Human DNA Replication-related Element Binding Factor (hDREF) Self-association via hATC Domain Is Necessary for Its Nuclear Accumulation and DNA Binding

Abstract

We previously demonstrated that hDREF, a human homologue of Drosophila DNA replication-related element binding factor (dDREF), is a DNA-binding protein predominantly distributed with granular structures in the nucleus. Here, glutathione S-transferase pulldown and chemical cross-linking assays showed that the carboxyl-terminal hATC domain of hDREF, highly conserved among hAT transposase family members, possesses self-association activity. Immunoprecipitation analyses demonstrated that hDREF self-associates in vivo, dependent on hATC domain. Moreover, analyses using a series of hDREF mutants carrying amino acid substitutions in the hATC domain revealed that conserved hydrophobic amino acids are essential for self-association. Immunofluorescence studies further showed that all hDREF mutants lacking self-association activity failed to accumulate in the nucleus. Self-associationdefective hDREF mutants also lost association with endogenous importin ␤1. Moreover, electrophoretic gel-mobility shift assays revealed that the mutations completely abolished the DNA binding activity of hDREF. These results suggest that selfassociation of hDREF via the hATC domain is necessary for its nuclear accumulation and DNA binding. We also found that ZBED4/KIAA0637, another member of the human hAT family, also self-associates, again dependent on the hATC domain, with deletion resulting in loss of efficient nuclear accumulation. Thus, hATC domains of human hAT family members appear to have conserved functions in self-association that are required for nuclear accumulation.

A Transcriptome Database for Astrocytes, Neurons, and Oligodendrocytes: A New Resource for Understanding Brain Development and Function

“…The exceptions to this trend included the two well established OPC markers, PDGFR␣ and NG2/Cspg4, but also some less well characterized genes such as Matrilin-4 (Matn4), Ligand-of-Numb protein X (Lnx1), protein kinase cGMP-dependent (Prkg2), and zinc finger BED domain containing 4 (Zbed4). Zbed4 is a member of the human hAT family that self-associates and acts as a transcriptional activator of cell cycle genes (Yamashita et al, 2007). G-protein-coupled receptor 17 (Gpr17) is strongly and specifically expressed by developing OLs and by adulthood appears specific to adult OPCs.…”

Molecular domestication of transposable elements: From detrimental parasites to useful host genes

“…DREF factors and hAT transposases share a Cterminal hATC (hAT C-terminal dimerization) domain that has been found to be a dimerization domain of Activator and Hermes transposases [81,82]. Similarly, the hATC domain is necessary for hDREF selfassociation in vivo, and is also required for nuclear accumulation, DNA-binding activity and granular pattern formation [83].…”

DAYSLEEPER: a nuclear and vesicular-localized protein that is expressed in proliferating tissues

“…The NLS is followed by a BED-type zinc finger and 6 identifiable hAT blocks (A to F), but lacks the amino acids essential for mobility [8,9]. hAT Blocks D, E and F make up a hAT dimerization domain [10,11]. These hAT blocks are defining characteristics of hAT transposases in all species, although not all transposases possess all six blocks [10].…”