A novel DNA-binding motif in the nuclear matrix attachment DNA-binding protein SATB1.

Abstract

The nuclear matrix attachment DNA (MAR) binding protein SATBI is a sequence context-specific binding protein that binds in the minor groove, making virtually no contact with the DNA bases. The SATB1 binding sites consist of a special AT-rich sequence context in which one strand is well-mixed A's, T's, and C's, excluding G's (ATC sequences), which is typically found in clusters within different MARs. To determine the extent of conservation of the SATB1 gene among different species, we cloned a mouse homolog of the human SATB1cDNA from a cDNA expression library of the mouse thymus, the tissue in which this protein is predominantly expressed. This mouse cDNA encodes a 764-amino-acid protein with a 98% homology in amino acid sequence to the human SATB1 originally cloned from testis. To characterize the DNA binding domain of this novel class of protein, we used the mouse SATBI cDNA and delineated a 150-amino-acid polypeptide as the binding domain. This region confers full DNA binding activity, recognizes the specific sequence context, and makes direct contact with DNA at the same nucleotides as the whole protein. This DNA binding domain contains a novel DNA binding motif: when no more than 21 amino acids at either the N-or C-terminal end of the binding domain are deleted, the majority of the DNA binding activity is lost. The concomitant presence of both terminal sequences is mandatory for binding. These two terminal regions consist of hydrophilic amino acids and share homologous sequences that are different from those of any known DNA binding motifs. We propose that the DNA binding region of SATB1 extends its two terminal regions toward DNA to make direct contact with DNA.

Interaction of the Nuclear Matrix-associated Region (MAR)-Binding Proteins, SATB1 and CDP/Cux, with a MAR Element (L2a) in an Upstream Regulatory Region of the Mouse CD8a Gene

“…5B). The size is consistent with that reported for mSATB1, and SATB1 has been reported to be phosphorylated (47). The failure to observe a corresponding band precipitated from A11 extract radiolabeled with either isotope may reflect the very low amount of SATB1 protein expressed.…”

“…2B). Although the SATB1 cDNA encodes a protein of molecular mass of 87 kDa, it is reported to migrate on SDS-PAGE as several closely spaced components of molecular mass of approximately 103 kDa, possibly due to phosphorylation or other post-translational modifications (47). This is comparable with the M r observed for the L2a-P1 protein (19).…”

“…Though only a very small amount of purified protein was obtained, the partial amino acid sequence of a peptide obtained from two incompletely resolved bands of fraction 54 showed significant homology in a data base search to SATB1, a protein whose cDNA was originally cloned from a testis cDNA library but that is reported to be expressed only in thymus (15,47) (Fig. 2B).…”

The Mouse Immunoglobulin Heavy Chain V-D Intergenic Sequence Contains Insulators That May Regulate Ordered V(D)J Recombination

“…Thus, HS4 and HS5 are strategically placed at the interface between two opposing histone modifications, a characteristic feature of insulators (39). Third, we have identified a 71-bp putative scaffold/matrix attachment region element 300 bp upstream of HS4, with 13 predicted SATB (special AT-rich sequence-binding protein) binding sites (52). Thus, HS4 may be associated with the nuclear matrix through SATB1/SATB2 interactions, which could stabilize interaction with other cis-acting elements.…”

Mapping the initial DNA breaks in apoptotic Jurkat cells using ligation-mediated PCR

“…SATB1 in its monomeric form is unable to bind to DNA. 29,33 Another MAR-binding protein, SAF-A, is also cleaved during apoptosis and loses its DNA-binding capacity, although in this case the cleavage occurs within the DNA-binding domain. 34 There have been a number of reports in the literature suggesting that initiation of apoptotic DNA cleavage occurs at matrix attachment regions.…”