Chromatin modulation at various cis-acting elements is critical for V(D)J 
recombination during T and B cell development. MARbeta, a matrix-associated 
region (MAR) located upstream of the T cell receptor beta (TCRbeta) enhancer 
(Ebeta), serves a crucial role in silencing Ebeta-mediated TCR activation. By 
DNaseI hypersensitivity assays, we show here that overexpression of the MAR 
binding proteins SMAR1 and Cux/CDP modulate the chromatin structure at MARbeta. 
We further demonstrate that the silencer function of MARbeta is mediated 
independently by SMAR1 and Cux/CDP as judged by their ability to repress 
Ebeta-dependent reporter gene expression. Moreover, the repressor activity of 
SMAR1 is strongly enhanced in the presence of Cux/CDP. These two proteins 
physically interact with each other and colocalize within the perinuclear region 
through a SMAR1 domain required for repression. The repression domain of SMAR1 
is separate from the MARbeta binding domain and contains a nuclear localization 
signal and an arginine-serine (RS)-rich domain, characteristic of pre-mRNA 
splicing regulators. Our data suggest that at the double positive stage of T 
cell development, cis-acting MARbeta elements recruit the strong negative 
regulators Cux and SMAR1 to control Ebeta-mediated recombination and 
transcription.