Comprehensive profiling of rhizome-associated alternative splicing and alternative polyadenylation in moso bamboo (Phyllostachys edulis).

Moso bamboo (Phyllostachys edulis) represents one of the fastest-spreading plants in the world, due in part to its well-developed rhizome system. However, the post-transcriptional mechanism for the development of the rhizome system in bamboo has not been comprehensively studied. We therefore used a combination of single-molecule long-read sequencing technology and polyadenylation site sequencing (PAS-seq) to re-annotate the bamboo genome, and identify genome-wide alternative splicing (AS) and alternative polyadenylation (APA) in the rhizome system. In total, 145 522 mapped full-length non-chimeric (FLNC) reads were analyzed, resulting in the correction of 2241 mis-annotated genes and the identification of 8091 previously unannotated loci. Notably, more than 42 280 distinct splicing isoforms were derived from 128 667 intron-containing full-length FLNC reads, including a large number of AS events associated with rhizome systems. In addition, we characterized 25 069 polyadenylation sites from 11 450 genes, 6311 of which have APA sites. Further analysis of intronic polyadenylation revealed that LTR/Gypsy and LTR/Copia were two major transposable elements within the intronic polyadenylation region. Furthermore, this study provided a quantitative atlas of poly(A) usage. Several hundred differential poly(A) sites in the rhizome-root system were identified. Taken together, these results suggest that post-transcriptional regulation may potentially have a vital role in the underground rhizome-root system

THE ROLE OF POLYADENYLATION IN SEED GERMINATION

Seed germination has many impacts on the uses of seeds, and is an important subject for study. Seed germination is regulated at both transcriptional and post-transcriptional levels. Therefore, it is important to study how polyadenylation regulates gene expression during seed germination. To this end, a modified Illumina GAIIx sequencing protocol (described in Chapter Two) was developed that allows deep coverage of poly(A) site position and distribution. Alternative polyadenylation (APA) regulates gene expression by choosing one potential poly(A) site on a precursor RNA consequentially shortening/lengthening the mRNA relative to other possible sites. To further explore this phenomenon, genes affected by APA during seed germination and other developmental stages were identified (Chapter Three). These genes were categorized based on the location of poly(A) sites. Several genes were chosen to demonstrate how APA, especially that occurring in the coding regions and 5’ untranslated regions, might down regulate gene expression by generating truncated transcripts. In animal oocytes, maternally-derived mRNAs are stored with short poly(A) tails and reactivated by the cytoplasmic polyadenylation complex. It has been reported that seeds also contain stored mRNAs. Moreover, germination and its completion are less sensitive to de novo transcription inhibitors than to poly(A) polymerase inhibitors. Together, these considerations suggest that stored RNA without or with a short poly(A) tail (stored, unadenylated RNA) may be present in dry seed and function in seed germination upon reactivation by cytoplasmic polyadenylation. To further explore this, in Chapter Four, mRNA polyadenylation was studied through the course of germination using a combination of transcriptional inhibitors and the modified sequencing protocol described in Chapter Two. 273 putative stored, unadenylated RNAs were identified. Gene ontology analysis revealed that genes whose products are involved in translation are overrepresented; these genes encode 21 60S- and 10 40S-ribosomal proteins. These results indicate that transcripts whose products are involved in translation might be a major component of the stored, unadenylated RNA pool and, more importantly, translation might be the first cellular process to be activated during seed germination

Role of alternative polyadenylation in epigenetic silencing and antisilencing

Epigenetic marks such as DNA methylation and histone modifications are widely involved in regulating different aspects of developmental and environmental responses (1). Meanwhile, DNA methylation and histone modification are also used constitutively to silence transposable elements and repeat elements (TREs) (2). Such TRE-mediated silencing should necessarily be limited to the intended targets only and not spread to adjacent genes and their regulatory elements. Higher eukaryotic organisms have evolved antisilencing mechanisms to keep the balance between silencing and antisilencing that is required for precise gene expression regulation.Research in the authors' laboratory is supported by US National Science Foundation Grant IOS–0817829 (to Q.Q.L.) and US National Institutes of Health Grant 1R15GM94732-1 A1 (to Q.Q.L.)

A comparative metabolomics analysis of domestic yak (Bos grunniens) milk with human breast milk

Yaks are tough animals living in Tibet’s hypoxic stress environment. However, the metabolite composition of yak milk and its role in hypoxic stress tolerance remains largely unexplored. The similarities and differences between yak and human milk in hypoxic stress tolerance are also unclear. This study explored yak colostrum (YC) and yak mature milk (YMM) using GC–MS, and 354 metabolites were identified in yak milk. A comparative metabolomic analysis of yak and human milk metabolites showed that over 70% of metabolites were species-specific. Yak milk relies mainly on essential amino acids- arginine and essential branched-chain amino acids (BCAAs): L-isoleucine, L-leucine, and L-valine tolerate hypoxic stress. To slow hypoxic stress, human breast milk relies primarily on the neuroprotective effects of non-essential amino acids or derivates, such as citrulline, sarcosine, and creatine. In addition, metabolites related to hypoxic stress were significantly enriched in YC than in YMM. These results reveal the unique metabolite composition of yak and human milk and provide practical information for applying yak and human milk to hypoxic stress tolerance

Direct sulphate-TOR signalling controls transcriptional reprogramming for shoot apex activation in <i>Arabidopsis</i>

AbstractPhotosynthetic plants play a primary role for the global sulphur cycle in the earth ecosystems by reduction of inorganic sulphate from the soil to organic sulphur-containing compounds. How plants sense and transduce the sulphate availability in soil to mediate their growth remains largely unclear. The target of rapamycin (TOR) kinase is an evolutionarily conserved master regulator of nutrient sensing and metabolic signalling to control cell proliferation and growth in all eukaryotes. Here, we discovered that inorganic sulphate exhibits higher potency than organic cysteine and glutathione for activation of TOR and cell proliferation in the leaf primordium to promote true leaf development in Arabidopsis. Chemical genetic analyses further revealed that this sulphate activation of TOR is independent of the sulphate-assimilation process and glucose-energy signalling. Significantly, tissue specific transcriptome analyses uncovered previously unknown sulphate-orchestrating genes involved in DNA replication, cell proliferation, autophagy and various secondary metabolism pathways, which are completely depending on TOR signalling. Systematic comparison between the sulphate- and glucose-TOR controlled transcriptome further revealed that, as the central growth integrator, TOR kinase can sense different upstream nutrient signals to control both shared and unique transcriptome networks, therefore, precisely modulate plant proliferation, growth and stress responses.</jats:p

Genome-wide analysis of GATA factors in moso bamboo (<i>Phyllostachys edulis</i>) unveils that PeGATAs regulate shoot rapid-growth and rhizome development

ABSTRACTBackgroundMoso bamboo is well-known for its rapid-growth shoots and widespread rhizomes. However, the regulatory genes of these two processes are largely unexplored. GATA factors regulate many developmental processes, but its role in plant height control and rhizome development remains unclear.ResultsHere, we found that bamboo GATA factors (PeGATAs) are involved in the growth regulation of bamboo shoots and rhizomes. Bioinformatics and evolutionary analysis showed that there are 31 PeGATA factors in bamboo, which can be divided into three subfamilies. Light, hormone, and stress-relatedcis-elements were found in the promoter region of thePeGATAgenes. Gene expression of 12PeGATAgenes was regulated by phytohormone-GA but there was no correlation between auxin andPeGATAgene expression. More than 27PeGATAgenes were differentially expressed in different tissues of rhizomes, and almost allPeGATAshave dynamic gene expression level during the rapid-growth of bamboo shoots. These results indicate thatPeGATAsregulate rhizome development and bamboo shoot growth partially via GA signaling pathway. In addition,PeGATA26,a rapid-growth negative regulatory candidate gene modulated by GA treatment, was overexpressed in Arabidopsis, and over-expression ofPeGATA26significantly repressed Arabidopsis primary root length and plant height. The PeGATA26 overexpressing lines were also resistant to exogenous GA treatment, further emphasizing that PeGATA26 inhibits plant height from Arabidopsis to moso bamboo via GA signaling pathway.ConclusionsOur results provide an insight into the function of GATA transcription factors in regulating shoot rapid-growth and rhizome development, and provide genetic resources for engineering plant height.</jats:sec