{"id":{"repo_id":"freiburg-diss","oai_identifier":"oai:freidok.uni-freiburg.de:1561"},"canonical_url":"https://search.dev.ndltd.org/etd/freiburg-diss/oai:freidok.uni-freiburg.de:1561","repository":{"repo_id":"freiburg-diss","name":"University of Freiburg","base_url":"https://freidok.uni-freiburg.de/oai/oai2.php"},"display":{"title":"The analysis of B cell antigen receptor (BCR) mutants deficient in antigen-independent signaling reveals novel BCR-associated proteins","abstract":"The B cell antigen receptor (BCR) is composed of the membrane-bound immunoglobulin (mIg)
and the Ig-alpha/Ig-beta heterodimer and plays a central role in development, survival, and activation
of B lymphocytes. During B cell development, the BCR seems to be able to generate signals in
an antigen-dependent and -independent fashion. Upon binding to antigen, the BCR emits a
negative selection signal for immature and an activation signal for mature B cells. In the
absence of antigen, the BCR signals positive selection of immature and maintenance for mature
B cells. The BCR-dependent, antigen-independent signaling mode can be analyzed by
stimulating B cells with the phosphatase inhibitor pervanadate (PV). Analysis of substrate
tyrosine phosphorylation after PV-treatment of different B cell lines expressing BCR mutants
revealed drastic differences in phosphorylation of two signaling proteins namely the SH2-
domain-containing inositolpolyphosphate 5’-phosphatase (SHIP) and the protein kinase C delta.
Tyrosine phosphorylation of SHIP in response to PV stimulation was critically dependent on
the integrity of the Ig-alpha C-terminus. SHIP could be phosphorylated by both BCR-proximal
protein tyrosine kinases, Lyn and Syk. The phosphorylation of SHIP by Syk, but not that by
Lyn, was strictly dependent on the presence of an integer BCR complex, indicating that Syk
requires BCR expression for the phosphorylation of SHIP. Furthermore, the absence of proper
SHIP activation led to a disturbed balance of PKB and MAPK pathway activation, suggesting
that SHIP is a crucial regulator of the antigen-independent maintenance signal in B cells.
In contrast to SHIP, PKC-delta was able to bind in vitro to phosphorylated peptides derived from
the Ig-alpha C-terminus. As observed by native gel electrophoresis PKC-delta can be purified with the
BCR complex. The PKC-delta/BCR binding was dependent on the C-terminus of Ig-alpha. Moreover,
by confocal studies, PKC-delta was found to influence BCR/Syk localization, probably by inducing
BCR/Syk internalization. These data suggest, together with published data from PKC-delta
deficient mice that PKC-delta can terminate BCR signaling by binding to the BCR.","abstract_html":"The B cell antigen receptor (BCR) is composed of the membrane-bound immunoglobulin (mIg) <br>and the Ig-alpha/Ig-beta heterodimer and plays a central role in development, survival, and activation <br>of B lymphocytes. During B cell development, the BCR seems to be able to generate signals in <br>an antigen-dependent and -independent fashion. Upon binding to antigen, the BCR emits a <br>negative selection signal for immature and an activation signal for mature B cells. In the <br>absence of antigen, the BCR signals positive selection of immature and maintenance for mature <br>B cells. The BCR-dependent, antigen-independent signaling mode can be analyzed by <br>stimulating B cells with the phosphatase inhibitor pervanadate (PV). Analysis of substrate <br>tyrosine phosphorylation after PV-treatment of different B cell lines expressing BCR mutants <br>revealed drastic differences in phosphorylation of two signaling proteins namely the SH2- <br>domain-containing inositolpolyphosphate 5’-phosphatase (SHIP) and the protein kinase C delta. <br>Tyrosine phosphorylation of SHIP in response to PV stimulation was critically dependent on <br>the integrity of the Ig-alpha C-terminus. SHIP could be phosphorylated by both BCR-proximal <br>protein tyrosine kinases, Lyn and Syk. The phosphorylation of SHIP by Syk, but not that by <br>Lyn, was strictly dependent on the presence of an integer BCR complex, indicating that Syk <br>requires BCR expression for the phosphorylation of SHIP. Furthermore, the absence of proper <br>SHIP activation led to a disturbed balance of PKB and MAPK pathway activation, suggesting <br>that SHIP is a crucial regulator of the antigen-independent maintenance signal in B cells. <br>In contrast to SHIP, PKC-delta was able to bind in vitro to phosphorylated peptides derived from <br>the Ig-alpha C-terminus. As observed by native gel electrophoresis PKC-delta can be purified with the <br>BCR complex. The PKC-delta/BCR binding was dependent on the C-terminus of Ig-alpha. Moreover, <br>by confocal studies, PKC-delta was found to influence BCR/Syk localization, probably by inducing <br>BCR/Syk internalization. These data suggest, together with published data from PKC-delta <br>deficient mice that PKC-delta can terminate BCR signaling by binding to the BCR.","abstract_has_math":false,"creators":["Pracht, Catrin"],"institution":null,"degree_name":null,"degree_level":null,"degree_discipline":null,"degree_department":null,"school":null,"contributors":["Reth, Michael"],"advisors":[],"committee_chairs":[],"committee_members":[],"year":null,"date_issued":"","date_published":null,"updated_at":"2026-07-24T02:22:22Z","subjects":["Tyrosin Phosphorylierung","tyrosine phosphorylation","transducer complex"],"languages":[],"rights":[],"rights_urls":[],"identifier_entries":[]},"links":{"outbound_url":"https://freidok.uni-freiburg.de/data/1561","outbound_label":"Repository record","outbound_source":"source_url"},"metadata_groups":[{"id":"people","label":"People","entries":[{"key":"dc:contributor","label":"Contributor","values":["Reth, Michael"]},{"key":"dc:creator","label":"Author","values":["Pracht, Catrin"]}]},{"id":"academic_context","label":"Academic Context","entries":[{"key":"dc:type","label":"Dc Type","values":["DoctoralThesis"]}]},{"id":"subjects_keywords","label":"Subjects and Keywords","entries":[{"key":"dc:subject","label":"Dc Subject","values":["Tyrosin Phosphorylierung","tyrosine phosphorylation","transducer complex"]}]},{"id":"additional","label":"Additional Metadata","entries":[{"key":"dc:description.abstract","label":"Abstract","values":["The B cell antigen receptor (BCR) is composed of the membrane-bound immunoglobulin (mIg)
and the Ig-alpha/Ig-beta heterodimer and plays a central role in development, survival, and activation
of B lymphocytes. During B cell development, the BCR seems to be able to generate signals in
an antigen-dependent and -independent fashion. Upon binding to antigen, the BCR emits a
negative selection signal for immature and an activation signal for mature B cells. In the
absence of antigen, the BCR signals positive selection of immature and maintenance for mature
B cells. The BCR-dependent, antigen-independent signaling mode can be analyzed by
stimulating B cells with the phosphatase inhibitor pervanadate (PV). Analysis of substrate
tyrosine phosphorylation after PV-treatment of different B cell lines expressing BCR mutants
revealed drastic differences in phosphorylation of two signaling proteins namely the SH2-
domain-containing inositolpolyphosphate 5’-phosphatase (SHIP) and the protein kinase C delta.
Tyrosine phosphorylation of SHIP in response to PV stimulation was critically dependent on
the integrity of the Ig-alpha C-terminus. SHIP could be phosphorylated by both BCR-proximal
protein tyrosine kinases, Lyn and Syk. The phosphorylation of SHIP by Syk, but not that by
Lyn, was strictly dependent on the presence of an integer BCR complex, indicating that Syk
requires BCR expression for the phosphorylation of SHIP. Furthermore, the absence of proper
SHIP activation led to a disturbed balance of PKB and MAPK pathway activation, suggesting
that SHIP is a crucial regulator of the antigen-independent maintenance signal in B cells.
In contrast to SHIP, PKC-delta was able to bind in vitro to phosphorylated peptides derived from
the Ig-alpha C-terminus. As observed by native gel electrophoresis PKC-delta can be purified with the
BCR complex. The PKC-delta/BCR binding was dependent on the C-terminus of Ig-alpha. Moreover,
by confocal studies, PKC-delta was found to influence BCR/Syk localization, probably by inducing
BCR/Syk internalization. These data suggest, together with published data from PKC-delta
deficient mice that PKC-delta can terminate BCR signaling by binding to the BCR.","Der B-Zell-Antigen-Rezeptor (BZR) besteht aus dem Membran-gebundenen Immunglobulin (mIg) und dem Ig-alpha/Ig-beta Heterodimer und spielt eine zentrale Rolle in Entwicklung, Überleben und Aktivierung von B-Lymphozyten. Während der B-Zell-Entwicklung scheint der BZR Signale in einer Antigen-abhängigen und -unabhängigen Weise zu erzeugen. Nach Bindung des Antigens sendet der BZR ein negatives Selektionssignal für unreife und ein Aktivierungssignal für reife B-Zellen aus. In Abwesenheit des Antigens dagegen übermittelt der BZR ein positives Selektionssignal für unreife und ein Überlebenssignal für reife B-Zellen. Der B-Zell-abhängige, Antigen-unabhängige Signal-Modus kann durch Stimulation der B-Zellen mit dem Phosphatase-Inhibitor Pervanadat (PV) untersucht werden. Nachdem verschiedene B-Zell-Linien, die BZR-Mutanten exprimierten, mit PV behandelt worden waren, zeigte sich ein großer Unterschied in der Substrat-Tyrosin-Phosphorylierung von zwei Signalproteinen, der SH2-enthaltenden Inositolpolyphosphat 5'-Phosphatase (SHIP) und der Proteinkinase C-delta (PKC-delta).
Die Tyrosin-Phosphorylierung von SHIP, als Antwort auf die PV-Behandlung, war abhängig von der Integrität des C-Terminus von Ig-alpha. SHIP konnte durch beide BZR-nahen Kinasen, Lyn und Syk, phosphoryliert werden. Die Phosphorylierung von SHIP durch Syk, jedoch nicht die durch Lyn, war stark von der Anwesenheit des intakten BZR-Komplexes abhängig. Dies legt nahe, daß Syk die Expression des BZRs benötigt, um SHIP phosphorylieren zu können. Zudem, das Fehlen der SHIP-Aktivierung führte dazu, daß das Gleichgewicht der Aktivierung der PKB- und MAPK-Signalwege gestört war. Dies läßt vermuten, daß SHIP ein wichtiger Regulator für das Antigen-unabhängige Überlebenssignals in B-Zellen ist. Im Gegensatz zu SHIP konnte PKC-delta in vitro an phosphorylierte Peptide, die den C-Terminus von Ig-alpha repräsentierten, binden. Durch Verwendung der Methode der nativen Gel-Elektrophorese wurde beobachtet, daß PKC-delta mit dem BZR aufgereinigt werden kann. Die Bindung von PKC-delta an den BZR war abhängig von dem C-Terminus von Ig-alpha. Darüber hinaus wurde durch konfokale Studien herausgefunden, daß PKC-delta die Lokalisation von dem BZR/Syk-Komplex beeinflußt, wahrscheinlich duch Induktion der BZR/Syk-Internalisierung. Diese Daten lassen vermuten, daß PKC-delta die BZR-Signalleitung durch Bindung an den BZR beendet."]},{"key":"dc:format.medium","label":"Dc Format Medium","values":["application/pdf"]},{"key":"dc:title","label":"Title","values":["The analysis of B cell antigen receptor (BCR) mutants deficient in antigen-independent signaling reveals novel BCR-associated proteins","Die Entdeckung neuer B-Zell-Rezeptor (BZR)-assoziierter Proteine durch die Analyse von BZR-Mutanten, die Defekte in der Antigen-unabhängigen Signalleitung aufweisen"]}]}],"canonical_facts":{"dc:contributor":["Reth, Michael"],"dc:creator":["Pracht, Catrin"],"dc:description.abstract":["The B cell antigen receptor (BCR) is composed of the membrane-bound immunoglobulin (mIg)
and the Ig-alpha/Ig-beta heterodimer and plays a central role in development, survival, and activation
of B lymphocytes. During B cell development, the BCR seems to be able to generate signals in
an antigen-dependent and -independent fashion. Upon binding to antigen, the BCR emits a
negative selection signal for immature and an activation signal for mature B cells. In the
absence of antigen, the BCR signals positive selection of immature and maintenance for mature
B cells. The BCR-dependent, antigen-independent signaling mode can be analyzed by
stimulating B cells with the phosphatase inhibitor pervanadate (PV). Analysis of substrate
tyrosine phosphorylation after PV-treatment of different B cell lines expressing BCR mutants
revealed drastic differences in phosphorylation of two signaling proteins namely the SH2-
domain-containing inositolpolyphosphate 5’-phosphatase (SHIP) and the protein kinase C delta.
Tyrosine phosphorylation of SHIP in response to PV stimulation was critically dependent on
the integrity of the Ig-alpha C-terminus. SHIP could be phosphorylated by both BCR-proximal
protein tyrosine kinases, Lyn and Syk. The phosphorylation of SHIP by Syk, but not that by
Lyn, was strictly dependent on the presence of an integer BCR complex, indicating that Syk
requires BCR expression for the phosphorylation of SHIP. Furthermore, the absence of proper
SHIP activation led to a disturbed balance of PKB and MAPK pathway activation, suggesting
that SHIP is a crucial regulator of the antigen-independent maintenance signal in B cells.
In contrast to SHIP, PKC-delta was able to bind in vitro to phosphorylated peptides derived from
the Ig-alpha C-terminus. As observed by native gel electrophoresis PKC-delta can be purified with the
BCR complex. The PKC-delta/BCR binding was dependent on the C-terminus of Ig-alpha. Moreover,
by confocal studies, PKC-delta was found to influence BCR/Syk localization, probably by inducing
BCR/Syk internalization. These data suggest, together with published data from PKC-delta
deficient mice that PKC-delta can terminate BCR signaling by binding to the BCR.","Der B-Zell-Antigen-Rezeptor (BZR) besteht aus dem Membran-gebundenen Immunglobulin (mIg) und dem Ig-alpha/Ig-beta Heterodimer und spielt eine zentrale Rolle in Entwicklung, Überleben und Aktivierung von B-Lymphozyten. Während der B-Zell-Entwicklung scheint der BZR Signale in einer Antigen-abhängigen und -unabhängigen Weise zu erzeugen. Nach Bindung des Antigens sendet der BZR ein negatives Selektionssignal für unreife und ein Aktivierungssignal für reife B-Zellen aus. In Abwesenheit des Antigens dagegen übermittelt der BZR ein positives Selektionssignal für unreife und ein Überlebenssignal für reife B-Zellen. Der B-Zell-abhängige, Antigen-unabhängige Signal-Modus kann durch Stimulation der B-Zellen mit dem Phosphatase-Inhibitor Pervanadat (PV) untersucht werden. Nachdem verschiedene B-Zell-Linien, die BZR-Mutanten exprimierten, mit PV behandelt worden waren, zeigte sich ein großer Unterschied in der Substrat-Tyrosin-Phosphorylierung von zwei Signalproteinen, der SH2-enthaltenden Inositolpolyphosphat 5'-Phosphatase (SHIP) und der Proteinkinase C-delta (PKC-delta).
Die Tyrosin-Phosphorylierung von SHIP, als Antwort auf die PV-Behandlung, war abhängig von der Integrität des C-Terminus von Ig-alpha. SHIP konnte durch beide BZR-nahen Kinasen, Lyn und Syk, phosphoryliert werden. Die Phosphorylierung von SHIP durch Syk, jedoch nicht die durch Lyn, war stark von der Anwesenheit des intakten BZR-Komplexes abhängig. Dies legt nahe, daß Syk die Expression des BZRs benötigt, um SHIP phosphorylieren zu können. Zudem, das Fehlen der SHIP-Aktivierung führte dazu, daß das Gleichgewicht der Aktivierung der PKB- und MAPK-Signalwege gestört war. Dies läßt vermuten, daß SHIP ein wichtiger Regulator für das Antigen-unabhängige Überlebenssignals in B-Zellen ist. Im Gegensatz zu SHIP konnte PKC-delta in vitro an phosphorylierte Peptide, die den C-Terminus von Ig-alpha repräsentierten, binden. Durch Verwendung der Methode der nativen Gel-Elektrophorese wurde beobachtet, daß PKC-delta mit dem BZR aufgereinigt werden kann. Die Bindung von PKC-delta an den BZR war abhängig von dem C-Terminus von Ig-alpha. Darüber hinaus wurde durch konfokale Studien herausgefunden, daß PKC-delta die Lokalisation von dem BZR/Syk-Komplex beeinflußt, wahrscheinlich duch Induktion der BZR/Syk-Internalisierung. Diese Daten lassen vermuten, daß PKC-delta die BZR-Signalleitung durch Bindung an den BZR beendet."],"dc:format.medium":["application/pdf"],"dc:subject":["Tyrosin Phosphorylierung","tyrosine phosphorylation","transducer complex"],"dc:title":["The analysis of B cell antigen receptor (BCR) mutants deficient in antigen-independent signaling reveals novel BCR-associated proteins","Die Entdeckung neuer B-Zell-Rezeptor (BZR)-assoziierter Proteine durch die Analyse von BZR-Mutanten, die Defekte in der Antigen-unabhängigen Signalleitung aufweisen"],"dc:type":["DoctoralThesis"]},"updated_at":"2026-07-24T02:22:22Z"}