No abstract

Drought stress impacts seedling establishment, survival and whole-plant productivity. Molecular responses to drought stress have been most extensively studied in herbaceous species, mostly considering only aboveground tissues. Coniferous tree species dominate boreal forests, which are predicted to be exposed to more frequent and acute drought as a result of ongoing climate change. The associated impact at all stages of the forest tree life cycle is expected to have large-scale ecological and economic impacts. However, the molecular response to drought has not been comprehensively profiled for coniferous species. We assayed the physiological and transcriptional response of Picea abies (L.) H. Karst seedling needles and roots after exposure to mild and severe drought. Shoots and needles showed extensive reversible plasticity for physiological measures indicative of drought response mechanisms, including changes in stomatal conductance (gs), shoot water potential and ABA (abscisic acid). In both tissues the most commonly observed expression profiles in response to drought were highly correlated with ABA levels. Still, root and needle transcriptional responses contrasted, with extensive root-specific downregulation of growth. Comparison between previously characterized A. thaliana drought-response genes and P. abies revealed both conservation and divergence of transcriptional response to drought. In P. abies, transcription factors belonging to the bZIP AREB/ABF (ABA Response Element Binding/ABRE Binding Factors) ABA-dependent pathway had a more limited role. These results highlight the importance of profiling both above- and below-ground tissues and provide a comprehensive framework to advance understanding of the drought response of P. abies. The results demonstrate that short term, severe drought induces severe physiological responses coupled to extensive transcriptome modulation and highlight the susceptibility of Norway spruce seedlings to such drought events.

One sentence summary: Analysis of the drought transcriptome of Norway spruce 31 reveals divergent molecular response pathways in conifers. 32 33 Abstract 34 Drought stress impacts on seedling establishment, survival and whole-plant 35 productivity. Drought stress responses have been extensively studied at the 36 physiological and molecular level in angiosperms, particularly in agricultural species 37 and the model Arabidopsis thaliana, with the vast majority of work performed on 38 aboveground tissues. Boreal forests are dominated by coniferous tree species and 39cover vast areas of the terrestrial surface. These areas are predicted to be particularly 40 influenced by ongoing climate change and will be exposed to more frequent and acute 41 drought. The associated impact at all stages of the forest tree life cycle is expected to 42 have large-scale ecological and economic impacts. To provide a comprehensive 43understanding of the drought response mechanisms of Picea abies seedlings, we 44 assayed the physiological response of needles and transcriptional responses of roots 45 and needles after exposure to mild and severe drought. Shoots and needles showed 46 extensive reversible plasticity for physiological measures indicative of drought 47 response mechanisms, including stomatal conductance (g s ) and shoot water potential. 48Root and needle transcriptional responses contrasted, with an extensive root-specific 49 down-regulation of growth. When we compared the responses of P. abies with 50 previously-characterised A. thaliana drought response genes, we found that the 51 majority of the genes were conserved across lineages. However, in P. abies, 52 transcription factors (TFs) previously identified as belonging to the ABA-dependent 53 pathway had a more limited role and most differentially expressed genes were 54 specific to the stress response of P. abies. These results highlight the importance of 55 profiling both above-and below-ground tissues and provide a comprehensive 56 framework to advance understanding of the drought response mechanism of P. abies. 57 58

Cold acclimation in plants is a complex phenomenon involving numerous stress-responsive transcriptional and metabolic pathways. Existing gene expression studies have primarily addressed cold acclimation responses in herbaceous plants, and few have focused on perennial evergreens, such as conifers, that survive extremely low temperatures during winter. Relative to Arabidopsis leaves, the main transcriptional response of Norway spruce (Picea abies (L.) H. Karst) needles exposed to cold was delayed, and this delay was associated with slower development of freezing tolerance. Despite this difference in timing, our results indicate that, similar to herbaceous species, Norway spruce principally utilizes early response transcription factors (TFs) of the APETALA 2/ethylene-responsive element binding factor (AP2/ERF) superfamily and NAM (no apical meristem)/ATAF (Arabidopsis Transcription Factors)/CUC (cup shaped cotyledon) (NACs). The needles and root of Norway spruce showed contrasting results, in keeping with their different metabolic and developmental states. Regulatory network analysis identified conserved TFs, including a root-specific bHLH101 homolog, and other members of the same TF family with a pervasive role in cold regulation, such as homologs of ICE1 and AKS3, and also homologs of the NAC (anac47 and anac28) and AP2/ERF superfamilies (DREB2 and ERF3), providing new functional insights into cold stress response strategies in Norway spruce.

Climate change in the conifer‐dominated boreal forest is expected to lead to warmer but more dynamic winter air temperatures, reducing the depth and duration of snow cover and lowering winter soil temperatures. To gain insight into the mechanisms that have enabled conifers to dominate extreme cold environments, we performed genome‐wide RNA‐Seq analysis from needles and roots of non‐dormant two‐year Norway spruce (Picea abies (L.) H. Karst), and contrasted these response to herbaceous model Arabidopsis We show that the main transcriptional response of Norway spruce needles exposed to cold was delayed relative to Arabidopsis, and this delay was associated with slower development of freezing tolerance. Despite this difference in timing, Norway spruce principally utilizes early response transcription factors (TFs) belonging to the same gene families as Arabidopsis, indicating broad evolutionary conservation of cold response networks. In keeping with their different metabolic and developmental states, needles and root of Norway spruce showed contrasting results. Regulatory network analysis identified both conserved TFs with known roles in cold acclimation (e.g. homologs of ICE1, AKS3, and of the NAC and AP2/ERF superfamilies), but also a root‐specific bHLH101 homolog, providing functional insights into cold stress response strategies in Norway spruce.

SummaryCold acclimation in plants is a complex phenomenon involving numerous stress-responsive transcriptional and metabolic pathways. Existing gene expression studies have primarily addressed short-term cold acclimation responses in herbaceous plants, while few have focused on perennial evergreens, such as conifers, that survive extremely low temperatures during winter. To characterize the transcriptome changes during cold acclimation in Picea abies (L.) H. Karst (Norway spruce), we performed RNA-Sequencing analysis of needles and roots subjected to a chilling progression (5 °C) followed by 10 days at freezing temperature (−5 °C). Comparing gene expression responses of needles against Arabidopsis thaliana L. (Arabidopsis) leaves, our results showed that early transient inductions were observed in both species but the transcriptional response of Norway spruce was delayed. Our results indicate that, similar to herbaceous species, Norway spruce principally utilizes early response transcription factors (TFs) that belong to the APETALA 2/ethylene-responsive element binding factor (AP2/ERF) superfamily and NACs. However, unique to the Norway spruce response was a large group of TFs that mounted a late transcriptional response to low temperature. A predicted regulatory network analysis identified key conserved TFs, including a root-specific bHLH101 homolog and other members of the same family with a pervasive role in cold regulation, such as homologs of ICE1 and AKS3 and also homologs of the NAC (anac47 and anac28) and AP2/ERF superfamilies (DREB2 and ERF3), providing new functional insights into cold stress response strategies in Norway spruce.One sentence summaryNorway spruce shares elements of the cold regulon described in herbaceous species but has undescribed components that contribute to the cold tolerance of this evergreen coniferous species.