We report an improved draft nucleotide sequence of the 2.3-gigabase genome of maize, an important crop plant and model for biological research. Over 32,000 genes were predicted, of which 99.8% were placed on reference chromosomes. Nearly 85% of the genome is composed of hundreds of families of transposable elements, dispersed nonuniformly across the genome. These were responsible for the capture and amplification of numerous gene fragments and affect the composition, sizes, and positions of centromeres. We also report on the correlation of methylation-poor regions with Mu transposon insertions and recombination, and copy number variants with insertions and/or deletions, as well as how uneven gene losses between duplicated regions were involved in returning an ancient allotetraploid to a genetically diploid state. These analyses inform and set the stage for further investigations to improve our understanding of the domestication and agricultural improvements of maize.
Complete and accurate reference genomes and annotations provide fundamental tools for characterization of genetic and functional variation 1 . These resources facilitate the determination of biological processes and support translation of research findings into improved and sustainable agricultural technologies. Many reference genomes for crop plants have been generated over the past decade, but these genomes are often fragmented and missing complex repeat regions 2 . Here we report the assembly and annotation of a reference genome of maize, a genetic and agricultural model species, using single-molecule real-time sequencing and high-resolution optical mapping. Relative to the previous reference genome 3 , our assembly features a 52-fold increase in contig length and notable improvements in the assembly of intergenic spaces and centromeres. Characterization of the repetitive portion of the genome revealed more than 130,000 intact transposable elements, allowing us to identify transposable element lineage expansions that are unique to maize. Gene annotations were updated using 111,000 full-length transcripts obtained by single-molecule real-time sequencing 4 . In addition, comparative optical mapping of two other inbred maize lines revealed a prevalence of deletions in regions of low gene density and maize lineage-specific genes.
Guanine-rich nucleic acid sequences can adopt noncanonical four-stranded secondary structures called guanine (G)-quadruplexes1. Bioinformatics analysis suggests that G-quadruplex motifs are prevalent in genomes2, which raises the need to elucidate their function. There is now evidence for the existence of DNA G-quadruplexes at telomeres with associated biological function3. A recent hypothesis supports the notion that gene promoter elements contain DNA G-quadruplex motifs that control gene expression at the transcriptional level4. We discovered a highly conserved, thermodynamically stable RNA G-quadruplex in the 5′ untranslated region (UTR) of the gene transcript of the human NRAS proto-oncogene. Using a cell-free translation system coupled to a reporter gene assay, we have demonstrated that this NRAS RNA G-quadruplex modulates translation. This is the first example of translational repression by an RNA Gquadruplex. Bioinformatics analysis has revealed 2,922 other 5′ UTR RNA G-quadruplex elements in the human genome. We propose that RNA G-quadruplexes in the 5′ UTR modulate gene expression at the translational level.The existence of RNA G-quadruplexes in vivo is more inevitable than the existence of DNA G-quadruplexes, given that (i) the former are generally more thermodynamically stable in the folded form than their DNA counterparts5, and (ii) RNA is single-stranded, which implies that quadruplex formation does not have to compete with hybridization to a complementary strand. In this study we have focused on the 5′ UTRs of mRNA, which are known to be involved in translational regulation, particularly for growth factors, transcription factors and oncoproteins6. The neuroblastoma RAS viral oncogene homolog (NRAS)-encoded protein p21 mediates both signal transduction across the plasma membrane and the intracellular signaling pathways responsible for cell proliferation and differentiation7. Activating mutations in the coding region of NRAS are responsible for increased cell proliferation8. The suppression of oncogenic NRAS by small interfering RNA causes apoptosis of tumor cells9, which suggests that inhibiting the expression of oncogenic NRAS is a potential therapeutic strategy. Using a computational search algorithm we developed for locating quadruplex sequence motifs2, we identified a putative G-quadruplex Fig. 1 online). This motif is highly conserved, in both its sequence and its position relative to the translation start site, across the 5′ UTRs of human, chimpanzee, macaque, mouse, rat and dog genes orthologous to NRAS (Table 1 and Supplementary Table 1 online).To confirm that the putative RNA G-quadruplex NRQ folds into a stable quadruplex, we carried out biophysical experiments on the synthetic oligonucleotide 5′-UGUGGGAGGGGCGGGUCUGGG-3′. Circular dichroism (CD) spectroscopy has been widely used to characterize the structure of folded nucleic acid quadruplexes11. At pH 7.4, 100 mM KCl, the CD spectrum of NRQ showed a positive peak at 263 nm and a negative peak at 241 nm ( Fig. 1a), which is the ch...
Molecular mechanisms that regulate gene expression can occur either before or after transcription. The information for post-transcriptional regulation can lie within the sequence or structure of the RNA transcript and it has been proposed that G-quadruplex nucleic acid sequence motifs may regulate translation as well as transcription. Here, we have explored the incidence of G-quadruplex motifs in and around the untranslated regions (UTRs) of mRNA. We observed a significant strand asymmetry, consistent with a general depletion of G-quadruplex-forming RNA. We also observed a positional bias in two distinct regions, each suggestive of a specific function. We observed an excess of G-quadruplex motifs towards the 5′-ends of 5′-UTRs, supportive of a hypothesis linking 5′-UTR RNA G-quadruplexes to translational control. We then analysed the vicinity of 3′-UTRs and observed an over-representation of G-quadruplex motifs immediately after the 3′-end of genes, especially in those cases where another gene is in close proximity, suggesting that G-quadruplexes may be involved in the termination of gene transcription.
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