The dataset of proteins specifically interacted with activated TICAM-1.

The presented data are related with our paper entitled "14-3-3-zeta participates in TLR3-mediated TICAM-1 signal-platform formation" (Funami et al., 2016) [1]. These data show the proteins which specifically bind to the activated (oligomerized) TICAM-1. Fifty-three proteins were identified as specifically interacted with oligomerized TICAM-1. Mutant TICAM-1 cannot form the active oligomer, so the proteins interacted with mutant TICAM-1 are dispensable for TICAM-1-signaling. Among 53 proteins, 14-3-3-zeta specifically interacts with oligomerized TICAM-1 to corroborate TICAM-1 signalosome.

Specifications Table

Value of the data

This data shows the components of TICAM-1 signalosome, which are important for the regulation of TICAM-1 functions.

This data first distinguishes a ‘functional binding’ to the activated TICAM-1 from ‘off-target binding’ to a non-functional mutant, TICAM-1-N+TIR-P434H.

This data shows that several types of 14-3-3 proteins are identified as TICAM-1-binding proteins. In addition to TICAM-1-signalosome formation we identified, 14-3-3 proteins may have other functions in the field of innate immunity.

Data

We show the strategy for identification of TICAM-1-signalosome component in Fig. 1. By mass spectrometry, we identify the proteins interacted with functional TICAM-1 and non-functional TICAM-1 mutant, and represent the list of the proteins in Table 1.

Fig. 1

Fetching TICAM-1-binding protein using LC-MS-MS analysis.

Experimental design, materials and methods

Full-length TICAM-1 and TICAM-1-N+TIR-P434H were expressed in HEK293 cells and TICAM-1-binding proteins were immunoprecipitated by Streptavidin Sepharose (GE healthcare) [1]. Eluted samples were separated in a 10% SDS-polyacrylamide gel for liquid chromatography coupled to tandem mass-spectrometry (LC/MS/MS). The raw data files obtained from the LC/MS/MS were analyzed as described previously, with minor modifications [2], [3]. Our scheme was depicted in Fig. 1.

The TICAM-1 construct having Tags (streptavidin-binding peptide (SBP) and calmodulin binding peptides (CBP)) and that lacking the RHIM domain with a mutated TIR domain were provided as reported previously [4]. The latter fails to oligomerize to activate RIP1 kinase in transfected cells. TICAM-1-signalosome-binding proteins were obtained by subtraction using proteome analyses (LC/MS/MS). NTD, N-terminal domain; TIR, toll-IL-1β-homology domain.

Subject area	Biology
More specific subject area	Innate immunity
Type of data	Table, figure
How data was acquired	Mass spectrometry analysis
Data format	Analyzed data in excel file
Experimental factors	TICAM-1 binding proteins were co-immunoprecipitated from HEK293 cells transfected with full-length TICAM-1 and non-specific binding was subtracted.
Experimental features	Immunoprecipitated samples were separated by SDS-PAGE and subjected to LC/MS/MS analysis.
Data source location	Sapporo, Hokkaido, Japan
Data accessibility	All data are accessible in this article.