus
provides a new reporter for productive TCR
engagement that is not subject to the interference issues that faced the biologic ligand Nck.
A
396
The APA1/1 antibody detected TCR accumulated in the immunologic synapse formed
by agonist MHC-peptide complexes, but not
by antagonist MHC-peptide complexes. The
APA1/1 antibody reacts with both mouse and
human TCR complexes, indicating that it has
broad potential applications for in vivo experi-
mental immunology and perhaps clinical applications to detect concurrent T-cell activation in short-term cultures or in biopsy specimens. Thus, the use of APA1/1 should greatly
extend the potential applications of the original finding by allowing detection of engaged
TCR in vitro and in vivo and should speed a
deeper understanding of the biologic importance of this conformational change. ■
REFERENCES
1. Gil D, Schamel WW, Montoya M, Sanchez-Madrid F,
Alarcon B. Recruitment of Nck by CD3 epsilon reveals a ligand-induced conformational change essential for T cell receptor signaling and synapse formation. Cell. 2002;109:
901-912.
2. Barda-Saad M, Braiman A, Titerence R, Bunnell SC,
Barr VA, Samelson LE. Dynamic molecular interactions
linking the T cell antigen receptor to the actin cytoskeleton.
Nat Immunol. 2005;6:80-89.
● ● ● HEMOSTASIS
Comment on Albrecht et al, page 542
An
ABC for Scott syndrome?
---------------------------------------------------------------------------------------------------------------Florence Toti and Jean-Marie Freyssinet
U.143 INSERM UNIVERSITÉ PARIS XI; UNIVERSITÉ LOUIS PASTEUR STRASBOURG
Scott syndrome is an extremely rare inherited hemorrhagic disorder linked to the
lack of exposure of procoagulant phosphatidylserine (PS) to the external leaflet of
the plasma membrane of activated platelets and other hematologic lineages.1-3 The
loss of membrane lipid asymmetry, however, occurs in virtually all eukaryotic cells
upon stimulation (including apoptosis)3 and is associated with more ancestral functions than hemostasis. Hence, it is of prime interest to identify effectors of this basic process.
o date, only 3 documented cases of Scott
syndrome have been reported, which is insufficient to allow the identification of positional
candidate genes. It should be emphasized that
none of these patients exhibited clinical symptoms other than bleeding, suggesting that the
rapid PS egress for an efficient platelet hemostatic response and slower membrane remodeling occurring during apoptosis are under different control mechanisms. This does not,
however, rule out that some or most of the actors
can be shared. On the one hand, several studies
have pointed to adenosine triphosphate (ATP)–
binding cassette (ABC) transporters as possible
phospholipid translocases (in particular, some
that fulfill multidrug efflux function), but none
has been confirmed as a true PS floppase. On the
other hand, a nonspecific and energy-indepen-
T
dent family of so-called scramblases has been
proposed. But canonical phospholipid scramblase 1 (PLSCR1) is normally expressed in Scott
cells, and other members or other candidate(s)
remain to be characterized as effectors of bidirectional phospholipid transmembrane redistribution. Another ABC family member, ABC transporter A1 (ABCA1), involved in phagocytosis
and cholesterol reverse transport and mutated in
Tangier dyslipidemia, has been considered after
the targeted deletion of the corresponding locus
resulted in a phenotype evocative of partial Scott
syndrome.4 In this issue of Blood, Albrecht and
colleagues precisely report 2 types of mutations
that may contribute to the Scott phenotype.
The patient was found heterozygous for a
novel missense substitution in ABCA1 R1925Q ,
which has not been previously reported in the
15 JULY 2005 I VOLUME 106, NUMBER 2
blood
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Membrane remodeling in normal and Scott cells. In the plasma membrane, dynamic maintenance or loss of asymmetric phospholipid distribution results from opposite fluxes under the control of specific transporters governing
inward (flip) or outward (flop) translocation. In the resting membrane, the flippase activity of aminophospholipid
translocase is prominent and subtained (left, yellow ellipse). Thus, aminophospholipids, phosphatidylserine (PS;
red negative polar heads), and phosphatidylethanolamine (PE; open polar heads) are sequestered into the inner
leaflet. Following stimulation, specific vectorial floppase and/or nonspecific bidirectional scramblase activities are
rapidly turned on, and PS and PE are moved to the outer leaflet (upper right). Flippase activity is shut down, leading to transient mass imbalance between the two leaflets. Membrane blebbing ultimately resolves into microvesicle or microparticle (MP) shedding after cytoskeleton degradation by calcium-dependent proteolysis. Stimulated cell membrane and MP therefore expose PS, which is particularly crucial in platelet procoagulant response
for the calcium-dependent assembly of the cascade of clotting enzyme complexes. The assembly of these complexes, composed of an enzyme (E) and a cofactor (C) in the activation of a substrate (S), culminates in the generation of sufficient thrombin for efficient hemostasis. In resting Scott cells, flippase activity is indeed operational;
stimulation remains without effect on the swift induction of floppase and/or scramblase and consecutive membrane shedding, explaining the resulting hemorrhagic phenotype (lower right). During the slower process of apoptotic cell death, however, PS egress is normal in Scott cells. As yet, floppase candidates have remained elusive.
relatively large cohort of Tangier patients described in the literature. In vitro expression experiments revealed an impaired traffic of
ABCA1 R1925Q to the plasma membrane, and
sequence comparisons pointed to a conserved
functional importance of this residue. The sec-
blood 1 5 J U L Y 2 0 0 5 I V O L U M E 1 0 6 , N U M B E R 2
ond mutation, which remains to be identified,
accounts for significant reduction of ABCA1
mRNA levels and may affect a trans-acting regulatory gene. Using a retroviral expression system, complementation of the defect of PS externalization proved feasible in Epstein-Barr virus
(EBV)–transformed Scott B cells. Unfortunately, no investigation of the protein expression
levels was possible, most likely as a result of the
lack of adequate antibody.
How these mutations relate to Scott syndrome may be debated, but this observation at
least provides the first link between a defect in
a transbilayer phospholipid transport pathway, that of ABCA1 here, and the bleeding
phenotype. In light of other genetic disorders,
other mutated or deleted elements may be
considered, including other ABC true transporters or regulators such as ABCA1 itself, as
already suggested for the latter.5
In Scott syndrome, bleeding episodes are
moderate when compared with spontaneous
hemorrhages in severe hemophilia, which affect a much greater number of patients. It has
to be kept in mind that the Scott defect most
likely can be traced back to a hematopoietic
stem cell line or even earlier. Hence, the incapacity of PS externalization may turn lethal
because of the prime role of PS as a recognition
determinant for phagocytosis, for instance.3
This could explain the surprising rarity of
Scott syndrome patients who might have survived as a result of the intervention of an opportune rescue mechanism. ■
REFERENCES
1. Weiss HJ. Scott syndrome: a disorder of platelet coagulant activity. Semin Hematol. 1994;31:312-319.
2. Toti F, Satta N, Fressinaud E, Meyer D, Freyssinet JM.
Scott syndrome, characterized by impaired transmembrane
migration of procoagulant phosphatidylserine and hemorrhagic complications, is an inherited disorder. Blood. 1996;
87:1409-1415.
3. Zwaal RF, Comfurius P, Bevers EM. Surface exposure
of phosphatidylserine in pathological cells. Cell Mol Life
Sci. Prepublished on March 9, 2005, as DOI 10.1007/
s00018-005-4527-3. (Now available as 2005;62:971-988).
4. Hamon Y, Broccardo C, Chambenoit O, et al. ABC1
promotes engulfment of apoptotic cells and transbilayer
redistribution of phosphatidylserine. Nat Cell Biol.
2000;2:399-406.
5. Szakacs G, Langmann T, Ozvegy C, et al. Characterization of the ATPase cycle of human ABCA1: implications for
its function as a regulator rather than an active transporter.
Biochem Biophys Res Commun. 2001;288:1258-1264.
397
From www.bloodjournal.org by guest on June 18, 2017. For personal use only.
2005 106: 396-397
doi:10.1182/blood-2005-04-1716
An ABC for Scott syndrome?
Florence Toti and Jean-Marie Freyssinet
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