Whole Genome Analysis of Gene Expression Reveals Coordinated Activation of Signaling and Metabolic Pathways during Pollen-Pistil Interactions in Arabidopsis

Plant reproduction depends on the concerted activation of many genes to ensure correct communication between pollen and pistil. Here, we queried the whole transcriptome of Arabidopsis (Arabidopsis thaliana) in order to identify genes with specific reproductive functions. We used the Affymetrix ATH1 whole genome array to profile wild-type unpollinated pistils and unfertilized ovules. By comparing the expression profile of pistils at 0.5, 3.5, and 8.0 h after pollination and applying a number of statistical and bioinformatics criteria, we found 1,373 genes differentially regulated during pollen-pistil interactions. Robust clustering analysis grouped these genes in 16 time-course clusters representing distinct patterns of regulation. Coregulation within each cluster suggests the presence of distinct genetic pathways, which might be under the control of specific transcriptional regulators. A total of 78% of the regulated genes were expressed initially in unpollinated pistil and/or ovules, 15% were initially detected in the pollen data sets as enriched or preferentially expressed, and 7% were induced upon pollination. Among those, we found a particular enrichment for unknown transcripts predicted to encode secreted proteins or representing signaling and cell wall-related proteins, which may function by remodeling the extracellular matrix or as extracellular signaling molecules. A strict regulatory control in various metabolic pathways suggests that fine-tuning of the biochemical and physiological cellular environment is crucial for reproductive success. Our study provides a unique and detailed temporal and spatial gene expression profile of in vivo pollen-pistil interactions, providing a framework to better understand the basis of the molecular mechanisms operating during the reproductive process in higher plants.

Whole genome sequence of Klebsiella pneumoniae U25, a hypermucoviscous, multidrug resistant, biofilm producing isolate from India

Klebsiella pneumoniae U25 is a multidrug resistant strain isolated from a tertiary care hospital in Chennai, India. Here, we report the complete annotated genome sequence of strain U25 obtained using PacBio RSII. This is the first report of the whole genome of K. pneumoniae species from Chennai. It consists of a single circular chromosome of size 5,491,870-bp and two plasmids of size 211,813 and 172,619-bp. The genes associated with multidrug resistance were identified. The chromosome of U25 was found to have eight antibiotic resistant genes [blaOXA-1, blaSHV-28, aac(6’)1b-cr, catB3, oqxAB, dfrA1]. The plasmid pMGRU25-001 was found to have only one resistant gene (catA1) while plasmid pMGRU25-002 had 20 resistant genes [strAB, aadA1, aac(6’)-Ib, aac(3)-IId, sul1,2, blaTEM-1A,1B, blaOXA-9, blaCTX-M-15, blaSHV-11, cmlA1, erm(B), mph(A)]. A mutation in the porin OmpK36 was identified which is likely to be associated with the intermediate resistance to carbapenems in the absence of carbapenemase genes. U25 is one of the few K. pneumoniae strains to harbour clustered regularly interspaced short palindromic repeats (CRISPR) systems. Two CRISPR arrays corresponding to Cas3 family helicase were identified in the genome. When compared to K. pneumoniae NTUHK2044, a transposase gene InsH of IS5-13 was found inserted.

Whole genome sequencing on the identification of pathogens involved with high mortality in piglets.

2016

Examining the cause of high inbreeding depression: analysis of whole-genome sequence data in 28 selfed progeny of Eucalyptus grandis.

2016

The Genomes of Oryza sativa: A history of duplications

We report improved whole-genome shotgun sequences for the genomes of indica and japonica rice, both with multimegabase contiguity, or almost 1,000-fold improvement over the drafts of 2002. Tested against a nonredundant collection of 19,079 full-length cDNAs, 97.7% of the genes are aligned, without fragmentation, to the mapped superscaffolds of one or the other genome. We introduce a gene identification procedure for plants that does not rely on similarity to known genes to remove erroneous predictions resulting from transposable elements. Using the available EST data to adjust for residual errors in the predictions, the estimated gene count is at least 38,000 - 40,000. Only 2% - 3% of the genes are unique to any one subspecies, comparable to the amount of sequence that might still be missing. Despite this lack of variation in gene content, there is enormous variation in the intergenic regions. At least a quarter of the two sequences could not be aligned, and where they could be aligned, single nucleotide polymorphism ( SNP) rates varied from as little as 3.0 SNP/kb in the coding regions to 27.6 SNP/kb in the transposable elements. A more inclusive new approach for analyzing duplication history is introduced here. It reveals an ancient whole-genome duplication, a recent segmental duplication on Chromosomes 11 and 12, and massive ongoing individual gene duplications. We find 18 distinct pairs of duplicated segments that cover 65.7% of the genome; 17 of these pairs date back to a common time before the divergence of the grasses. More important, ongoing individual gene duplications provide a never-ending source of raw material for gene genesis and are major contributors to the differences between members of the grass family.

酵母基因组进化中一些重要新性状的产生及其效应

本论文用生物信息学的方法对酵母基因组进化中产生的新性状进行了系统 深入的研究。首先，在大多数的真核生物中，线粒体是生物能量生成所必需的细 胞器。但当葡萄糖的含量丰富的时候，即使是在有氧条件下，经过基因组重复 （WGD，whole genome duplication）后的大多酵母也都可以不需要线粒体而执行 发酵过程，而且甚至在线粒体基因组缺陷的情况下仍可以生存。在本次研究中， 我们揭示核编码的线粒体相关基因的进化速率在基因组重复后的物种中比其在 基因组重复前的物种中显著加快。而且这些基因的密码子使用偏好也在基因组重 复后的物种中减弱。密码子使用偏好的模式和一个特殊转录调控因子的分布显示 在基因组重复后的进化支系中，有效的有氧发酵过程的起源时间大致是在 Kluyveromyces polysporus 和 Saccharomyces castellii 从它们的共同祖先分化之 后。根据上述结果我们得出结论，可能正是这种新的能量策略的产生导致了线粒 体相关基因的功能在基因组重复后的物种中选择性放松。 其次，我们系统地研究了一个多细胞真菌Ashbya gossypii 和九个单细胞酵母 之间密码子使用偏好性的差异。细胞周期调控基因一直被认为是它们形态差异的 关键基因。由于A. gossypii 和典型的单细胞酵母Saccharomyces cerevisiae 有几乎 完全一样的细胞周期调控基因，因此形态上的差异可能是由于直系同源基因的表 达调控差异造成的。我们发现在A. gossypii 中细胞周期基因的翻译效率比在其他 单细胞酵母中显著增高，同时也发现单细胞酵母中的新陈代谢基因比其在A. gossypii 中有显著增高的翻译效率。因为基因的翻译效率和该基因在物种中的重 要性密切相关，所以我们观察到的这些基因翻译效率的显著差异可能可以阐明 A. gossypii 和单细胞酵母的形态差异的原因。同时我们的结果对理解真核生物多 细胞的起源过程也有提示意义。

Identification of dopamine receptors across the extant avian family tree and analysis with other clades uncovers a polyploid expansion among vertebrates.

Dopamine is an important central nervous system transmitter that functions through two classes of receptors (D1 and D2) to influence a diverse range of biological processes in vertebrates. With roles in regulating neural activity, behavior, and gene expression, there has been great interest in understanding the function and evolution dopamine and its receptors. In this study, we use a combination of sequence analyses, microsynteny analyses, and phylogenetic relationships to identify and characterize both the D1 (DRD1A, DRD1B, DRD1C, and DRD1E) and D2 (DRD2, DRD3, and DRD4) dopamine receptor gene families in 43 recently sequenced bird genomes representing the major ordinal lineages across the avian family tree. We show that the common ancestor of all birds possessed at least seven D1 and D2 receptors, followed by subsequent independent losses in some lineages of modern birds. Through comparisons with other vertebrate and invertebrate species we show that two of the D1 receptors, DRD1A and DRD1B, and two of the D2 receptors, DRD2 and DRD3, originated from a whole genome duplication event early in the vertebrate lineage, providing the first conclusive evidence of the origin of these highly conserved receptors. Our findings provide insight into the evolutionary development of an important modulatory component of the central nervous system in vertebrates, and will help further unravel the complex evolutionary and functional relationships among dopamine receptors.

Pervasive positive selection on duplicated and nonduplicated vertebrate protein coding genes.

A stringent branch-site codon model was used to detect positive selection in vertebrate evolution. We show that the test is robust to the large evolutionary distances involved. Positive selection was detected in 77% of 884 genes studied. Most positive selection concerns a few sites on a single branch of the phylogenetic tree: Between 0.9% and 4.7% of sites are affected by positive selection depending on the branches. No functional category was overrepresented among genes under positive selection. Surprisingly, whole genome duplication had no effect on the prevalence of positive selection, whether the fish-specific genome duplication or the two rounds at the origin of vertebrates. Thus positive selection has not been limited to a few gene classes, or to specific evolutionary events such as duplication, but has been pervasive during vertebrate evolution.

Algorithmes pour la reconstruction de génomes ancestraux

L’inférence de génomes ancestraux est une étape essentielle pour l’étude de l’évolution des génomes. Connaissant les génomes d’espèces éteintes, on peut proposer des mécanismes biologiques expliquant les divergences entre les génomes des espèces modernes. Diverses méthodes visant à résoudre ce problème existent, se classant parmis deux grandes catégories : les méthodes de distance et les méthodes de synténie. L’état de l’art des distances génomiques ne permettant qu’un certain répertoire de réarrangements pour le moment, les méthodes de synténie sont donc plus appropriées en pratique. Nous proposons une méthode de synténie pour la reconstruction de génomes ancestraux basée sur une définition relaxée d’adjacences de gènes, permettant un contenu en gène inégal dans les génomes modernes causé par des pertes de gènes de même que des duplications de génomes entiers (DGE). Des simulations sont effectuées, démontrant une capacité de former une solution assemblée en un nombre réduit de régions ancestrales contigües par rapport à d’autres méthodes tout en gardant une bonne fiabilité. Des applications sur des données de levures et de plantes céréalières montrent des résultats en accord avec d’autres publications, notamment la présence de fusion imbriquée de chromosomes pendant l’évolution des céréales.

Étude de l’évolution des génomes par duplications, pertes et réarrangements

La duplication est un des évènements évolutifs les plus importants, car elle peut mener à la création de nouvelles fonctions géniques. Durant leur évolution, les génomes sont aussi affectés par des inversions, des translocations (incluant des fusions et fissions de chromosomes), des transpositions et des délétions. L'étude de l'évolution des génomes est importante, notamment pour mieux comprendre les mécanismes biologiques impliqués, les types d'évènements qui sont les plus fréquents et quels étaient les contenus en gènes des espèces ancestrales. Afin d'analyser ces différents aspects de l'évolution des génomes, des algorithmes efficaces doivent être créés pour inférer des génomes ancestraux, des histoires évolutives, des relations d'homologies et pour calculer les distances entre les génomes. Dans cette thèse, quatre projets reliés à l'étude et à l'analyse de l'évolution des génomes sont présentés : 1) Nous proposons deux algorithmes pour résoudre des problèmes reliés à la duplication de génome entier : un qui généralise le problème du genome halving aux pertes de gènes et un qui permet de calculer la double distance avec pertes. 2) Nous présentons une nouvelle méthode pour l'inférence d'histoires évolutives de groupes de gènes orthologues répétés en tandem. 3) Nous proposons une nouvelle approche basée sur la théorie des graphes pour inférer des gènes in-paralogues qui considère simultanément l'information provenant de différentes espèces afin de faire de meilleures prédictions. 4) Nous présentons une étude de l'histoire évolutive des gènes d'ARN de transfert chez 50 souches de Bacillus.

Systematic Variation in the Pattern of Gene Paralog Retention between the Teleost Superorders Ostariophysi and Acanthopterygii

Teleost fish underwent whole-genome duplication around 450 Ma followed by diploidization and loss of 80-85% of the duplicated genes. To identify a deep signature of this teleost-specific whole-genome duplication (TSGD), we searched for duplicated genes that were systematically and uniquely retained in one or other of the superorders Ostariophysi and Acanthopterygii. TSGD paralogs comprised 17-21% of total gene content. Some 2.6% (510) of TSGD paralogs were present as pairs in the Ostariophysi genomes of Danio rerio (Cypriniformes) and Astyanax mexicanus (Characiformes) but not in species from four orders of Acanthopterygii (Gasterosteiformes, Gasterosteus aculeatus; Tetraodontiformes, Tetraodon nigroviridis; Perciformes, Oreochromis niloticus; and Beloniformes, Oryzias latipes) where a single copy was identified. Similarly, 1.3% (418) of total gene number represented cases where TSGD paralogs pairs were systematically retained in the Acanthopterygian but conserved as a single copy in Ostariophysi genomes. We confirmed the generality of these results by phylogenetic and synteny analysis of 40 randomly selected linage-specific paralogs (LSPs) from each superorder and completed with the transcriptomes of three additional Ostariophysi species (Ictalurus punctatus [Siluriformes], Sinocyclocheilus species [Cypriniformes], and Piaractus mesopotamicus [Characiformes]). No chromosome bias was detected in TSGD paralog retention. Gene ontology (GO) analysis revealed significant enrichment of GO terms relative to the human GO SLIM database for growth, Cell differentiation, and Embryo development in Ostariophysi and for Transport, Signal Transduction, and Vesicle mediated transport in Acanthopterygii. The observed patterns of paralog retention are consistent with different diploidization outcomes having contributed to the evolution/diversification of each superorder.

Characterization of the transcriptome of fast and slow muscle myotomal fibres in the pacu (Piaractus mesopotamicus)

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Fgfrl1, a fibroblast growth factor receptor-like gene, is found in the cephalochordate Branchiostoma floridae but not in the urochordate Ciona intestinalis

FGFRL1 is a novel member of the fibroblast growth factor receptor family that controls the formation of musculoskeletal tissues. Some vertebrates, including man, cow, dog, mouse, rat and chicken, possess a single copy the FGFRL1 gene. Teleostean fish have two copies, fgfrl1a and fgfrl1b, because they have undergone a whole genome duplication. Vertebrates belong to the chordates, a phylum that also includes the subphyla of the cephalochordates (e.g. Branchiostoma floridae) and urochordates (tunicates, e.g. Ciona intestinalis). We therefore investigated whether other chordates might also possess an FGFRL1 related gene. In fact, a homologous gene was found in B. floridae (amphioxus). The corresponding protein showed 60% sequence identity with the human protein and all sequence motifs identified in the vertebrate proteins were also conserved in amphioxus Fgfrl1. In contrast, the genome of the urochordate C. intestinalis and those from more distantly related invertebrates including the insect Drosophila melanogaster and the nematode Caenorhabditis elegans did not appear to contain any related sequences. Thus, the FGFRL1 gene might have evolved just before branching of the vertebrate lineage from the other chordates.

NFAT5 genes are part of the osmotic regulatory system in Atlantic salmon (Salmo salar)

Acknowledgements This study was supported by a grant from the Biotechnology and Biological Sciences Research Council (BBSRC, BB/H008063/1), UK to DGH and SAM. Funding also came from Research Council Norway for project number 241016 for DGH and EJ. This work was carried out as part of a PhD thesis funded by the Marine Alliance of Science and Technology Scotland (MASTS).

Gene duplication in the genome of parasitic Giardia lamblia

Background: Giardia are a group of widespread intestinal protozoan parasites in a number of vertebrates. Much evidence from G. lamblia indicated they might be the most primitive extant eukaryotes. When and how such a group of the earliest branching unicellular eukaryotes developed the ability to successfully parasitize the latest branching higher eukaryotes (vertebrates) is an intriguing question. Gene duplication has long been thought to be the most common mechanism in the production of primary resources for the origin of evolutionary novelties. In order to parse the evolutionary trajectory of Giardia parasitic lifestyle, here we carried out a genome-wide analysis about gene duplication patterns in G. lamblia. Results: Although genomic comparison showed that in G. lamblia the contents of many fundamental biologic pathways are simplified and the whole genome is very compact, in our study 40% of its genes were identified as duplicated genes. Evolutionary distance analyses of these duplicated genes indicated two rounds of large scale duplication events had occurred in G. lamblia genome. Functional annotation of them further showed that the majority of recent duplicated genes are VSPs (Variant-specific Surface Proteins), which are essential for the successful parasitic life of Giardia in hosts. Based on evolutionary comparison with their hosts, it was found that the rapid expansion of VSPs in G. lamblia is consistent with the evolutionary radiation of placental mammals. Conclusions: Based on the genome-wide analysis of duplicated genes in G. lamblia, we found that gene duplication was essential for the origin and evolution of Giardia parasitic lifestyle. The recent expansion of VSPs uniquely occurring in G. lamblia is consistent with the increment of its hosts. Therefore we proposed a hypothesis that the increment of Giradia hosts might be the driving force for the rapid expansion of VSPs.