The effects of molecular crowding and CpG hypermethylation on DNA G-quadruplexes formed by the C9orf72 nucleotide repeat expansion

A nucleotide repeat expansion (NRE), (G4C2)n, located in a classically noncoding region of C9orf72 (C9), is the most common genetic mutation associated with ALS/FTD. There is increasing evidence that nucleic acid structures formed by the C9-NRE may both contribute to ALS/FTD, and serve as therapeutic targets, but there is limited characterization of these nucleic acid structures under physiologically and disease relevant conditions. Here we show in vitro that the C9-NRE DNA can form both parallel and antiparallel DNA G-quadruplex (GQ) topological structures and that the structural preference of these DNA GQs can be dependent on the molecular crowding conditions. Additionally, 5-methylcytosine DNA hypermethylation, which is observed in the C9-NRE locus in some patients, has minimal effects on GQ topological preferences. Finally, molecular dynamic simulations of methylated and nonmethylated GQ structures support in vitro data showing that DNA GQ structures formed by the C9-NRE DNA are stable, with structural fluctuations limited to the cytosine-containing loop regions. These findings provide new insight into the structural polymorphic preferences and stability of DNA GQs formed by the C9-NRE in both the methylated and nonmethylated states, as well as reveal important features to guide the development of upstream therapeutic approaches to potentially attenuate C9-NRE-linked diseases.

Identification, distribution and comparative analysis of microsatellites in the chloroplast genome of Oryza species

Chloroplast genome is important because of its maternally inherited, conserved within species and stable in structure. This allows elucidation of inter-specific comparison, identification and establishment of evolutionary relationship among different species. Different classes of DNA elements were present in chloroplast genome of Oryza species, among them chloroplast microsatellites were supposed to be highly variable. In this study, a total of 102 sequences of chloroplast genome of 23 species of Oryza genus belonging to ten different genome types were downloaded from NCBI nucleotide database. The chloroplast DNA length varied from 134,401bp to 136,133bp. Microsatellite analysis revealed that total SSR number varied from 268 in O. coarctata to 281 in O. granulata. Comparative analysis of repeat type revealed that mononucleotide and trinucleotide were least present, while penta and hexa nucleotide motif were the most common type among all the Oryza species. Frequency analysis of the repeat revealed that T repeat among mono, AT among di, AAT among tri, AATA among tetra, AATTC among penta and AAAGAA among hexa nucleotide repeat was found to be common among most of the Oryza species. Total of five set of SSR markers flanking to penta and hexa nucleotide repeat types were developed which uniquely present among species, namely, O. barthi, O. sativa Japonica, O. cocarctata, O. rhizomatis and O. ridley. Phylogeny based on SSR markers indicate separate evolution of different SSR markers among individual species.