Cloning of a novel T-cell protein FYB that binds FYN and SH2-domain-containing leukocyte protein 76 and modulates interleukin 2 production

Abstract

T cell receptor (TcR)͞CD3 ligation initiates a signaling cascade that involves src kinases p56 lck and -associated protein 70, leading to the phosphorylation of substrates such as TcR, Vav, SH2-domain-containing leukocyte protein 76 (SLP-76), cbl, and p120͞130. FYN binding protein (FYB or p120͞130) associates with p59 fyn , the TcR͞ CD3 complex, and becomes tyrosine-phosphorylated in response to receptor ligation. In this study, we report the cDNA cloning of human and murine FYB and show that it is restricted in expression to T cells and myeloid cells and possesses an overall unique hydrophilic sequence with several tyrosine-based motifs, proline-based type I and type II SH3 domain binding motifs, several putative lysine͞glutamic acidrich nuclear localization motifs, and a SH3-like domain. In addition to binding the src kinase p59 fyn , FYB binds specifically to the hematopoietic signaling protein SLP-76, an interaction mediated by the SLP-76 SH2 domain. In keeping with this, expression of FYB augmented interleukin 2 secretion from a T cell hybridoma, DC27.10, in response to TcR͞CD3 ligation. FYB is therefore a novel hematopoietic protein that acts as a component of the FYN and SLP-76 signaling cascades in T cells.

ARAP, a Novel Adaptor Protein, Is Required for TCR Signaling and Integrin-Mediated Adhesion

“…These results may suggest that ARAP participates with Y491 phosphorylation in the proximal signaling event by other mechanisms in addition to SLP-76 binding. The intervening phosphorylation site Y521 in ARAP may have a different tyrosine consensus sequence, perhaps more similar to the YDGI in ADAP that was previously identified to mediate the association of ADAP with the Src kinase Fyn (43). This might explain why Y521 was essentially irrelevant for SLP-76 binding in the present studies.…”

Adapter proteins SLP-76 and BLNK both are expressed by murine macrophages and are linked to signaling via Fcγ receptors I and II/III

“…SLP-76 [Src homology 2 (SH2) domain-containing leukocyte protein of 76 kDa] (5) is an adapter molecule crucial in TCR signaling. SLP-76 has numerous direct and indirect associations, including Grb2 (5), the Grb2-related adapter downstream of Shc (Gads) that links SLP-76 to LAT (linker for activation of T cells) (6), phospholipase C (PLC)-␥ (7), Vav (8), and the Fyn binding protein (9), also known as SLP-76-associated protein of 130 kDa (SLAP-130) (10), the SH2 domain-containing phosphatase-1 (11), and Nck (12). PLC-␥ activation leads to release of intracellular calcium and stimulation of calcium-dependent pathways.…”

In vivo disruption of T cell development by expression of a dominant‐negative polypeptide designed to abolish the SLP‐76/Gads interaction

“…The N terminus harbors three tyrosines that become phos- phorylated upon TCR engagement [5,6] and have been reported to serve as docking sites for the guanine nucleotide exchange factor Vav, the adaptor protein Nck, the Tec family kinase Itk, and the p85 subunit of PI3K [7][8][9][10][11][12][13][14][15][16]. A single SH2 domain resides in the C-terminal portion of SLP-76 and links it to the adhesion and degranulation-promoting adaptor protein (ADAP) and to the hematopoietic progenitor kinase 1 [17][18][19]. Hematopoietic progenitor kinase 1, in turn, has been shown to phosphorylate SLP-76, creating a binding site for 14-3-3, a negative regulator of TCR signaling [20,21].…”