Abstract
BackgroundThe ZAS family is composed of proteins that regulate transcription via specific gene regulatory elements. The amino-DNA binding domain (ZAS-N) and the carboxyl-DNA binding domain (ZAS-C) of a representative family member, named κB DNA binding and recognition component (KRC), were expressed as fusion proteins and their target DNA sequences were elucidated by site selection amplification binding assays, followed by cloning and DNA sequencing. The fusion proteins-selected DNA sequences were analyzed by the MEME and MAST computer programs to obtain consensus motifs and DNA elements bound by the ZAS domains.ResultsBoth fusion proteins selected sequences that were similar to the κB motif or the canonical elements of the V(D)J recombination signal sequences (RSS) from a pool of degenerate oligonucleotides. Specifically, the ZAS-N domain selected sequences similar to the canonical RSS nonamer, while ZAS-C domain selected sequences similar to the canonical RSS heptamer. In addition, both KRC fusion proteins selected oligonucleoties with sequences identical to heptamer and nonamer sequences within endogenous RSS.ConclusionsThe RSS are cis-acting DNA motifs which are essential for V(D)J recombination of antigen receptor genes. Due to its specific binding affinity for RSS and κB-like transcription enhancer motifs, we hypothesize that KRC may be involved in the regulation of V(D)J recombination.
Highlights
The ZAS family is composed of proteins that regulate transcription via specific gene regulatory elements
Amplification of KRC's DNA targets with a site selection amplification binding assay In this study, sequences bound by the DNA binding domains of KRC were identified in a site selection PCR amplification DNA binding assay
Because the pentameric motifs derived from the KRC/ZAS-N dataset more closely resemble the canonical recombination signal sequences (RSS) nonamer and the motif derived from the KRC/ZAS-C dataset more closely resemble the canonical RSS heptamer and a canonical sequence was derived from the majority of sequences, we propose that the ZAS-N domain of KRC binds RSS nonamers more frequently than the ZAS-C domain, and vice versa for the RSS heptamer in vivo
Summary
The ZAS family is composed of proteins that regulate transcription via specific gene regulatory elements. Each ZAS gene encodes large sequence-specific DNAbinding proteins with Mr >250,000 that contain two widely separated of C2H2-type zinc finger pairs. The amino acid sequence and relative location of the two zinc finger pairs are highly conserved among ZAS proteins from invertebrate to vertebrate [Reviewed in [25]]. The zinc finger is a major structural motif involved in protein-nucleic acid interactions and is present in the largest superfamily of transcription factors, few proteins contain separate zinc finger pairs. Each ZAS protein contains a sequence similar to the serine stripe present in basonuclin, in which eight serines are located on one side of a putative α-helix [13,27]
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