Differential protein–protein interactions of full length human FasL and FasL fragments generated by proteolysis

Fas ligand (FasL) is a death factor of the tumor necrosis factor superfamily. Like other members of this family of type II transmembrane proteins, FasL is subject to ectodomain shedding by a disintegrin and metalloproteinases (ADAMs) liberating soluble FasL and leaving membrane-integral N-terminal f...

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Published in:Experimental cell research Vol. 320; no. 2; pp. 290 - 301
Main Authors: Lettau, Marcus, Voss, Matthias, Ebsen, Henriette, Kabelitz, Dieter, Janssen, Ottmar
Format: Journal Article
Language:English
Published: United States Elsevier Inc 15.01.2014
Elsevier BV
Subjects:
Animals
Biomedical research
Cells, Cultured
Fas Ligand Protein - chemistry
Fas Ligand Protein - metabolism
FasL
HEK293 Cells
Humans
Intramembrane proteolysis
Jurkat Cells
K562 Cells
Mice
Mice, Inbred BALB C
Monoclonal antibody
PCH proteins
Peptide Fragments - chemistry
Peptide Fragments - metabolism
Protein Binding - physiology
Protein expression
Protein Interaction Domains and Motifs - physiology
Protein Interaction Mapping
Protein–protein interaction
Proteolysis
Proteomics
Signal transduction
Sorting nexins
Studies
Substrate Specificity
T lymphocytes
Tumors
T lymphocytes
NTF
PCH proteins
(F-)BAR
PRD
ESCRT
SPPL2a
ICD
Monoclonal antibody
TNF
Intramembrane proteolysis
Protein–protein interaction
mab
ADAM
Signal transduction
Sorting nexins
PCH
FasL
Proteolysis
RIP
SNX
I-CLiP
SH3
Fas ligand
(Fer/CIP4 homology-)Bin–Amphiphysin–Rvs
Pombe Cdc15 homology
intracellular domain
endosomal sorting complexes required for transport
intramembrane-cleaving protease
proline-rich domain
Src homology 3
regulated intramembrane proteolysis
sorting nexin
N-terminal fragment
a disintegrin and metalloproteinase
signal peptide peptidase like 2a
tumor necrosis factor
ISSN:0014-4827, 1090-2422, 1090-2422
Online Access:Get full text
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Summary:Fas ligand (FasL) is a death factor of the tumor necrosis factor superfamily. Like other members of this family of type II transmembrane proteins, FasL is subject to ectodomain shedding by a disintegrin and metalloproteinases (ADAMs) liberating soluble FasL and leaving membrane-integral N-terminal fragments (NTFs). These NTFs are further processed by intramembrane proteolysis through signal peptide peptidase-like 2a (SPPL2a), releasing intracellular domains (ICDs) which might translocate to the nucleus to regulate transcription. Previous work established that the proline-rich domain within the cytosolic N-terminus of FasL is required for protein–protein interactions with different Src homology 3 (SH3) or WW domain proteins. Distinct binding partners regulate FasL storage and surface appearance or are involved in other aspects of FasL biology. Given the large number of FasL interactors, we asked whether proteolytically processed FasL fragments associate with the same or distinct sets of SH3 domain proteins. To address this, we performed co-precipitation experiments using a monoclonal antibody directed against the FasL N-terminus for subsequent protein detection of full length FasL and NTFs/ICDs in Western blots. We demonstrate that members of the sorting nexin (SNX) family bind full length FasL and its N-terminal fragments whereas members of the Pombe Cdc15 homology (PCH) protein family bind full length FasL, but fail to associate with processed FasL. Thus, we provide first evidence that full length FasL and FasL fragments display selectivity regarding their association with intracellular binding partners. The differential binding most likely governs the fate and function of the intracellular FasL fragments. •Proteolytic processing of FasL can be visualized using the newly established mab Fc2.•FasL fragments are constitutively produced in transfectants and inducible in T cells.•PCH proteins and sorting nexins were analyzed as known FasL interactors.•Sorting nexins co-precipitate full length FasL and its N-terminal fragments.•PCH proteins bind full length FasL but do not co-precipitate N-terminal fragments.
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ISSN:0014-4827
1090-2422
1090-2422
DOI:10.1016/j.yexcr.2013.11.016