A high-quality reference genome is critical for understanding genome structure, genetic variation and evolution of an organism. Here we report the de novo assembly of an indica rice genome Shuhui498 (R498) through the integration of single-molecule sequencing and mapping data, genetic map and fosmid sequence tags. The 390.3 Mb assembly is estimated to cover more than 99% of the R498 genome and is more continuous than the current reference genomes of japonica rice Nipponbare (MSU7) and Arabidopsis thaliana (TAIR10). We annotate high-quality protein-coding genes in R498 and identify genetic variations between R498 and Nipponbare and presence/absence variations by comparing them to 17 draft genomes in cultivated rice and its closest wild relatives. Our results demonstrate how to de novo assemble a highly contiguous and near-complete plant genome through an integrative strategy. The R498 genome will serve as a reference for the discovery of genes and structural variations in rice.
Purple acid phosphatases (PAPs) are members of the metallo-phosphoesterase family. They are characterized by the presence of seven conserved amino acid residues involved in coordinating the dimetal nuclear center in their reactive site. We compared the 29 PAPs predicted for Arabidopsis thaliana in their varieties of potential metal-ligating residues. Although 24 members possessed sets of metal-ligating residues typical of known PAPs, 1 member lacked four of the seven residues. For the remaining four members, potential metal-ligating residues were generally more similar to those in metal-dependent exonucleases and related proteins. Evidence was obtained for the expression of the majority of the 29 PAPs. To facilitate future investigations, a scheme for naming Arabidopsis PAPs and a system for classifying the 29 PAPs are proposed. The cDNA sequences and the responses to phosphate deprivation of seven Arabidopsis PAPs (AtPAP7-AtPAP13) were characterized. For some AtPAPs analyzed, there were fully processed transcripts as well as splice variants. The splice variants of AtPAP10 were found to associate with polyribosomes and may be translated into a NH 2 -terminal truncated protein. Phylogenetic investigations showed that AtPAPs 7 and 8, together with similar enzymes from other plant species, formed the low molecular weight plant PAP group. Members of this group were more closely related to PAPs from mammalian cells. AtPAPs 9 -13, together with kidney bean PAP, formed the high molecular weight PAP group. In phosphate deprivation experiments, gene transcription of AtPAP11 and AtPAP12 was induced and increased, respectively, whereas that of the remaining five AtPAPs was not affected by phosphate deprivation. The present work demonstrates that structure variation and expression regulation of plant PAPs are more complex than previously described and provides a framework for comprehensive molecular genetic and biochemical studies of all Arabidopsis PAPs in the future.
Purple acid phosphatases (PAPs) are metallo-phosphoesterases. Their expression and function have not been systematically investigated in higher plants. In this work, we compared the transcript levels of 28 Arabidopsis PAP (AtPAP) genes in five Arabidopsis organs. The 28 members, although differed in their expression patterns in vegetative organs, were all transcribed in flower. Furthermore, the transcription of seven members (AtPAPs 6, 11, 14, 19, 23, 24 and 25) occurred predominantly in the flower. To begin dissecting the role of AtPAP genes in flower development, further expression and functional analyses were conducted using AtPAP23. Histochemical staining of transgenic plants expressing AtPAP23 promoter-beta-glucuronidase (GUS) gene construct revealed that AtPAP23 transcription was strong in flower apical meristems, but became restricted to petals and anther filaments in fully developed flower. A GST (glutathione S-transferase) fusion protein of AtPAP23 (GST:AtPAP23) was expressed in bacterial cells, and was found to contain significant amounts of Fe and Mn (whereas the control GST protein contained none). In biochemical tests, GST:AtPAP23 showed typical acid phosphatase activities. The fusion protein was also highly active on phosphoserine, but not phosphotyrosine. Despite its highly specific expression pattern and the demonstrated biochemical function of its protein product, the RNAi (RNA interference), T-DNA knock-out and overexpression lines of AtPAP23 were indistinguishable from wild type plants in the development of flower (or other organs). Interestingly, the Fe and Mn contents were found significantly increased in AtPAP23 overexpression lines, which may offer a new direction for further functional studies of AtPAPs in Arabidopsis.
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