Histones and DNA Compete for Binding Polyphosphoinositides in Bilayers

Abstract

Recent discoveries on the presence and location of phosphoinositides in the eukaryotic cell nucleoplasm and nuclear membrane prompted us to study the putative interaction of chromatin components with these lipids in model membranes (liposomes). Turbidimetric studies revealed that a variety of histones and histone combinations (H1, H2AH2B, H3H4, octamers) caused a dose-dependent aggregation of phosphatidylcholine vesicles (large unilamellar vesicle or small unilamellar vesicle) containing negatively charged phospholipids. 5 mol % phosphatidylinositol-4-phosphate (PIP) was enough to cause extensive aggregation under our conditions, whereas with phosphatidylinositol (PI) at least 20 mol % was necessary to obtain a similar effect. Histone binding to giant unilamellar vesicle and vesicle aggregation was visualized by confocal microscopy. Histone did not cause vesicle aggregation in the presence of DNA, and the latter was able to disassemble the histone-vesicle aggregates. At DNA/H1 weight ratios 0.1-0.5 DNA- and PIP-bound H1 appear to coexist. Isothermal calorimetry studies revealed that the PIP-H1 association constant was one order of magnitude higher than that of PI-H1, and the corresponding lipid/histone stoichiometries were ~0.5 and ~1, respectively. The results suggest that, in the nucleoplasm, a complex interplay of histones, DNA, and phosphoinositides may be taking place, particularly at the nucleoplasmic reticula that reach deep within the nucleoplasm, or during somatic and nonsomatic nuclear envelope assembly. The data described here provide a minimal model for analyzing and understanding the mechanism of these interactions.

Biology, role and therapeutic potential of circulating histones in acute inflammatory disorders

“…Positively charged histones preferentially bind anionic phospholipids such as cardiolipin or phosphatidylserine, but not zwitterionic phospholipids like phosphatidylcholine 132. Furthermore, adding negative charge (eg, a phosphate head group as in phosphatidylinositol bis‐phosphate) increases the binding capacity of histones as measured by calorimetry 128. Histones have also been shown to expose phosphatidylserine on the surface of red blood cells in a dose‐dependent manner47; however, it is unclear whether this is as a result of altering flippase kinetics or via induction of apoptosis pathways.…”

“…Once integrated, histones induce permeabilization of membranes to cations, disruptions of cellular calcium signalling112 and cell death by necrosis. Negatively charged acute‐phase proteins (such as C‐reactive protein, CRP),14 DNA,128 innate polysaccharides (heparin)120 or synthetic macromolecules126 compete with membrane phospholipids and prevent histone integration and toxicity. Bactericidal properties of histone fragments are dependent on their ability to form amphipathic α‐helices—potentially membrane spanning domains—however no such structural analyses have been performed on mammalian cells to date 89…”

Histones Cause Aggregation and Fusion of Lipid Vesicles Containing Phosphatidylinositol-4-Phosphate

“…1, 2, 3, 4, and 5) demonstrate that histones cause effects that in combination are diagnostic for membrane fusion. In our previous study (13), we showed that PIP could compete with DNA for histones. Together, our previous and current data suggest a complex multiple equilibrium in which free and bound DNA, free and bound histones (specifically H1) (44) and membranes, and nonfused and fused vesicles may coexist in the close vicinity of the nucleoplasmic reticulum.…”

“…Isothermal calorimetry data for the lipidprotein interaction were compatible with phosphoinositide/ histone stoichiometries of~0.5 (PIP) and~1.0 (PI), respectively. Moreover, competition experiments involving histones and negatively charged vesicles and DNA demonstrated the possibility of coexisting DNA-and PIP-bound histones (13).…”

“…In fact, several authors (10)(11)(12) have studied the interaction of histones with phosphatidylserine and other negatively charged lipids that exist in very minor amounts in the NE. In a previous study (13), we found that a variety of histone and histone combinations (H1, H2AH2B, H3H4, and octamers) caused a dose-dependent aggregation of phospholipid vesicles composed mainly of phosphatidylcholine (PC) to which 5-20 mol % phosphoinositides was added. We observed that 5 mol % phosphatidylinositol-4-phosphate (PIP) was enough to cause extensive vesicle aggregation under our conditions, which we interpreted as a (mostly electrostatic) interaction between histones and PIP.…”