Kit- and Fc epsilonRI-induced differential phosphorylation of the transmembrane adaptor molecule NTAL/LAB/LAT2 allows flexibility in its scaffolding function in mast cells.
The transmembrane adaptor protein (TRAP), NTAL, is phosphorylated in mast cells following FcvarepsilonRI aggregation whereby it cooperates with LAT to induce degranulation. The Kit ligand, stem cell factor (SCF), enhances antigen-induced degranulation and this also appears to be NTAL-dependent. However, Kit and FcvarepsilonRI appear to utilize different mechanisms to induce NTAL phosphorylation. Thus, we examined whether the responsible kinases selectively phosphorylated distinct tyrosines in NTAL and explored the implications for downstream signaling. Whereas FcvarepsilonRI required Lyn and Syk for NTAL phosphorylation, Kit appeared to directly phosphorylate NTAL. Furthermore, co-transfection studies with NTAL constructs revealed that Lyn, Syk, and Kit phosphorylate different tyrosines in NTAL. The tyrosines principally phosphorylated by Syk were recognized as Grb2-binding sites, whereas Lyn and Kit phosphorylated other tyrosines, both inside and outside of these motifs. Pull down studies revealed that PLCgamma1 associated with the two terminal Syk-phosphorylated Grb2-binding sites, which would help to explain the observed decrease in antigen-induced calcium signal and degranulation in NTAL-knock down-human mast cells. The observations reported herein support the conclusion that NTAL may be differentially utilized by specific receptors for relaying alternative signals and this suggests a flexibility in the function of TRAPs not previously appreciated.
Iwaki
Shoko
S
Laboratory of Allergic Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, 10 Center Drive, MSC 1881, Bethesda, MD 20892-1881, USA.
Spicka
Jiri
J
Tkaczyk
Christine
C
Jensen
Bettina M
BM
Furumoto
Yasuko
Y
Charles
Nicolas
N
Kovarova
Martina
M
Rivera
Juan
J
Horejsi
Vaclav
V
Metcalfe
Dean D
DD
Gilfillan
Alasdair M
AM
D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism*; D000818:Animals; D002478:Cells, Cultured; D003599:Cytoskeleton / Q000378:metabolism*; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000235:genetics / Q000378:metabolism; D008407:Mast Cells / Q000378:metabolism*; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D051966:Phospholipase C gamma / Q000378:metabolism; D010766:Phosphorylation; D011505:Protein-Tyrosine Kinases / Q000235:genetics / Q000378:metabolism; D017455:Receptors, IgE / Q000378:metabolism*; D019089:Stem Cell Factor / Q000502:physiology*; D000072377:Syk Kinase; D014162:Transfection; D019061:src-Family Kinases / Q000235:genetics / Q000378:metabolism
D048868:Adaptor Proteins, Signal Transducing; D047908:Intracellular Signaling Peptides and Proteins; C419196:LAT2 protein, human; D017455:Receptors, IgE; D019089:Stem Cell Factor; D011505:Protein-Tyrosine Kinases; C000606896:SYK protein, human; D000072377:Syk Kinase; C000606897:Syk protein, mouse; C060902:lyn protein-tyrosine kinase; D019061:src-Family Kinases; D051966:Phospholipase C gamma