Transcriptome Sequencing of Essential Marine Brown and Red
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Acta Oceanol. Sin., 2014, Vol. 33, No. 2, P. 1–12 DOI: 10.1007/s13131-014-0435-4 http://www.hyxb.org.cn E-mail: [email protected] Transcriptome sequencing of essential marine brown and red algal species in China and its significance in algal biology and phylogeny WU Shuangxiu1,3†, SUN Jing1,3,4†, CHI Shan2†, WANG Liang1,3,4†, WANG Xumin1,3, LIU Cui2, LI Xingang1,3, YIN Jinlong1, LIU Tao2*, YU Jun1,3* 1 CAS Key Laboratory of Genome Sciences and Information, Beijing Key Laboratory of Genome and Precision Medicine Technologies, Beijing Institute of Genomics, Chinese Academy of Sciences, Beijing 100101, China 2 College of Marine Life Science, Ocean University of China, Qingdao 266003, China 3 Beijing Key Laboratory of Functional Genomics for Dao-di Herbs, Beijing Institute of Genomics, Chinese Academy of Sciences, Beijing 100101, China 4 University of Chinese Academy of Sciences, Beijing 100049, China Received 3 April 2013; accepted 26 July 2013 ©The Chinese Society of Oceanography and Springer-Verlag Berlin Heidelberg 2014 Abstract Most phaeophytes (brown algae) and rhodophytes (red algae) dwell exclusively in marine habitats and play important roles in marine ecology and biodiversity. Many of these brown and red algae are also important resources for industries such as food, medicine and materials due to their unique metabolisms and me- tabolites. However, many fundamental questions surrounding their origins, early diversification, taxonomy, and special metabolisms remain unsolved because of poor molecular bases in brown and red algal study. As part of the 1 000 Plant Project, the marine macroalgal transcriptomes of 19 Phaeophyceae species and 21 Rhodophyta species from China's coast were sequenced, covering a total of 2 phyla, 3 classes, 11 orders, and 19 families. An average of 2 Gb per sample and a total 87.3 Gb of RNA-seq raw data were generated. Approxi- mately 15 000 to 25 000 unigenes for each brown algal sample and 5 000 to 10 000 unigenes for each red algal sample were annotated and analyzed. The annotation results showed obvious differences in gene expres- sion and genome characteristics between red algae and brown algae; these differences could even be seen between multicellular and unicellular red algae. The results elucidate some fundamental questions about the phylogenetic taxonomy within phaeophytes and rhodophytes, and also reveal many novel metabolic pathways. These pathways include algal CO2 fixation and particular carbohydrate metabolisms, and related gene/gene family characteristics and evolution in brown and red algae. These findings build on known algal genetic information and significantly improve our understanding of algal biology, biodiversity, evolution, and potential utilization of these marine algae. Key words: Phaeophyceae, brown algae, Rhodophyta, red algae, marine macroalgae, transcriptome sequencing, secondary generation sequencing Citation: Wu Shuangxiu, Sun Jing, Chi Shan, Wang Liang, Wang Xumin, Liu Cui, Li Xingang, Yin Jinlong, Liu Tao, Yu Jun. 2014. Tran- scriptome sequencing of essential marine brown and red algal species in China and its significance in algal biology and phylogeny. Acta Oceanologica Sinica, 33(2): 1–12, doi: 10.1007/s13131-014-0435-4 1 Introduction multicellular (Grosberg and Strathmann, 2007). Algae are a highly diverse group of organisms that live in Both brown and red algae exhibit a range of different hap- a range of aquatic and terrestrial environments (Grossman, loid-diploid life cycles, house a variety of novel metabolic path- 2007). Dwelling exclusively in particular marine habitats, in- ways, and synthesize various unique chemical compounds of cluding some harsh environments, are the phaeophytes, known both ecological and commercial importance (Grossman, 2007). as brown algae belonging to Class Phaeophyceae of Phylum These marine algae serve as major carbon-fixation producers Ochrophyta, and rhodophytes, known as red algae of Phylum and play essential roles in stabilizing different marine ecosys- Rhodophyta. These organisms are morphologically diverse, tems, forming submerged forests or creating niches for a broad varying from unicells about 1 µm in diameter, such as Cyanidi- range of other marine organisms (Cock et al., 2012). As a result, oschyzon merolae (Matsuzaki et al., 2004) and Galdieria sulphu- these environmental tolerance characteristics make brown and raria (Schönknecht et al., 2013), to complex multicellular forms red algae ideal candidates for mechanism study and novel gene reaching lengths of more than 30 m, such as Macrocystis Pyrif- discovery (Misumi et al., 2008). era (Tirichine and Bowler, 2011); though, most phaeophytes are In particular, special polysaccharides, such as alginates and Foundation item: The National Natural Science Foundation of China under contract Nos 31140070, 31271397 and 41206116; the algal transcrip- tome sequencing was supported by 1KP Project (www.onekp.com). *Corresponding author, E-mail: [email protected], [email protected] †Contributed equally. 2 WU Shuangxiu et al. Acta Oceanol. Sin., 2014, Vol. 33, No. 2, P. 1–12 fucoids in brown algae and agars in red algae, as well as their chiangii, which was nominated as Prionitis divaricata previous- numerous and various derivatives, are valuable resources in the ly, and genera Grateloupia and Gracilaria (Wang et al., 2001). production of antitumours, anticoagulants, solid matrices in Algal evolution study is complicated and difficult given cur- medicines, and additives for foods and cosmetics (Berteau and rently available genome data because of multiple methods of Mulloy, 2003; Drury et al., 2003; Matsubara, 2004; Grossman, gene acquisition by algae. Nuclear genomes are mosaics of 2007). Recently, red and brown algae have also attracted grow- genes acquired over long periods of time, not only by vertical ing interest as potential resources for biofuel production due to descent but also by endosymbiotic gene transfer (EGT) and their huge biomass storages (Bartsch et al., 2008). Therefore, the horizontal gene transfer (HGT) during both the primary and corresponding novel carbohydrate metabolism pathways have the secondary endosymbiosis processes (Green, 2011; Tirich- become long-term areas of focus in research. In addition, there ine and Bowler, 2011). Algal evolution study is further compli- is a long-standing debate on the existence of a C4 photosyn- cated by the dearth of existing sequenced red and brown algal thetic pathway during CO2-fixation in marine phytoplankton genomes. Within red algae, C. merolae and G. sulphuraria are (Falkowski and Raven, 1997). However, so far only a few carbo- the only unicellular species that have been sequenced, and Py- hydrate metabolism genes, such as the genes encoding GDP- ropia yezoensis and Chondrus crispus are the only multicellular mannose dehydrogenase of Ectocarpus silicuiosus (Tenhaken et species that have been sequenced. For brown algae, a compre- al., 2011) and mannuronan C-5-epimerase of Laminaria digita- hensive view of genetic characteristics was not available until ta (Nyvall et al., 2003) in the alginate biosynthesis pathway, and 2010, when the complete genome sequence of E. silicilosus, a one gene encoding the first enzyme, mannitol-1-phosphate de- small multicellular brown alga from the order Ectocarpales, hydrogenase in the mannitol biosynthesis pathway (Rousvoal was published (Cock et al., 2010). In addition, expressed se- et al., 2011), have been characterized by molecular biological quence tag (EST) libraries of G. sulphuraria (Weber et al., 2004) experiments. and RNA-seq data of Pyropia yezoensis of Rhodophyta (Liang The origin and evolution of phaeophytes and rhodophytes et al., 2010), Saccharina japonica (Deng et al., 2012), S. latis- is also a research hotspot. Rhodophytes are believed to have sima (Heinrich et al., 2012) and E. siliculosus (Dittami et al., originated from a non-photosynthetic unicellular eukaryote 2009) of Phaeophyceae were the only molecular data available engulfing a photosynthetic cyanobacterium 1.5–1.8 billion for studies in brown algae and red algae until now. Therefore, years ago (Gould et al., 2008; Kutschera and Niklas, 2005; Parker more genome information on more species is needed to solve et al., 2008). Termed the primary endosymbiosis, this event these questions. gave rise to the extant Plantae (or Archaeplastida), consisting In November 2009, a NESCent/iPlant-sponsored 1 000 Plant of three photosynthetic lineages: Glaucophyta, Rhodophyta (1KP) Analysis Workshop was held in Phoenix to initiate the 1 000 (red algae), and a collective group of Chlorophyta (green algae) Plant Transcriptome Sequencing Project (1KP Project, www. and land plants, whose chloroplasts have double layered mem- onekp.com). The project aimed to resolve relationships across branes (Simon et al., 2009). After the primary endosymbiosis, the green plant phylogeny and elucidate processes contributing a second heterotrophic eukaryote engulfed a unicellular green to diversification and biological innovations, including origins or red photosynthetic eukaryote, resulting in a variety of sec- of multicellularity, colonization of land, the evolution of vascu- ondary-endosymbiosis photosynthetic eukaryotes. These sec- lar systems, and the origins of seeds and flowers. The 1KP Proj- ondary-endosymbiosis photosynthetic eukaryotes have three ect will generate unparalleled plant sequence databases for in- or four membraned chloroplasts and include cryptophytes, vestigating the evolution of gene families, regulatory networks haptophytes,