1. Heat Stress (HS) in Blueberry Final report 11 April, 2013

2. cyclic nucleotide-gated calcium channel Arabidopsis has two paralogs (CNG2 and CNG4) Temperature Sensor Andrija Finka, America Farinia Henriquez Cuendet, Frans J.M. Maathuis, Younousse Saidi, and Pierre Goloubinoffa.. Plasma Membrane Cyclic Nucleotide Gated Calcium Channels Control Land Plant Thermal Sensing and Acquired Thermotolerance. The Plant Cell, Vol. 24: 3333–3348, August 2012.

3. Conserved Family of Genes

4. Deletions Grow Slower 20° C Andrija Finka, America Farinia Henriquez Cuendet, Frans J.M. Maathuis, Younousse Saidi, and Pierre Goloubinoffa.. Plasma Membrane Cyclic Nucleotide Gated Calcium Channels Control Land Plant Thermal Sensing and Acquired Thermotolerance. The Plant Cell, Vol. 24: 3333–3348, August 2012. ArabidopsisMoss WT CNGC2 -/-

5. CNGC2 -/- Hyper-Thermosensitive Moss HSP promoter + Reporter

6. More Calcium Enters CNGC2 -/- Moss WTCNGC2 -/-

7. 3 Temp-Sensitive Ca 2+ Channels Moss conductance in picosiemens (1 – 100 range) WT CNGC2 -/-

8. Prime Plants for Thermotolerance Arabidopsis WT CNGC2 -/-

9. Full HSR at Lower Temps Arabidopsis WT CNGC2 -/-

10. Survive Extreme Heat Treatment WTCNGC2 -/- Arabidopsis

11. Post Temperature Sensor 1.calcium channel in PM (primary pathway) 2.histone sensor in the nucleus 3.unfolded protein sensor (UPR) in the endoplasmic reticulum 4.unfolded protein sensor (UPR) in the cytosol

12. tolerant NCED4 allele is responsible for germination at high temperatures in lettuce encodes 9-cis-epoxycarotenoid dioxygenase 4 (1.13.11.51) Catalyzes the first step of abscisic-acid biosynthesis from carotenoids in chloroplasts, in response to water stress Seed Germination

13. NCED4 Determines Heat Inhibition wt NCED4 inhib NCED4 + RNAi tolerant NCED4 heat tolerant seeds heat inhibited seeds Heqiang Huo, Peetambar Dahal, Keshavulu Kunusoth, Claire M. McCallum, and Kent J. Bradford.2013 ePublication. Expression of 9-cis-EPOXYCAROTENOID DIOXYGENASE4 Is Essential for Thermoinhibition of Lettuce Seed Germination but Not for Seed Development or Stress Tolerance. www.plantcell.org/cgi/doi/10.1105/tpc.112.108902.www.plantcell.org/cgi/doi/10.1105/tpc.112.108902

14. NCED4 Determines Heat Inhibition wt NCED4 inhib NCED4 + RNAi tolerant NCED4 heat tolerant seeds heat inhibited seeds Heqiang Huo, Peetambar Dahal, Keshavulu Kunusoth, Claire M. McCallum, and Kent J. Bradford.2013 ePublication. Expression of 9-cis-EPOXYCAROTENOID DIOXYGENASE4 Is Essential for Thermoinhibition of Lettuce Seed Germination but Not for Seed Development or Stress Tolerance. www.plantcell.org/cgi/doi/10.1105/tpc.112.108902.www.plantcell.org/cgi/doi/10.1105/tpc.112.108902

15. HS Pathway reverts when cooled and HsfB antagonism wins HS response requires heterotrimer of HsfA+B+C HsfA1 (family) is the master regulator

16. HsfA1-independent Pathway heat may be able to activate HS response without HsfA1

17. genes involved in thermotolerance

18. List of Genes with 51 SSRs Primer Pairs xn WRKY 3scaffold00003 x 3 RBOHD & Escaffold00029 x 3 BIP1 scaffold00039 x3 MBF1 scaffold00048 x3 H2A.Z aka HTA11scaffold00069 x4 CNGC2 & CNGC4scaffold00079 x3 CAT1 scaffold00093 x3 HSFA2 scaffold00109 x3 ETR1 scaffold00112 x3 FAD6 scaffold00117 x3 HSFB1scaffold00122 x3 FAD3 scaffold00126 x3 PLD scaffold00152 x3 ABI3 scaffold00173 x3 UVH6 scaffold00209 x3 CAD2 scaffold00210 x3 VTC2 scaffold00252 x3 NPQ1 scaffold00260 x3 ABI1 scaffold00338 x3 RBOHA & C & G & Hscaffold02317 x2 FAD5 scaffold00455 x3 ACD2 scaffold00474 x3 RBOHB scaffold00539 x3 FAD4 scaffold00588 x3 BZIP28 scaffold00604 x3 BZIP17 scaffold00617 x3 IRE1 scaffold00646 x3 DREB scaffold00667 x3 MS1 scaffold00725 x3 BIP2 scaffold00773 x3 NCED4 scaffold00790 x2 VTC1 scaffold00867 x3 CAM3 scaffold00875 x3 CDPK 2scaffold00916 x3 ETO1scaffold00956 x3 AXR1 scaffold01037 x3 PIPK scaffold01039 x3 HSFA1 scaffold01062 x3 RBOHF scaffold01076 x1 UBP2 scaffold01164 x3 ABI2 scaffold01374 x3 HOT1 scaffold01438 x3 ETO2 & 3scaffold01451 x3 UVH3 scaffold01548 x2 UVH1 scaffold02430 x2 ETO1 #2scaffold01626 x2 FAD2 scaffold01825 x2 EIN2scaffold02031 CD UBP1 scaffold02114 x2 FAD7 scaffold02449 x1 UTR3 scaffold02795 x1 BZIP60 no scaffold NPR1 scaffold12427 x0

19. References Andrija Finka et al. 2012. Plasma Membrane Cyclic Nucleotide Gated Calcium Channels Control Land Plant Thermal Sensing and Acquired Thermotolerance. The Plant Cell. Vol. 24: 3333–3348. Heqiang Huo et al.2013 ePublication. Expression of 9-cis- EPOXYCAROTENOID DIOXYGENASE4 Is Essential for Thermoinhibition of Lettuce Seed Germination but Not for Seed Development or Stress Tolerance. www.plantcell.org/cgi/doi/10.1105/tpc.112.108902. www.plantcell.org/cgi/doi/10.1105/tpc.112.108902 Jane Larkindale et al. 2005. Heat Stress Phenotypes of Arabidopsis Mutants Implicate Multiple Signaling Pathways in the Acquisition of Thermotolerance. Plant Physiology. Vol. 138: 882– 897. Sachin Kotak et al. 2007. A Novel Transcriptional Cascade Regulating Expression of Heat Stress Proteins duringSeedDevelopmentof Arabidopsis. The Plant Cell, Vol. 19: 182–195. Ching-Hui Yeha et al. 2012. Some like it hot, some like it warm: Phenotyping to explore thermotolerance diversity. Plant Science. Vol. 195: 10-23. Yong-Xiang Lin et al. 2011. Genome-wide identification, classification and analysis of heat shock transcription factor family in maize. BMC Genomics. Vol. 12: 76. Hsiang-chin Liu and Yee-yung Charng. 2012. Acquired thermotolerance independent of heat shock factor A1 (HsfA1), the master regulator of the heat stress response. Plant Signaling & Behavior 7(5): 547–550. Jason Argyris, et al. 2011. A gene encoding an abscisic acid biosynthetic enzyme (LsNCED4) collocates with the high temperature germination locus Htg6.1 in lettuce (Lactuca sp.) Theor Appl Genet. Vol. 122:95–108. Younousse Saidi, et al. 2009. The Heat Shock Response in Moss Plants Is Regulated by Specific Calcium-Permeable Channels in the Plasma Membrane. The Plant Cell. Vol. 21: 2829–2843. Ondrej Krinke, et al. 2009. Phospholipase D Activation Is an Early Component of the Salicylic Acid Signaling Pathway in Arabidopsis Cell Suspensions. Plant Physiology. Vol. 150, pp. 424–436.