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杨树HSF家族基因生物信息学与胁迫应答表达分析

王雪怡 顾咏梅 张雪梅 姜廷波 刘焕臻

王雪怡, 顾咏梅, 张雪梅, 姜廷波, 刘焕臻. 杨树HSF家族基因生物信息学与胁迫应答表达分析[J]. 北京林业大学学报, 2021, 43(2): 34-45. doi: 10.12171/j.1000-1522.20200159
引用本文: 王雪怡, 顾咏梅, 张雪梅, 姜廷波, 刘焕臻. 杨树HSF家族基因生物信息学与胁迫应答表达分析[J]. 北京林业大学学报, 2021, 43(2): 34-45. doi: 10.12171/j.1000-1522.20200159
Wang Xueyi, Gu Yongmei, Zhang Xuemei, Jiang Tingbo, Liu Huanzhen. Bioinformatics and stress response expression analysis of poplar HSF family genes[J]. Journal of Beijing Forestry University, 2021, 43(2): 34-45. doi: 10.12171/j.1000-1522.20200159
Citation: Wang Xueyi, Gu Yongmei, Zhang Xuemei, Jiang Tingbo, Liu Huanzhen. Bioinformatics and stress response expression analysis of poplar HSF family genes[J]. Journal of Beijing Forestry University, 2021, 43(2): 34-45. doi: 10.12171/j.1000-1522.20200159

杨树HSF家族基因生物信息学与胁迫应答表达分析

doi: 10.12171/j.1000-1522.20200159
基金项目: 转基因生物新品种培育重大专项(2018ZX08020002)
详细信息
    作者简介:

    王雪怡。主要研究方向:林木抗性育种研究。Email:1368736882@qq.com 地址:150040 黑龙江省哈尔滨市香坊区和兴路26号东北林业大学林学院

    责任作者:

    刘焕臻,讲师。主要研究方向:林木抗性育种研究。Email:zhenzhen_0522@163.com 地址:同上

  • 中图分类号: S722.3+6

Bioinformatics and stress response expression analysis of poplar HSF family genes

  • 摘要:   目的  探究小黑杨热激转录因子HSF在应答高温和高盐胁迫时发挥的关键作用。保守结构域和顺式作用元件预测等对杨树HSF转录因子家族基因进行生物信息学分析。本研究以小黑杨为材料,经过37 ℃高温胁迫半个月后观察其形态变化;将小黑杨在37 ℃下分别处理0、12、24、48 h,采用RT-qPCR对小黑杨组织中的PsnHSFs基因进行时空表达分析;将小黑杨于150 mmol/L NaCl胁迫分别处理0、24 h,通过RNA-seq分析PsnHSFs基因的相对表达量变化,并通过RT-qPCR进行验证。  结果  通过结构特征和系统发育比较将29个HSF转录因子家族基因分成A、B和C三个亚家族,各亚家族分别包含18、10和1个HSF基因;HSF编码的氨基酸序列长度介于209 ~ 595之间,均为亲水性蛋白;其N端具有高度保守的DBD结构域,由三个保守基序构成;HSF基因启动子序列中包含DRE core、ABRE和TC-rich等多种顺式作用元件。小黑杨经37 ℃高温处理后其株高仅为对照的76.51%,叶片呈卷曲状,叶表面粗糙,叶面积显著减小且苗干多侧枝柔软无韧性。RT-qPCR与RNA-seq结果表明,PsnHSFs被高温、高盐胁迫诱导表达。高温处理后其株高仅为对照的76.51%,叶片呈卷曲状,叶表面粗糙,叶面积显著减小且苗干多侧枝柔软无韧性。RT-qPCR与RNA-seq结果表明,PsnHSFs被高温、高盐胁迫诱导表达。家族基因以及揭示HSF参与木本植物胁迫应答的分子机制调控具有参考意义。

     

  • 图  1  杨树与拟南芥HSF家族基因系统进化树

    Figure  1.  Phylogenetic tree of HSF family in poplar and Arabidopsis thaliana

    图  2  PtrHSFs结构域分析

    A. 杨树PtrHSFs基本结构图;B. 杨树PtrHSFs中DBD保守结构域;C. 杨树PtrHSFs氨基酸序列20个保守基序。A, basic structure of PtrHSFs of poplar; B, DBD conservative domain in PtrHSFs of poplar; C, there are 20 conserved amino acid sequences of PtrHSFs in poplar.

    Figure  2.  PtrHSFs domain analysis

    图  3  小黑杨高温胁迫生长状态

    A. 小黑杨非胁迫和胁迫生长状态;a为对照,b、c、d为37 ℃高温处理小黑杨;B. 小黑杨对照和高温胁迫处理半个月的株高。数据均为平均值 ± 标准差;星号表示胁迫处理植株与非胁迫处理植株之间的差异性,*、**、***分别表示在P < 0.05、P < 0.01、P < 0.001水平上差异显著。下同。A, non-stress and stress growth state of P. simonii × P. nigra; a, control; b, c, d, P. simonii × P. nigra trees under 37 ℃ high temperature treatment; B, plant height of P. simonii × P. nigra under control and HT for half a month. The data annotation in the picture is average value ± SD; the asterisk indicates that the difference between stressed and non-stressed plants. *, **, *** indicate significant difference at P < 0.05, P < 0.01, P < 0.001 level, respectively. The same below.

    Figure  3.  Growth state of P. simonii × P. nigra under high temperature stress

    图  4  小黑杨高温胁迫叶片状态

    A.形态学上端向下第3片到第11片小黑杨叶片;B.小黑杨第6片叶表面;a.对照;b、c、d. 37 ℃高温处理小黑杨。A, the 3rd to 11th leaves of P. simonii × P. nigra from apical side; B, surface of the 6th leaf of P. simonii × P. nigra; a, control; b, c, d, P. simonii × P. nigra under 37 ℃ high temperature treatment.

    Figure  4.  Leaf state of P. simonii × P. nigra under high temperature stress

    图  5  小黑杨PsnHSFs转录因子基因高温胁迫下相对表达量

    Figure  5.  Relative expression level of PsnHSFs transcription factor genes in P. simonii × P. nigra under high temperature stress

    图  6  小黑杨PsnHSFs应答盐胁迫热谱图分析

    C1 ~ C4. 对照组的4个生物学重复;N1 ~ N4. 处理组的4个生物学重复。C1−C4, four biological repeats in control; N1−N4, four biological repeats in treatment group.

    Figure  6.  Thermogram analysis of PsnHSFs genes responsing to salt stress of P. simonii × P. nigra

    图  7  小黑杨PsnHSFs基因应答盐胁迫定量分析

    Figure  7.  Quantitative analysis of PsnHSFs genes responsing to salt stress in P. simonii × P. nigra

    表  1  小黑杨PsnHSFs转录因子基因定量引物

    Table  1.   Quantitative primers for PsnHSFs transcription factor genes of Populus simonii × P. nigra

    基因名称 Gene name基因编号 Gene ID正向引物(5′—3′) Forward primer (5′−3′)反向引物(3′—5′) Reverse primer (3′−5′)
    PsnHSF1 Potri.001G108100.1 CTACGATGGCGGCATCAGCTG CAGCTGATGCCGCCATCGTAG
    PsnHSF2 Potri.001G138900.1 GCTTAGGACTATCAGTCGGCG CCTCTTAAGCCTCTCTACCTC
    PsnHSF3 Potri.001G273700.1 CCCAACACCATCACCAGCACT GATTATCTTCTGAGAGTGCAG
    PsnHSF4 Potri.001G320900.1 CCTACTTTTGTTGAACACCTT TACTGTGATTTTCCACAAGAC
    PsnHSF5 Potri.002G048200.1 ATGAATCCATATCTAACAGTG GTATCATGTAAACCTTCCATC
    PsnHSF6 Potri.002G124800.1 CAGCTCAGCCACAAGTAGCCA GTGAATCACACCAGTTGTTGG
    PsnHSF7 Potri.003G095000.1 GCTTAGGATTATCAGTCGGCG CTTCCTCGAGCCCAAATTTCC
    PsnHSF8 Potri.004G042600.1 GCGAGCCCAGCACGTTTCCAG CTAGTTCCATTTTCTCTCTTC
    PsnHSF9 Potri.004G062300.1 GTATCTTTTGTGACCCGAGTG GAAGTCACCGTCTGATTATCC
    PsnHSF10 Potri.005G214800.1 GATCTAGTAGAGGTGGTGGTG CATACCCTTGCATTTCCAACG
    PsnHSF11 Potri.006G049200.1 GGACTAACAAGCAACAACCTA CTAGTAAGCTCAGTGCTCAAG
    PsnHSF12 Potri.006G115700.1 GGAGAGATTGGTTCATCAAGG GTGATGTTTCTCCAATCAGGC
    PsnHSF13 Potri.006G148200.1 ATGAATACAAGAGAAATGCAG CTTCAACTCATTGAGACTATC
    PsnHSF14 Potri.006G226800.1 GACATCTCTCACAGACCACAC CTAAGCCTTACAATCTCTGCC
    PsnHSF15 Potri.007G043800.1 GACAGCGGCTGCGTCTCCGAC GACGTTGACGTGGACCCCAGG
    PsnHSF16 Potri.008G157600.1 CATCTTCTCAAGAGTATTAGG CTTGCTTGTCTCGCCTTAATC
    PsnHSF17 Potri.009G068000.1 CAATATCCCAGCACCATCACC CTGAGAGGGCAGTTAGGATAT
    PsnHSF18 Potri.010G082000.1 CTCCTCAAACTCAGACTTCTC CTTCACTAGTTCCACCATTAG
    PsnHSF19 Potri.010G104300.1 CTCAGGGCACAGACAATCGAA CTCAACTTCCTTCCAGAGACC
    PsnHSF20 Potri.011G051600.1 GTGAGCCTAGTATGTTTCCAG CAGTTCAAGTTTCTCTCTTCG
    PsnHSF21 Potri.011G071700.1 CATACAGCAAACTATAGTGTC GTATAGATAACCAATTCTAGG
    PsnHSF22 Potri.012G138900.1 GCCATGGTGAAAACGTCGTCG CTCTTCATCTCCGTCAACTCC
    PsnHSF23 Potri.013G079800.1 CTGTTCATGGTAACCTACCAC TCTAACAAGTTCCTGCATGAG
    PsnHSF24 Potri.014G027100.1 CAGCTCAGCCACAAGTAGCTA GAGAGATGCTGGTTCTGCTTG
    PsnHSF25 Potri.014G141400.1 GTTCTCCAATTGCAAACCTGG CAACATGGCACACTTCTTCAC
    PsnHSF26 Potri.015G141100.1 GTGGCGACGGTGGCGCAAGTG CCGGCGAAGAACTCCTCGTCG
    PsnHSF27 Potri.016G056500.1 CATGGTCTAGCAAGCAACAAC GTAAGCTCAGTGCTCAAGACC
    PsnHSF28 Potri.017G059600.1 CTGCTTTTGTTGAACATCTTG AGAACTACAGTGATTTTCGAC
    PsnHSF29 Potri.T137400.1 CTGAAGAATATAGTTCGCAGG CATAATTATATCTTCATCATC
    下载: 导出CSV

    表  2  杨树HSF转录因子基因理化性质

    Table  2.   Physical and chemical properties of HSF transcription factor genes in poplar

    基因名称
    Gene name
    基因编号
    Gene ID
    基因群
    Gene
    group
    氨基酸个数
    Amino acid
    number
    连锁群
    Linkage
    group
    蛋白分子质量
    Molecular mass
    of protein/Da
    等电点
    Isoelectric
    point (PI)
    脂肪指数
    Aliphatic
    index (AI)
    不稳定系数
    Instability
    index (II)
    总平均亲水性
    Grand average of
    hydropathicity
    (GRAVY)
    PtrHSF1 Potri.001G108100.1 B2b 343 Chr01 36 826.6 4.71 69.15 55.66 −0.559
    PtrHSF2 Potri.001G138900.1 A1d 595 Chr01 65 369.3 5.51 74.57 64.08 −0.532
    PtrHSF3 Potri.001G273700.1 B4d 270 Chr01 31 292.3 7.15 70.37 56.72 −0.610
    PtrHSF4 Potri.001G320900.1 A5b 490 Chr01 54 702.7 6.01 70.67 56.61 −0.770
    PtrHSF5 Potri.002G048200.1 A7b 359 Chr02 41 148.3 5.13 67.41 57.79 −0.808
    PtrHSF6 Potri.002G124800.1 B4a 364 Chr02 40 440.7 8.15 73.90 51.74 −0.468
    PtrHSF7 Potri.003G095000.1 A1a 507 Chr03 55 694.9 4.61 70.81 58.55 −0.601
    PtrHSF8 Potri.004G042600.1 A1 209 Chr04 24 028.4 9.50 70.53 57.40 −0.655
    PtrHSF9 Potri.004G062300.1 A4a 407 Chr04 46 642.1 4.91 62.04 57.47 −0.843
    PtrHSF10 Potri.005G214800.1 A7a 359 Chr05 40 692.4 5.37 61.42 66.48 −0.914
    PtrHSF11 Potri.006G049200.1 B3 226 Chr06 26 377.0 9.06 76.77 51.80 −0.753
    PtrHSF12 Potri.006G115700.1 A3 444 Chr06 49 881.0 4.86 66.94 61.80 −0.634
    PtrHSF13 Potri.006G148200.1 A9 430 Chr06 48 496.8 5.40 73.23 51.38 −0.626
    PtrHSF14 Potri.006G226800.1 A2 388 Chr06 43 856.9 4.70 73.30 56.80 −0.592
    PtrHSF15 Potri.007G043800.1 B1 285 Chr07 31 018.9 4.57 59.58 37.61 −0.853
    PtrHSF16 Potri.008G157600.1 A6b 348 Chr08 40 084.0 4.90 75.89 57.84 −0.692
    PtrHSF17 Potri.009G068000.1 B4b 272 Chr09 31 530.7 7.19 72.68 57.32 −0.586
    PtrHSF18 Potri.010G082000.1 A6a 358 Chr10 41 335.3 4.90 67.21 55.43 −0.788
    PtrHSF19 Potri.010G104300.1 A8b 392 Chr10 44 690.9 4.42 70.61 43.14 −0.705
    PtrHSF20 Potri.011G051600.1 A1 211 Chr11 24 383.6 9.79 63.84 53.41 −0.772
    PtrHSF21 Potri.011G071700.1 A4a 406 Chr11 46 266.6 5.02 66.28 54.95 −0.793
    PtrHSF22 Potri.012G138900.1 B2a 301 Chr12 33 366.2 4.77 67.94 50.83 −0.667
    PtrHSF23 Potri.013G079800.1 A1b 499 Chr13 55 091.5 5.62 65.03 57.66 −0.619
    PtrHSF24 Potri.014G027100.1 B4c 368 Chr14 41 038.2 8.17 68.10 52.12 −0.564
    PtrHSF25 Potri.014G141400.1 A4b 443 Chr14 50 781.7 6.17 65.15 62.50 −0.806
    PtrHSF26 Potri.015G141100.1 B2c 286 Chr15 31 751.8 4.86 80.66 50.80 −0.470
    PtrHSF27 Potri.016G056500.1 B3b 228 Chr16 26 486.0 7.30 71.84 55.59 −0.732
    PtrHSF28 Potri.017G059600.1 A5a 485 Chr17 54 386.4 5.84 69.01 57.90 −0.752
    PtrHSF29 Potri.T137400.1 C1 339 Scaffold-294 37 981.9 5.27 77.05 51.81 −0.409
    下载: 导出CSV

    表  3  杨树HSF家族基因启动子顺式作用元件

    Table  3.   Cis-acting elements of HSF family gene promoter in poplar

    作用元件 Acting element序列 Sequence功能 Function
    Gap-box CAAATGAA(A/G)A 光响应元件的一部分 Part of light responsive element
    Box4 ATTAAT 与光相关的保守DNA模块的一部分 Part of a conserved DNA module involved in light
    DRE core GCCGAC 脱水反应元件 Dehydration responsive element
    MYC CATTTG 参与低温反应 Involved in chilling response
    MYB CAACCA MYB 结合位点 MYB binding site
    ARE AAACCA 无氧诱导所必需的 Essential for anaerobic induction
    ABRE ACGTG 参与脱落酸反应 Involved in abscisic acid responsiveness
    TGACG-motif TGACG 参与了MeJA反应 Involved in MeJA-responsiveness
    TC-rich repeats ATTCTCTAAC 参与防御和压力反应 Involved in defense and stress responsiveness
    TCA-element CCATCTTTTT 参与水杨酸反应 Involved in salicylic acid responsiveness
    LTR CCGAAA 参与低温反应 Involved in low-temperature responsiveness
    MBS CAACTG 参与干旱诱导 Involved in drought-inducibility
    MBS І aaaAaaC(G/C)GTTA 参与类黄酮生物合成基因调控 Involved in flavonoid biosynthetic gene regulation
    GARE-motif TCTGTTG 赤霉素反应元件 Gibberellins-responsive element
    AE-box AGAAACTT 光响应模块的一部分 Part of a module for light response
    WUN-motif AAATTACT 创伤反应元件 Wound-responsive element
    P-box CCTTTTG 赤霉素反应元件 Gibberellin-responsive element
    TGA-box 生长素反应元件的一部分 Part of an auxin-responsive element
    ERE ATTTTAAA 乙烯反应元件 Ethylene-responsive element
    G-box CACGTC/TACGTG 光响应性和结合其他特定压力调节的元件
    Light responsiveness and combines with other regulatory elements under specific stress
    下载: 导出CSV
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出版历程
  • 收稿日期:  2020-05-25
  • 修回日期:  2020-07-01
  • 网络出版日期:  2021-01-29
  • 刊出日期:  2021-02-24

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