TOM1L1 is a Lyn substrate involved in FcepsilonRI signaling in mast cells.
Protein-tyrosine kinase Lyn and Syk are critical for antigen-receptor-induced signal transduction in mast cells. To identify novel Lyn/Syk substrates, we screened an RBL-2H3 bacterial expression library for proteins that were tyrosine phosphorylated with baculoviral expressed Lyn or Syk. Five clones as potential Lyn substrates and eight clones as Syk substrates were identified including known substrates such as SLP-76, LAT, and alpha-tubulin. A potential substrate of Lyn identified was the molecule TOM1L1, which has several domains thought to be important for membrane trafficking and protein-protein interactions. Because the function of TOM1L1 is unclear, the rat TOM1L1 full-length cDNA was isolated and used to express the protein in COS-1 and RBL-2H3 mast cells. In COS-1 cells, the co-transfection of TOM1L1 and Lyn, but not Syk, resulted in the tyrosine phosphorylation of TOM1L1. In RBL-2H3 mast cells, the overexpressed TOM1L1 was strongly tyrosine phosphorylated in non-stimulated cells, and this phosphorylation was enhanced by FcepsilonRI aggregation. By subcellular fractionation, wild-type TOM1L1 was mainly in the cytoplasm with a small fraction constitutively associated with the membrane; this association was markedly reduced in deletion mutants lacking several of the protein interaction domains. The overexpression of TOM1L1 enhanced antigen-induced tumor necrosis factor (TNF) alpha generation and release. Both protein interaction domains (VHS and the coiled-coil domains) were required for the increased TNFalpha release, but not the increased TNFalpha generation. These results suggest that TOM1L1 is a novel protein involved in the FcepsilonRI signal transduction for the generation of cytokines.
Zhang
Juan
J
Receptors and Signal Transduction Section, Oral Infection and Immunity Branch, NIDCR, National Institutes of Health, Bethesda, MD 20892, USA.
Suzuki
Katsuhiro
K
Hitomi
Tomohiro
T
Siraganian
Reuben P
RP
D048868:Adaptor Proteins, Signal Transducing / Q000737:chemistry / Q000502:physiology*; D000818:Animals; D019556:COS Cells; D045744:Cell Line, Tumor; D002522:Chlorocebus aethiops; D016207:Cytokines / Q000378:metabolism; D005786:Gene Expression Regulation; D008407:Mast Cells / Q000378:metabolism*; D010766:Phosphorylation; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D051381:Rats; D017455:Receptors, IgE / Q000378:metabolism*; D015398:Signal Transduction; D019061:src-Family Kinases / Q000737:chemistry*
D048868:Adaptor Proteins, Signal Transducing; D016207:Cytokines; D017455:Receptors, IgE; C060902:lyn protein-tyrosine kinase; D019061:src-Family Kinases