Spermine Condenses DNA, but Not RNA Duplexes

Abstract

Interactions between the polyamine spermine and nucleic acids drive important cellular processes. Spermine condenses DNA and some RNAs, such as poly(rA):poly(rU). A large fraction of the spermine present in cells is bound to RNA but apparently does not condense it. Here, we study the effect of spermine binding to short duplex RNA and DNA, and compare our findings with predictions of molecular-dynamics simulations. When small numbers of spermine are introduced, RNA with a designed sequence containing a mixture of 14 GC pairs and 11 AU pairs resists condensation relative to DNA of an equivalent sequence or to 25 bp poly(rA):poly(rU) RNA. A comparison of wide-angle x-ray scattering profiles with simulation results suggests that spermine is sequestered deep within the major groove of mixed-sequence RNA. This prevents condensation by limiting opportunities to bridge to other molecules and stabilizes the RNA by locking it into a particular conformation. In contrast, for DNA, simulations suggest that spermine binds externally to the duplex, offering opportunities for intermolecular interaction. The goal of this study is to explain how RNA can remain soluble and available for interaction with other molecules in the cell despite the presence of spermine at concentrations high enough to precipitate DNA.

Towards Improved Oligonucleotide Therapeutics Through Faster Target Binding Kinetics

“…[34][35][36] Also, positioning of an aminohexyl group into the major groove of duplex RNA considerably increases T M values. [39] The findings are in agreement with previous studies demonstrating ah igh electronegative potentiali nt he major groove of an Aform helix. [38] Recent modeling studies have described how polyamines bury into the major groove of duplex RNA, stabilizing the duplex structure and conformationally restricting its mobility.…”

“…[39] The findings are in agreement with previous studies demonstrating ah igh electronegative potentiali nt he major groove of an Aform helix. [39,40] It was conceivable that the differences between major and minor groove location of the conjugated polyamine might have resulted from changes in duplex geometry.C D spectroscopy is as tandard method used to investigate helix conformations. [41] Characteristic features of A-form helicesa re a strongp ositive band at 260 nm, as mall local maximum at 225 nm, and as trongn egative band at 210 nm.…”

“…[21,37] Our data is consistentw ith evidence that free polyamines bind favorably to A-form helical structures, in line with the stronger effects of free spermine on DNA/RNA heteroduplexes than on DNA/DNA duplexes, [17] and the preference of polyamines for cellular RNA over DNA. [39,40] It was conceivable that the differences between major and minor groove location of the conjugated polyamine might have resulted from changes in duplex geometry.C D spectroscopy is as tandard method used to investigate helix conformations. [39] The findings are in agreement with previous studies demonstrating ah igh electronegative potentiali nt he major groove of an Aform helix.…”

“…[39,40] It was conceivable that the differences between major and minor groove location of the conjugated polyamine might have resulted from changes in duplex geometry.C D spectroscopy is as tandard method used to investigate helix conformations. [39] Next, we synthesized as econd library of polyamine-conjugated 7n ucleotide-long (7-mer), 9-mer,a nd 10-mer 2'-OMe oligoribonucleotides( ORN 20-32:T able 2; SupportingI nformation), positioning the polyamine fragment at variouss ites within the sequence. Ac hange in duplexc onformationt owards aB -form would shift the strong positive band to higher wavelengths, increase the local maximum at 225 nm with as econd strongp ositive band, and decrease the intensity of the negative band at 210 nm.…”

“…[39,46] Given the dramatic effects of conjugated polyamines bound(presumably) in the major groove of our duplexes,a nd the high intracellular concentrations of endogenous polyamines, [38] we wondered whether natural polyamines mayi nterferew ith the conjugated fragments during hybridization of modified oligonucleotides to RNAs in cells. [39,46] Given the dramatic effects of conjugated polyamines bound(presumably) in the major groove of our duplexes,a nd the high intracellular concentrations of endogenous polyamines, [38] we wondered whether natural polyamines mayi nterferew ith the conjugated fragments during hybridization of modified oligonucleotides to RNAs in cells.…”