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Responses to winter dormancy, temperature, and plant hormones share gene networks

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Abstract

Gene networks modulated in winter dormancy (WD) in relation to temperature and hormone responses were analyzed in tea [Camellia sinensis (L.) O. Kuntze]. Analysis of subtracted cDNA libraries prepared using the RNA isolated from the apical bud and the associated two leaves (two and a bud, TAB) of actively growing (AG) and winter dormant plant showed the downregulation of genes involved in cell cycle/cell division and upregulation of stress-inducible genes including those encoding chaperons during WD. Low temperature (4°C) modulated gene expression in AG cut-shoots in similar fashion as observed in TAB during WD. In tissue harvested during WD, growth temperature (25°C) modulated gene expression in the similar way as observed during the period of active growth (PAG). Abscisic acid (ABA) and gibberellic acid (GA3) modulated expression of selected genes, depending upon if the tissue was harvested during PAG or WD. Tissue preparedness was critical for ABA- and GA3-mediated response, particularly for stress-responsive genes/chaperons. Data identified the common gene networks for winter dormancy, temperature, and plant hormone responses.

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Acknowledgments

The authors thank Dr. P. S. Ahuja, Director, IHBT, for encouragement and providing necessary facilities. Authors acknowledge Council of Scientific and Industrial Research (CSIR) for providing financial assistance under Major Lab Project. The project was also funded through grant-in-aid projects by the National Tea Research Foundation, India, and the Department of Biotechnology, Government of India. AP is grateful to CSIR for awarding Junior and Senior Research Fellowships. Authors thank Mr. Digvijay Singh Naruka and Ms. Anupama Saini for their help in sequencing and plasmid isolation. MS represents IHBT publication number 1030.

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Authors and Affiliations

  1. Biotechnology Division, Council of Scientific and Industrial Research-Institute of Himalayan Bioresource Technology, Palampur, Himachal Pradesh, 176061, India

    Asosii Paul & Sanjay Kumar

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  1. Asosii Paul
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  2. Sanjay Kumar
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Correspondence to Sanjay Kumar.

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Supplementary Fig. S1

Close-up view of tea bushes (a) in the field during the period of active growth (PAG), winter dormancy (WD), and dormancy release (DR). Mean maximum and minimum air temperatures for two consecutive years are shown in (b). Data for air temperature were procured from the neighboring agriculture university (Chaudhary Sarwan Kumar Himachal Pradesh Krishi Vishvavidyalaya, Palampur, India). Observatory was within 3 km of the tea farm in the university campus. Each value is a mean of 1 month data with ±SE (JPEG 124 kb)

Supplementary Fig. S2

Functional classification of clones obtained in forward- (FL) and reverse- (RL)-subtracted libraries. FL and RL represent the genes upregulated during winter dormancy (WD) and the period of active growth (PAG), respectively. The clones were grouped into functional classes based on the MIPS classification system (Ruepp et al. 2004 (JPEG 106 kb)

Supplementary Fig. S3

Slot blot (reverse northern) analysis for differential screening. The inserts were PCR amplified and blotted on nylon membrane in duplicate and hybridized with radio-labeled probe synthesized from unsubtracted driver cDNA and tester cDNA seperately. (a) Genes represented in the forward library (FL), (b) Genes represented in the reverse library (RL) (DOCX 1614 kb)

Supplementary Fig. S4

Gene expression during the period of active growth (PAG), winter dormancy (WD) and dormancy release (DR) for two consecutive years in TAB (apical bud and the associated two leaves) and ML (mature leaves) harvested from field-grown tea bushes. 26S rRNA gene was used as control. Name of the gene is shown on right side of the panel (DOC 169 kb)

Supplementary Fig. S5

Effect of abscisic acid (100 μM ABA; b) and gibberellic acid (100 μM GA3; c) on gene expression in TAB harvested during period of active growth (PAG) and winter dormancy (WD) and placed at two temperatures [4 ± 2°C (low temperature, LT) and 25 ± 3°C (growth temperature, GT)]. Experiment without exogenous hormone was also set up for the two temperatures (a). Experiment was carried out over a period of 96 hrs and the analysis was performed at different time points as shown above the panel. Numeral above the lane indicates the hour of the treatment. 26S rRNA gene was used as an internal control for equal loading. Name of the gene is shown above the panel (DOC 613 kb)

Supplementary Table S1

Primer sequences and PCR conditions used in RT-PCR analysis (DOCX 15 kb)

Supplementary Table S2

List of genes classified by putative function in the forward library (FL) along with their annotation, size of insert, redundancy and Blast X score values. FL represents the genes upregulated during winter dormancy. Genes were grouped into functional classes based on the MIPS classification system (Ruepp et al. 2004 (XLS 75 kb)

Supplementary Table S3

List of genes classified by putative function in the reverse library (RL) along with their annotation, size of insert, redundancy, and Blast X values. RL represents the genes upregulated during the period of active growth. Genes were grouped into functional classes based on the MIPS classification system (Ruepp et al. 2004 (XLS 134 kb)

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Paul, A., Kumar, S. Responses to winter dormancy, temperature, and plant hormones share gene networks. Funct Integr Genomics 11, 659–664 (2011). https://doi.org/10.1007/s10142-011-0233-4

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