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. 2013 Jul 8:14:457.
doi: 10.1186/1471-2164-14-457.

Evolutionary origins, molecular cloning and expression of carotenoid hydroxylases in eukaryotic photosynthetic algae

Hongli Cui  1 , Xiaona Yu, Yan Wang, Yulin Cui, Xueqin Li, Zhaopu Liu, Song Qin
Affiliations

Evolutionary origins, molecular cloning and expression of carotenoid hydroxylases in eukaryotic photosynthetic algae

Hongli Cui et al. BMC Genomics. .

Abstract

Background: Xanthophylls, oxygenated derivatives of carotenes, play critical roles in photosynthetic apparatus of cyanobacteria, algae, and higher plants. Although the xanthophylls biosynthetic pathway of algae is largely unknown, it is of particular interest because they have a very complicated evolutionary history. Carotenoid hydroxylase (CHY) is an important protein that plays essential roles in xanthophylls biosynthesis. With the availability of 18 sequenced algal genomes, we performed a comprehensive comparative analysis of chy genes and explored their distribution, structure, evolution, origins, and expression.

Results: Overall 60 putative chy genes were identified and classified into two major subfamilies (bch and cyp97) according to their domain structures. Genes in the bch subfamily were found in 10 green algae and 1 red alga, but absent in other algae. In the phylogenetic tree, bch genes of green algae and higher plants share a common ancestor and are of non-cyanobacterial origin, whereas that of red algae is of cyanobacteria. The homologs of cyp97a/c genes were widespread only in green algae, while cyp97b paralogs were seen in most of algae. Phylogenetic analysis on cyp97 genes supported the hypothesis that cyp97b is an ancient gene originated before the formation of extant algal groups. The cyp97a gene is more closely related to cyp97c in evolution than to cyp97b. The two cyp97 genes were isolated from the green alga Haematococcus pluvialis, and transcriptional expression profiles of chy genes were observed under high light stress of different wavelength.

Conclusions: Green algae received a β-xanthophylls biosynthetic pathway from host organisms. Although red algae inherited the pathway from cyanobacteria during primary endosymbiosis, it remains unclear in Chromalveolates. The α-xanthophylls biosynthetic pathway is a common feature in green algae and higher plants. The origination of cyp97a/c is most likely due to gene duplication before divergence of green algae and higher plants. Protein domain structures and expression analyses in green alga H. pluvialis indicate that various chy genes are in different manners response to light. The knowledge of evolution of chy genes in photosynthetic eukaryotes provided information of gene cloning and functional investigation of chy genes in algae in the future.

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Figures

Figure 1
Figure 1
Distribution of putative genes encoding CHYs and hypothesized xanthophylls biosynthetic pathway in algae. A] The distribution of genes encoding BCH, CrtR and CYP97 homologs across 18 algal genome sequences. The number of putative homologs or paralogs in each corresponding genome is indicated by "+" with color codes, respectively. B] Hypothesized xanthophylls biosynthetic pathway in algae. The genes encoding putative BCH-, CrtR- or CYP97-homologs identified from different algae genomes were indicated with color arrows. Lutein and α-carotene are absent in red algae and Chromalveolates according to our results. Question mark indicates enzymes involved in xanthophylls biosynthesis are unclear.
Figure 2
Figure 2
Domain structure of BCH from green algae and CrtR from cyanobacteria and red algae. Ten BCH-type CHYs from green algae, one CrtR-type CHY from red algae and five CrtRs from cyanobacteria are included. A partial protein sequence (position: 110–400) has been selected for domain structure analysis. The predicted trans-membrane segments are shaded in red. The histidine boxes are shaded in green and Black stars indicate the positions of conserved histidine residues. Ten BCH-type CHYs from green algae includes Prasionphyceae [Micromonas pusilla, Micromonas sp. RCC299, Ostreococcus sp. RCC809, Ostreococcus tauri, and Ostreococcus lucimarinus]. One CrtR-type CHY from the red alga is Cyanidioschyzon merolae. The information of BCH genes from algae is as in Table 1. Five CrtRs from cyanobacteria includes Synechococcus sp. JA-2-3B'a (2–13) [Cyanobase: CYB_0102], Synechococcus sp. JA-3-3Ab [Cyanobase: CYA_1931], Cyanothece sp. PCC 7425 [Cyanobase: Cyan7425_1008], Acaryochloris marina MBIC11017 [Cyanobase: AM1_3637] and Thermosynechococcus elongatus BP-1 [Cyanobase: tlr1900].
Figure 3
Figure 3
A un-rooted maximum likelihood tree of our BCH database and some other BCHs from bacteria, higher plants and cyanobacteria. The sequences information of BCH from cyanobacteria, bacteria and higher plants was downloaded from Cyanobase or NCBI database respectively and summarized in an additional file [see Additional file 3: Table S2]. A partial protein sequence (position: 110–400) has been selected for phylogenetic analysis. A maximum likelihood phylogenetic tree (loglk = −12176.58485) as inferred from amino acid sequences (291 amino acid characters) of BCH and CrtR proteins was computed using LG model for amino acid substitution (selected by PROTTEST) with discrete gamma distribution in four categories. All parameters (gamma shape = 1.963; proportion of invariants = 0.010; number of categories: 4) were estimated from the dataset. Numbers above branches indicate ML bootstrap supports. ML bootstraps were computed using the above mentioned model in 300 replicates. Stars indicate where later gene duplications led to creation of paralogs genes found within one species. Major groups of organisms are labeled to allow comparison between the phylogeny of BCH and algae evolution.
Figure 4
Figure 4
Domain structure of CYP97 from algae. A partial protein sequence (position: 272–926) has been selected for domain structure analysis. The red shades indicated conserved amino acid residues in CYP97A and CYP97C homologs from green algae. The green shades indicated conserved amino acid residues in all CYP97B homologs from all algae. The P450s active site components were found in the amino acid sequences of all CYP97A/B/C, including I-helix involved in oxygen binding (CD6 in CYP97B and CD5 in CYP97A and CYP97C), ERR triad (CD7 in CYP97A/B/C) involved in locking the heme pockets into position and to assure stabilization of the conserved core structure, and CD10 involved in heme binding and a conserved cysteine (the circle with blue color). The abbreviations used are: Chl-C, CYP97C from Chlorophyta C. reinhardtii, V. carteri, M. sp. RCC299, O. RCC809, and Cercozoa B. natans CCMP2755; Chl-A, CYP97A from Chlorophyta C. reinhardtii, V. carteri, M. sp. RCC299, and O. RCC809; Chl-B, CYP97B from Chlorophyta M. sp. RCC299, O. RCC809, and V. carteri; Cry-B, CYP97B from Cryptophyta G. theta; Bac-B, CYP97B from Bacillariophyta T. pseudonana, P. tricornutum, F. cylindrus, and Stramenopiles A. anophagefferens; Hap-B, CYP97B from E. huxleyi. The information of BCH genes from algae is as in Table 1.
Figure 5
Figure 5
A maximum likelihood tree of our CYP97 database and some other CYP97s from higher plants. The sequences information of CYP97s from higher plants was downloaded from NCBI database and summarized as follow: Arabidopsis thaliana [GenBank: CYP97A3, gb|AEE31394.1, CYP97B3, gb|AEE83557.1, CYP97C1, sp|Q6TBX7.1 and CYP86A, AED97111.1], Zea mays [GenBank: CYP97A16, ACG28871.1], Glycine max [GenBank: carotene epsilon-monooxygenase, XP_003537025.1], Solanum lycopersicum [GenBank: CYP97C11, NP_001234058.1] and Oryza sativa Japonica Group [GenBank: carotene epsilon-monooxygenase, AAK20054.1]. A partial protein sequence (position: 272–926) has been selected for phylogenetic analysis. A maximum likelihood phylogenetic tree (loglk = −27808.68723) as inferred from amino acid sequences (655 amino acid characters) of CYP97 proteins was computed using LG model for amino acid substitution (selected by PROTTEST) with discrete gamma distribution in four categories. All parameters (gamma shape = 1.924; proportion of invariants = 0.011; number of categories: 4) were estimated from the dataset. Numbers above branches indicate ML bootstrap supports. ML bootstraps were computed using the above mentioned model in 300 replicates. The arrow indicates an ancient gene duplication event creating CYP97A/C, respectively. Stars indicate where later gene duplications led to creation of paralogs genes found within one species. Black circle indicate two genes belonged to no one subfamily of CYP97. Major groups of organisms are labeled to allow comparison between the phylogeny of CYP97A/B/C and algae evolution.
Figure 6
Figure 6
mRNA levels of xanthophylls biosynthesis-related genes upon white or blue high light stimulation. The exponentially growing cultures (cell density approximately 5 × 107 cells ml–1) were harvested and transferred cells to 500-ml erlenmeyer flasks (named, 1–9), each containing 250-ml BBM (fresh medium) under continuous white light (390–770 nm) or blue light (420–500 nm) with light intensity of 1,000 μmol photons m–2 s–1 without a day/night cycle, respectively. Collected algal cells (20-mL sample at some selected time) were rinsed with PBS, stored at −80°C if not immediately used. The relative transcript levels of bch[A], Haecyp97a[B], Haecyp97b[C] and Haecyp97c[D] were determined after 2, 4, 6, 10, 13, 24, 28, 34, 48, 54 and 72 h by qRT-PCR using actin as a reference gene. The values were normalized to the transcript levels in the normal light condition. Data are averages of triplicate measurements. The error bars represent standard deviation. Length of the distance in x-axis did not correspond to length of induced time (hours).
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References

    1. Lichtenthaler HK. The 1-Deoxy-D-Xylulose-5-Phosphate pathway of isoprenoid biosynthesis in plants. Annu Rev Plant Physiol Plant Mol Biol. 1999;50(1):47–65. doi: 10.1146/annurev.arplant.50.1.47. - DOI - PubMed
    1. Hirschberg J, Cohen M, Harker M, Lotan T, Mann V, Pecker I. Molecular genetics of the carotenoid biosynthesis pathway in plants and algae. Pure Appl Chem. 1997;69(10):2151–2158. doi: 10.1351/pac199769102151. - DOI
    1. Niyogi KK. Photoprotection revisited: genetic and molecular approaches. Annu Rev Plant Biol. 1999;50(1):333–359. doi: 10.1146/annurev.arplant.50.1.333. - DOI - PubMed
    1. Davison PA, Hunter CN, Horton P. Overexpression of beta-carotene hydroxylase enhances stress tolerance in Arabidopsis. Nature. 2002;418(6894):203–206. doi: 10.1038/nature00861. - DOI - PubMed
    1. DellaPenna D, Pogson BJ. Vitamin synthesis in plants: tocopherols and carotenoids. Annu Rev Plant Biol. 2006;57:711–738. doi: 10.1146/annurev.arplant.56.032604.144301. - DOI - PubMed

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