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De Novo Transcriptome Analysis of Warburgia ugandensis to Identify Genes Involved in Terpenoids and Unsaturated Fatty Acids Biosynthesis.

Wang X, Zhou C, Yang X, Miao D, Zhang Y - PLoS ONE (2015)

Bottom Line: In addition, the expression of 12 DEGs with putative roles in terpenoid and unsaturated fatty acid metabolic pathways was confirmed by qRT-PCRs, which was consistent with the data of the RNA-sequencing.In conclusion, we constructed a comprehensive transcriptome dataset derived from the bark and leaf of W. ugandensis, which forms the basis for functional genomics studies on this plant species.Particularly, the comparative analysis of the transcriptome data between the bark and leaf will provide critical clues to reveal the regulatory mechanisms underlying the biosynthesis of terpenoids and PUFAs in W. ugandensis.

View Article: PubMed Central - PubMed

Affiliation: CAS Key Laboratory of Plant Germplasm Enhancement and Specialty Agriculture, Wuhan Botanical Garden, Chinese Academy of Sciences, Wuhan, Hubei, China.

ABSTRACT
The bark of Warburgia ugandensis (Canellaceae family) has been used as a medicinal source for a long history in many African countries. The presence of diverse terpenoids and abundant polyunsaturated fatty acids (PUFAs) in this organ contributes to its broad range of pharmacological properties. Despite its medicinal and economic importance, the knowledge on the biosynthesis of terpenoid and unsaturated fatty acid in W. ugandensis bark remains largely unknown. Therefore, it is necessary to construct a genomic and/or transcriptomic database for the functional genomics study on W. ugandensis. The chemical profiles of terpenoids and fatty acids between the bark and leaves of W. ugandensis were compared by gas chromatography-mass spectrometry (GC-MS) analysis. Meanwhile, the transcriptome database derived from both tissues was created using Illumina sequencing technology. In total, about 17.1 G clean nucleotides were obtained, and de novo assembled into 72,591 unigenes, of which about 38.06% can be aligned to the NCBI non-redundant protein database. Many candidate genes in the biosynthetic pathways of terpenoids and unsaturated fatty acids were identified, including 14 unigenes for terpene synthases. Furthermore, 2,324 unigenes were discovered to be differentially expressed between both tissues; the functions of those differentially expressed genes (DEGs) were predicted by gene ontology enrichment and metabolic pathway enrichment analyses. In addition, the expression of 12 DEGs with putative roles in terpenoid and unsaturated fatty acid metabolic pathways was confirmed by qRT-PCRs, which was consistent with the data of the RNA-sequencing. In conclusion, we constructed a comprehensive transcriptome dataset derived from the bark and leaf of W. ugandensis, which forms the basis for functional genomics studies on this plant species. Particularly, the comparative analysis of the transcriptome data between the bark and leaf will provide critical clues to reveal the regulatory mechanisms underlying the biosynthesis of terpenoids and PUFAs in W. ugandensis.

No MeSH data available.


Phylogenetic tree of terpene synthases.Phylogenetic analysis of 14 putative W. ugandensis WuTPS protein sequences with their homologs from other plants indicates that they are clustered into three main clades including: monoterpenoid synthase (WuMts), sesquiterpenoid synthase (WuSps), and diterpenoid synthase (WuDts). WarbTPS-c (ACJ46047.1, putative sesquiterpene synthase, W. ugandensis); WarbTPS-g (ACJ46048.1, putative sesquiterpene synthase, W. ugandensis); WuSps1, CL29873Contig1; WuSps2, CL29511Contig1; WuSps3, CL3178Contig1; WuSps4, CL4160Contig1; WuSps5, comp68897_c0_seq1; WuSps6, CL24969Contig1; WuSps7, CL30258Contig1; WuMts1, CL27268Contig1; WuMts2, CL27339Contig1; WuMts3, CL30385Contig1; WuMts4, CL276Contig2; WuMts5, CL1Contig9269; WuMts6, CL29966Contig1; WuMts7, CL14869Contig1; WuDts1, CL9128Contig1; WuDts2, CL28648Contig1; Citsi_Germacrene_D (XP_006494713.1, (-)-germacrene D synthase-like isoform X2, Citrus sinensis); Popeu_Valencene (XP_011015484.1, valencene synthase-like, Populus euphratica); Nelnu_Germacrene_D (XP_010258444.1, (-)-germacrene D synthase-like, Nelumbo nucifera); Eletr_Copaene (ADK94034.1, alpha-copaene synthase, Eleutherococcus trifoliatus); Gosar_Germacrene_D (KHG04103.1, (-)-germacrene D synthase, Gossypium arboretum); Vitvi_Germacrene_D (XP_010644711.1, (-)-germacrene D synthase, Vitis vinifera); Vitvi_Germacrene_A (ADR66821.1, Germacrene A synthase, Vitis vinifera); Citja_Elemene (BAP74389.1, delta-elemene synthase, Citrus jambhiri); Theca_Cadinene (EOY12648.1, Delta-cadinene synthase isozyme A, Theobroma cacao); Ricco_Cadinene (EEF38721.1, (+)-delta-cadinene synthase isozyme A, Ricinus communis); Aqusi_Guaiene (AIT75875.1, putative delta-guaiene synthase, Aquilaria sinensis); Vitvi_Caryophyllene (AEP17005.1, (E)-beta-caryophyllene synthase, Vitis vinifera); Maggr_Cubebene (ACC66281.1, beta-cubebene synthase, Magnolia grandiflora); Cinos_Linalool (AFK09265.1, S-(+)-linalool synthase, Cinnamomum osmophloeum); Nelnu_Nerolidol (XP_010248179.1, (3S,6E)-nerolidol synthase 1-like, Nelumbo nucifera); Vitvi_Nerolidol (XP_010646919.1, (3S,6E)-nerolidol synthase 1, chloroplastic-like isoform X1, Vitis vinifera); Vitvi_Linalool (ADR74212.1, (3S)-linalool/(E)-nerolidol synthase, Vitis vinifera); Actpo_Linalool (ADD81295.1, linalool synthase, Actinidia polygama); Nelnu_Ent-copalyl (XP_010277558.1, ent-copalyl diphosphate synthase, chloroplastic-like, Nelumbo nucifera); Theca_Ent-copalyl (XP_007050589.1, Copalyl diphosphate synthase, Theobroma cacao); Morno_Ent-copalyl (XP_010090409.1, Ent-copalyl diphosphate synthase, Morus notabilis); Gosar_Ent-copalyl (KHG01750.1, Ent-copalyl diphosphate synthase, chloroplastic, Gossypium arboreum); Nelnu_Ent-kaurene (XP_010260722.1, ent-kaur-16-ene synthase, chloroplastic isoform X1, Nelumbo nucifera); Phoda_Ent-kaurene (XP_008809130.1, ent-kaur-16-ene synthase, chloroplastic, Phoenix dactylifera); Ricico_Ent-kaurene (XP_002533694.1, Ent-kaurene synthase B, chloroplast precursor, Ricinus communis); Popeu_Ent-kaurene (XP_011014299.1, ent-kaur-16-ene synthase, chloroplastic, Populus euphratica); Nicta_Epi-Aristolochene (3M02.A, 5-Epi-Aristolochene Synthase, Nicotiana tabacum); Soltu_Vetispiradiene (Q9XJ32.1, vetispiradiene synthase 1, Solanum tuberosum); Litcu_Ocimene (AEJ91554.1, trans-ocimene synthase, Litsea cubeba); Litcu_Thujene (AEJ91555.1, alpha-thujene synthase, Litsea cubeba); Citli_Limonene (AAM53946.1, (+)-limonene synthase 2, Citrus limon); Vitvi_Ocimene/Myrcene (ADR74206.1, (E)-beta-ocimene/myrcene synthase, Vitis vinifera); Queil_Pinene (CAK55186.1, pinene synthase, Quercus ilex).
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pone.0135724.g005: Phylogenetic tree of terpene synthases.Phylogenetic analysis of 14 putative W. ugandensis WuTPS protein sequences with their homologs from other plants indicates that they are clustered into three main clades including: monoterpenoid synthase (WuMts), sesquiterpenoid synthase (WuSps), and diterpenoid synthase (WuDts). WarbTPS-c (ACJ46047.1, putative sesquiterpene synthase, W. ugandensis); WarbTPS-g (ACJ46048.1, putative sesquiterpene synthase, W. ugandensis); WuSps1, CL29873Contig1; WuSps2, CL29511Contig1; WuSps3, CL3178Contig1; WuSps4, CL4160Contig1; WuSps5, comp68897_c0_seq1; WuSps6, CL24969Contig1; WuSps7, CL30258Contig1; WuMts1, CL27268Contig1; WuMts2, CL27339Contig1; WuMts3, CL30385Contig1; WuMts4, CL276Contig2; WuMts5, CL1Contig9269; WuMts6, CL29966Contig1; WuMts7, CL14869Contig1; WuDts1, CL9128Contig1; WuDts2, CL28648Contig1; Citsi_Germacrene_D (XP_006494713.1, (-)-germacrene D synthase-like isoform X2, Citrus sinensis); Popeu_Valencene (XP_011015484.1, valencene synthase-like, Populus euphratica); Nelnu_Germacrene_D (XP_010258444.1, (-)-germacrene D synthase-like, Nelumbo nucifera); Eletr_Copaene (ADK94034.1, alpha-copaene synthase, Eleutherococcus trifoliatus); Gosar_Germacrene_D (KHG04103.1, (-)-germacrene D synthase, Gossypium arboretum); Vitvi_Germacrene_D (XP_010644711.1, (-)-germacrene D synthase, Vitis vinifera); Vitvi_Germacrene_A (ADR66821.1, Germacrene A synthase, Vitis vinifera); Citja_Elemene (BAP74389.1, delta-elemene synthase, Citrus jambhiri); Theca_Cadinene (EOY12648.1, Delta-cadinene synthase isozyme A, Theobroma cacao); Ricco_Cadinene (EEF38721.1, (+)-delta-cadinene synthase isozyme A, Ricinus communis); Aqusi_Guaiene (AIT75875.1, putative delta-guaiene synthase, Aquilaria sinensis); Vitvi_Caryophyllene (AEP17005.1, (E)-beta-caryophyllene synthase, Vitis vinifera); Maggr_Cubebene (ACC66281.1, beta-cubebene synthase, Magnolia grandiflora); Cinos_Linalool (AFK09265.1, S-(+)-linalool synthase, Cinnamomum osmophloeum); Nelnu_Nerolidol (XP_010248179.1, (3S,6E)-nerolidol synthase 1-like, Nelumbo nucifera); Vitvi_Nerolidol (XP_010646919.1, (3S,6E)-nerolidol synthase 1, chloroplastic-like isoform X1, Vitis vinifera); Vitvi_Linalool (ADR74212.1, (3S)-linalool/(E)-nerolidol synthase, Vitis vinifera); Actpo_Linalool (ADD81295.1, linalool synthase, Actinidia polygama); Nelnu_Ent-copalyl (XP_010277558.1, ent-copalyl diphosphate synthase, chloroplastic-like, Nelumbo nucifera); Theca_Ent-copalyl (XP_007050589.1, Copalyl diphosphate synthase, Theobroma cacao); Morno_Ent-copalyl (XP_010090409.1, Ent-copalyl diphosphate synthase, Morus notabilis); Gosar_Ent-copalyl (KHG01750.1, Ent-copalyl diphosphate synthase, chloroplastic, Gossypium arboreum); Nelnu_Ent-kaurene (XP_010260722.1, ent-kaur-16-ene synthase, chloroplastic isoform X1, Nelumbo nucifera); Phoda_Ent-kaurene (XP_008809130.1, ent-kaur-16-ene synthase, chloroplastic, Phoenix dactylifera); Ricico_Ent-kaurene (XP_002533694.1, Ent-kaurene synthase B, chloroplast precursor, Ricinus communis); Popeu_Ent-kaurene (XP_011014299.1, ent-kaur-16-ene synthase, chloroplastic, Populus euphratica); Nicta_Epi-Aristolochene (3M02.A, 5-Epi-Aristolochene Synthase, Nicotiana tabacum); Soltu_Vetispiradiene (Q9XJ32.1, vetispiradiene synthase 1, Solanum tuberosum); Litcu_Ocimene (AEJ91554.1, trans-ocimene synthase, Litsea cubeba); Litcu_Thujene (AEJ91555.1, alpha-thujene synthase, Litsea cubeba); Citli_Limonene (AAM53946.1, (+)-limonene synthase 2, Citrus limon); Vitvi_Ocimene/Myrcene (ADR74206.1, (E)-beta-ocimene/myrcene synthase, Vitis vinifera); Queil_Pinene (CAK55186.1, pinene synthase, Quercus ilex).

Mentions: TPSs are the primary enzymes responsible for catalyzing the conversion of prenyl diphosphates (GPP or FPP) into monoterpenes (C10), sesquiterpenes (C15), or diterpenes (C20). The multiple diversities of terpene carbon skeletons can be largely attributed to a great number of different TPSs [15]. The PFAM motif PF01397 (N-terminal TPS domain) [31] was used to search against the assembled W. ugandensis unigenes. As a result, a total of 16 unigenes were found to encode putative TPS, of which 9 unigenes were predicted to contain full-length coding sequences (S3 Table). To infer their possible functions and better describe the evolutionary relationships, phylogenetic analysis of the putative TPSs protein sequences and their homologs in other plants was performed, which indicated that they were divided into three main clades (Fig 5). In the first group, seven TPSs (WuSps1-7) are likely to be involved in sesquiterpene biosynthesis, which are clustered with two available W. ugandensis TPSs sequences (WargTPS-c, ACJ46047.1; WargTPS-g, ACJ46048.1) and a β-cubebene synthasefrom Magnolia grandiflora (ACC66281.1) in the same subclass. Seven unigenes (WuMts1-7) were found to be probably involved in monoterpene biosynthesis, and scattered into different subgroups. For example, four WuMts (WuMts1-4) are clustered with a Litsea cubeba thujene synthase (AEJ91555.1), while WuMts5 and WuMts7 forms a separate subgroup with several linalool- or nerolidol synthase homologs from other plants. The remaining two TPSs (WuDts1 and WuDts2) with full-length sequence were predicted to participate in diterpene biosynthesis, exhibiting closer relationships with several ent-copalyl diphosphate synthases and ent-kaurene synthases.


De Novo Transcriptome Analysis of Warburgia ugandensis to Identify Genes Involved in Terpenoids and Unsaturated Fatty Acids Biosynthesis.

Wang X, Zhou C, Yang X, Miao D, Zhang Y - PLoS ONE (2015)

Phylogenetic tree of terpene synthases.Phylogenetic analysis of 14 putative W. ugandensis WuTPS protein sequences with their homologs from other plants indicates that they are clustered into three main clades including: monoterpenoid synthase (WuMts), sesquiterpenoid synthase (WuSps), and diterpenoid synthase (WuDts). WarbTPS-c (ACJ46047.1, putative sesquiterpene synthase, W. ugandensis); WarbTPS-g (ACJ46048.1, putative sesquiterpene synthase, W. ugandensis); WuSps1, CL29873Contig1; WuSps2, CL29511Contig1; WuSps3, CL3178Contig1; WuSps4, CL4160Contig1; WuSps5, comp68897_c0_seq1; WuSps6, CL24969Contig1; WuSps7, CL30258Contig1; WuMts1, CL27268Contig1; WuMts2, CL27339Contig1; WuMts3, CL30385Contig1; WuMts4, CL276Contig2; WuMts5, CL1Contig9269; WuMts6, CL29966Contig1; WuMts7, CL14869Contig1; WuDts1, CL9128Contig1; WuDts2, CL28648Contig1; Citsi_Germacrene_D (XP_006494713.1, (-)-germacrene D synthase-like isoform X2, Citrus sinensis); Popeu_Valencene (XP_011015484.1, valencene synthase-like, Populus euphratica); Nelnu_Germacrene_D (XP_010258444.1, (-)-germacrene D synthase-like, Nelumbo nucifera); Eletr_Copaene (ADK94034.1, alpha-copaene synthase, Eleutherococcus trifoliatus); Gosar_Germacrene_D (KHG04103.1, (-)-germacrene D synthase, Gossypium arboretum); Vitvi_Germacrene_D (XP_010644711.1, (-)-germacrene D synthase, Vitis vinifera); Vitvi_Germacrene_A (ADR66821.1, Germacrene A synthase, Vitis vinifera); Citja_Elemene (BAP74389.1, delta-elemene synthase, Citrus jambhiri); Theca_Cadinene (EOY12648.1, Delta-cadinene synthase isozyme A, Theobroma cacao); Ricco_Cadinene (EEF38721.1, (+)-delta-cadinene synthase isozyme A, Ricinus communis); Aqusi_Guaiene (AIT75875.1, putative delta-guaiene synthase, Aquilaria sinensis); Vitvi_Caryophyllene (AEP17005.1, (E)-beta-caryophyllene synthase, Vitis vinifera); Maggr_Cubebene (ACC66281.1, beta-cubebene synthase, Magnolia grandiflora); Cinos_Linalool (AFK09265.1, S-(+)-linalool synthase, Cinnamomum osmophloeum); Nelnu_Nerolidol (XP_010248179.1, (3S,6E)-nerolidol synthase 1-like, Nelumbo nucifera); Vitvi_Nerolidol (XP_010646919.1, (3S,6E)-nerolidol synthase 1, chloroplastic-like isoform X1, Vitis vinifera); Vitvi_Linalool (ADR74212.1, (3S)-linalool/(E)-nerolidol synthase, Vitis vinifera); Actpo_Linalool (ADD81295.1, linalool synthase, Actinidia polygama); Nelnu_Ent-copalyl (XP_010277558.1, ent-copalyl diphosphate synthase, chloroplastic-like, Nelumbo nucifera); Theca_Ent-copalyl (XP_007050589.1, Copalyl diphosphate synthase, Theobroma cacao); Morno_Ent-copalyl (XP_010090409.1, Ent-copalyl diphosphate synthase, Morus notabilis); Gosar_Ent-copalyl (KHG01750.1, Ent-copalyl diphosphate synthase, chloroplastic, Gossypium arboreum); Nelnu_Ent-kaurene (XP_010260722.1, ent-kaur-16-ene synthase, chloroplastic isoform X1, Nelumbo nucifera); Phoda_Ent-kaurene (XP_008809130.1, ent-kaur-16-ene synthase, chloroplastic, Phoenix dactylifera); Ricico_Ent-kaurene (XP_002533694.1, Ent-kaurene synthase B, chloroplast precursor, Ricinus communis); Popeu_Ent-kaurene (XP_011014299.1, ent-kaur-16-ene synthase, chloroplastic, Populus euphratica); Nicta_Epi-Aristolochene (3M02.A, 5-Epi-Aristolochene Synthase, Nicotiana tabacum); Soltu_Vetispiradiene (Q9XJ32.1, vetispiradiene synthase 1, Solanum tuberosum); Litcu_Ocimene (AEJ91554.1, trans-ocimene synthase, Litsea cubeba); Litcu_Thujene (AEJ91555.1, alpha-thujene synthase, Litsea cubeba); Citli_Limonene (AAM53946.1, (+)-limonene synthase 2, Citrus limon); Vitvi_Ocimene/Myrcene (ADR74206.1, (E)-beta-ocimene/myrcene synthase, Vitis vinifera); Queil_Pinene (CAK55186.1, pinene synthase, Quercus ilex).
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pone.0135724.g005: Phylogenetic tree of terpene synthases.Phylogenetic analysis of 14 putative W. ugandensis WuTPS protein sequences with their homologs from other plants indicates that they are clustered into three main clades including: monoterpenoid synthase (WuMts), sesquiterpenoid synthase (WuSps), and diterpenoid synthase (WuDts). WarbTPS-c (ACJ46047.1, putative sesquiterpene synthase, W. ugandensis); WarbTPS-g (ACJ46048.1, putative sesquiterpene synthase, W. ugandensis); WuSps1, CL29873Contig1; WuSps2, CL29511Contig1; WuSps3, CL3178Contig1; WuSps4, CL4160Contig1; WuSps5, comp68897_c0_seq1; WuSps6, CL24969Contig1; WuSps7, CL30258Contig1; WuMts1, CL27268Contig1; WuMts2, CL27339Contig1; WuMts3, CL30385Contig1; WuMts4, CL276Contig2; WuMts5, CL1Contig9269; WuMts6, CL29966Contig1; WuMts7, CL14869Contig1; WuDts1, CL9128Contig1; WuDts2, CL28648Contig1; Citsi_Germacrene_D (XP_006494713.1, (-)-germacrene D synthase-like isoform X2, Citrus sinensis); Popeu_Valencene (XP_011015484.1, valencene synthase-like, Populus euphratica); Nelnu_Germacrene_D (XP_010258444.1, (-)-germacrene D synthase-like, Nelumbo nucifera); Eletr_Copaene (ADK94034.1, alpha-copaene synthase, Eleutherococcus trifoliatus); Gosar_Germacrene_D (KHG04103.1, (-)-germacrene D synthase, Gossypium arboretum); Vitvi_Germacrene_D (XP_010644711.1, (-)-germacrene D synthase, Vitis vinifera); Vitvi_Germacrene_A (ADR66821.1, Germacrene A synthase, Vitis vinifera); Citja_Elemene (BAP74389.1, delta-elemene synthase, Citrus jambhiri); Theca_Cadinene (EOY12648.1, Delta-cadinene synthase isozyme A, Theobroma cacao); Ricco_Cadinene (EEF38721.1, (+)-delta-cadinene synthase isozyme A, Ricinus communis); Aqusi_Guaiene (AIT75875.1, putative delta-guaiene synthase, Aquilaria sinensis); Vitvi_Caryophyllene (AEP17005.1, (E)-beta-caryophyllene synthase, Vitis vinifera); Maggr_Cubebene (ACC66281.1, beta-cubebene synthase, Magnolia grandiflora); Cinos_Linalool (AFK09265.1, S-(+)-linalool synthase, Cinnamomum osmophloeum); Nelnu_Nerolidol (XP_010248179.1, (3S,6E)-nerolidol synthase 1-like, Nelumbo nucifera); Vitvi_Nerolidol (XP_010646919.1, (3S,6E)-nerolidol synthase 1, chloroplastic-like isoform X1, Vitis vinifera); Vitvi_Linalool (ADR74212.1, (3S)-linalool/(E)-nerolidol synthase, Vitis vinifera); Actpo_Linalool (ADD81295.1, linalool synthase, Actinidia polygama); Nelnu_Ent-copalyl (XP_010277558.1, ent-copalyl diphosphate synthase, chloroplastic-like, Nelumbo nucifera); Theca_Ent-copalyl (XP_007050589.1, Copalyl diphosphate synthase, Theobroma cacao); Morno_Ent-copalyl (XP_010090409.1, Ent-copalyl diphosphate synthase, Morus notabilis); Gosar_Ent-copalyl (KHG01750.1, Ent-copalyl diphosphate synthase, chloroplastic, Gossypium arboreum); Nelnu_Ent-kaurene (XP_010260722.1, ent-kaur-16-ene synthase, chloroplastic isoform X1, Nelumbo nucifera); Phoda_Ent-kaurene (XP_008809130.1, ent-kaur-16-ene synthase, chloroplastic, Phoenix dactylifera); Ricico_Ent-kaurene (XP_002533694.1, Ent-kaurene synthase B, chloroplast precursor, Ricinus communis); Popeu_Ent-kaurene (XP_011014299.1, ent-kaur-16-ene synthase, chloroplastic, Populus euphratica); Nicta_Epi-Aristolochene (3M02.A, 5-Epi-Aristolochene Synthase, Nicotiana tabacum); Soltu_Vetispiradiene (Q9XJ32.1, vetispiradiene synthase 1, Solanum tuberosum); Litcu_Ocimene (AEJ91554.1, trans-ocimene synthase, Litsea cubeba); Litcu_Thujene (AEJ91555.1, alpha-thujene synthase, Litsea cubeba); Citli_Limonene (AAM53946.1, (+)-limonene synthase 2, Citrus limon); Vitvi_Ocimene/Myrcene (ADR74206.1, (E)-beta-ocimene/myrcene synthase, Vitis vinifera); Queil_Pinene (CAK55186.1, pinene synthase, Quercus ilex).
Mentions: TPSs are the primary enzymes responsible for catalyzing the conversion of prenyl diphosphates (GPP or FPP) into monoterpenes (C10), sesquiterpenes (C15), or diterpenes (C20). The multiple diversities of terpene carbon skeletons can be largely attributed to a great number of different TPSs [15]. The PFAM motif PF01397 (N-terminal TPS domain) [31] was used to search against the assembled W. ugandensis unigenes. As a result, a total of 16 unigenes were found to encode putative TPS, of which 9 unigenes were predicted to contain full-length coding sequences (S3 Table). To infer their possible functions and better describe the evolutionary relationships, phylogenetic analysis of the putative TPSs protein sequences and their homologs in other plants was performed, which indicated that they were divided into three main clades (Fig 5). In the first group, seven TPSs (WuSps1-7) are likely to be involved in sesquiterpene biosynthesis, which are clustered with two available W. ugandensis TPSs sequences (WargTPS-c, ACJ46047.1; WargTPS-g, ACJ46048.1) and a β-cubebene synthasefrom Magnolia grandiflora (ACC66281.1) in the same subclass. Seven unigenes (WuMts1-7) were found to be probably involved in monoterpene biosynthesis, and scattered into different subgroups. For example, four WuMts (WuMts1-4) are clustered with a Litsea cubeba thujene synthase (AEJ91555.1), while WuMts5 and WuMts7 forms a separate subgroup with several linalool- or nerolidol synthase homologs from other plants. The remaining two TPSs (WuDts1 and WuDts2) with full-length sequence were predicted to participate in diterpene biosynthesis, exhibiting closer relationships with several ent-copalyl diphosphate synthases and ent-kaurene synthases.

Bottom Line: In addition, the expression of 12 DEGs with putative roles in terpenoid and unsaturated fatty acid metabolic pathways was confirmed by qRT-PCRs, which was consistent with the data of the RNA-sequencing.In conclusion, we constructed a comprehensive transcriptome dataset derived from the bark and leaf of W. ugandensis, which forms the basis for functional genomics studies on this plant species.Particularly, the comparative analysis of the transcriptome data between the bark and leaf will provide critical clues to reveal the regulatory mechanisms underlying the biosynthesis of terpenoids and PUFAs in W. ugandensis.

View Article: PubMed Central - PubMed

Affiliation: CAS Key Laboratory of Plant Germplasm Enhancement and Specialty Agriculture, Wuhan Botanical Garden, Chinese Academy of Sciences, Wuhan, Hubei, China.

ABSTRACT
The bark of Warburgia ugandensis (Canellaceae family) has been used as a medicinal source for a long history in many African countries. The presence of diverse terpenoids and abundant polyunsaturated fatty acids (PUFAs) in this organ contributes to its broad range of pharmacological properties. Despite its medicinal and economic importance, the knowledge on the biosynthesis of terpenoid and unsaturated fatty acid in W. ugandensis bark remains largely unknown. Therefore, it is necessary to construct a genomic and/or transcriptomic database for the functional genomics study on W. ugandensis. The chemical profiles of terpenoids and fatty acids between the bark and leaves of W. ugandensis were compared by gas chromatography-mass spectrometry (GC-MS) analysis. Meanwhile, the transcriptome database derived from both tissues was created using Illumina sequencing technology. In total, about 17.1 G clean nucleotides were obtained, and de novo assembled into 72,591 unigenes, of which about 38.06% can be aligned to the NCBI non-redundant protein database. Many candidate genes in the biosynthetic pathways of terpenoids and unsaturated fatty acids were identified, including 14 unigenes for terpene synthases. Furthermore, 2,324 unigenes were discovered to be differentially expressed between both tissues; the functions of those differentially expressed genes (DEGs) were predicted by gene ontology enrichment and metabolic pathway enrichment analyses. In addition, the expression of 12 DEGs with putative roles in terpenoid and unsaturated fatty acid metabolic pathways was confirmed by qRT-PCRs, which was consistent with the data of the RNA-sequencing. In conclusion, we constructed a comprehensive transcriptome dataset derived from the bark and leaf of W. ugandensis, which forms the basis for functional genomics studies on this plant species. Particularly, the comparative analysis of the transcriptome data between the bark and leaf will provide critical clues to reveal the regulatory mechanisms underlying the biosynthesis of terpenoids and PUFAs in W. ugandensis.

No MeSH data available.