Nucleic‐acid‐binding properties of hnRNP‐U/SAF‐A, a nuclear‐matrix protein which binds DNA and RNA in vivo and in vitro

Abstract

We show that SAF-A, a nuclear protein which specifically binds vertebrate scaffold-attachmentregion (SAR) elements with high affinity is identical with hnRNP-U, assumed to be involved in packaging of hnRNA in ribonucleoprotein particles. Ultraviolet cross-linking experiments show that the protein, referred to as hnRNP-U/SAF-A, is bound to chromosomal DNA in vivo. Zn vitro, the isolated protein binds to double-stranded and single-stranded DNA and forms higher ordered nucleic-acid -protein complexes. Filter-binding experiments performed with different types of natural and synthetic nucleic acids as substrates show that the protein binds DNA and RNA with different affinities and most likely at different binding sites. We conclude that hnRNP-U/SAF-A thus may have functions in the organisation of chromosomal DNA in addition to its suggested role in hnRNA metabolism.It is widely believed that DNA in eukaryotic interphase nuclei is organized into loop domains with average lengths of 50-100 kb and that these chromatin loops are bound to internal nuclear structures via the scaffold-attachment region (SAR; for review see [l]) or the matrix-associated region (MAR; for review see 12, 31) DNA elements. Many DNA fragments which specifically bind to internal nuclear structures have been identified, but only little is known about their function in vivo and how they exert this function. In addition, only a few proteins ([4, 51 and references therein) are known which possibly are involved in the binding of these DNA fragments to internal nuclear structures.We have therefore attempted to analyse for nuclear proteins which specifically bind SAR DNA elements and we reported recently on the purification of a DNA binding protein with this property 161. This protein, SAF-A (scaffoldattachment factor A), is a component of the nuclear matrix and of scaffolds; it specifically binds vertebrate SAR DNA elements and reconstitutes looped structures with these DNAs [6]. SAF-A is an abundant protein and the large number of molecules presenthucleus indicate a structural function of this protein. In this communication, we identify SAF-A as hnRNP-U [7], a protein component of heterogeneous nuclear ribonucleoprotein complexes (for review see [S-lo]). We refer to this protein as hnRNP-U/SAF-A and characterize the nucleic-acid binding properties of the protein. We further show by ultraviolet (UV) cross-linking experiments that the protein is bound to DNA in vivo and disCorrespondence to A. Richter,

hnRNP U Enhances Caspase-9 Splicing and Is Modulated by AKT-dependent Phosphorylation of hnRNP L

“…Previously, hnRNP U has been reported to preferentially bind G/U-rich motifs in RNA (9,10). In line with these reports, hnRNP U was demonstrated to specifically interact with the G-rich sequence, GAGGG, in C9/E3.…”

“…hnRNP U, also known as SP120 or scaffold attachment factor A, is capable of binding to both DNA and RNA through the acidic region in the N terminus or the RGG box in the C terminus (9,10). hnRNP U has been reported to have various biological functions from regulation of gene transcription (11)(12)(13)(14) to controlling RNA stability (15) and even participating in the X inactivation process (16,17) and telomere length control (18).…”

Mouse Geminin Inhibits Not Only Cdt1-MCM6 Interactions but Also a Novel Intrinsic Cdt1 DNA Binding Activity

“…However, mCdt1 does not bind to yeast transfer RNA or to DNA that is treated with high concentrations of ethidium bromide (36), suggesting that mCdt1 specifically recognizes DNA (data not shown). This binding profile is reminiscent of that of members of the HMG protein family (37), the RNP-U protein (38), and some initiator proteins such as DnaA, O (39), or yeast ORC (40,41). HMG proteins and the RNP-U protein can associate with a wide range of DNA structures and are considered to modulate chromatin structure (37,38).…”

Domain Structure of Human Nuclear DNA Helicase II (RNA Helicase A)

“…In the case of hnRNP U, the RGG domain is the only nucleic acid binding domain; nevertheless, it mediates binding to ssDNA even in the presence at 0.5-1 M NaCl (14). Such a high affinity for ssDNA may be a prerequisite for the observed binding of hnRNP U to nuclear matrix elements (SAR/MAR) (41,42); it may also help to unwind SAR/MAR elements (43). Similar to hnRNP U, the RGG-box of human NDH II bound to ssDNAagarose at salt concentrations of more than 0.5 M NaCl.…”