Massive production of small RNAs from a non-coding region of Cauliflower mosaic virus in plant defense and viral counter-defense

To successfully infect plants, viruses must counteract small RNA-based host defense responses. During infection of Arabidopsis, Cauliflower mosaic pararetrovirus (CaMV) is transcribed into pregenomic 35S and subgenomic 19S RNAs. The 35S RNA is both reverse transcribed and also used as an mRNA with highly structured 600 nt leader. We found that this leader region is transcribed into long sense- and antisense-RNAs and spawns a massive quantity of 21, 22 and 24 nt viral small RNAs (vsRNAs), comparable to the entire complement of host-encoded small-interfering RNAs and microRNAs. Leader-derived vsRNAs were detected bound to the Argonaute 1 (AGO1) effector protein, unlike vsRNAs from other viral regions. Only negligible amounts of leader-derived vsRNAs were bound to AGO4. Genetic evidence showed that all four Dicer-like (DCL) proteins mediate vsRNA biogenesis, whereas the RNA polymerases Pol IV, Pol V, RDR1, RDR2 and RDR6 are not required for this process. Surprisingly, CaMV titers were not increased in dcl1/2/3/4 quadruple mutants that accumulate only residual amounts of vsRNAs. Ectopic expression of CaMV leader vsRNAs from an attenuated geminivirus led to increased accumulation of this chimeric virus. Thus, massive production of leader-derived vsRNAs does not restrict viral replication but may serve as a decoy diverting the silencing machinery from viral promoter and coding regions

Massive production of small RNAs from a non-coding region of Cauliflower mosaic virus in plant defense and viral counter-defense

To successfully infect plants, viruses must counteract small RNA-based host defense responses. During infection of Arabidopsis, Cauliflower mosaic pararetrovirus (CaMV) is transcribed into pregenomic 35S and subgenomic 19S RNAs. The 35S RNA is both reverse transcribed and also used as an mRNA with highly structured 600 nt leader. We found that this leader region is transcribed into long sense- and antisense-RNAs and spawns a massive quantity of 21, 22 and 24 nt viral small RNAs (vsRNAs), comparable to the entire complement of host-encoded small-interfering RNAs and microRNAs. Leader-derived vsRNAs were detected bound to the Argonaute 1 (AGO1) effector protein, unlike vsRNAs from other viral regions. Only negligible amounts of leader-derived vsRNAs were bound to AGO4. Genetic evidence showed that all four Dicer-like (DCL) proteins mediate vsRNA biogenesis, whereas the RNA polymerases Pol IV, Pol V, RDR1, RDR2 and RDR6 are not required for this process. Surprisingly, CaMV titers were not increased in dcl1/2/3/4 quadruple mutants that accumulate only residual amounts of vsRNAs. Ectopic expression of CaMV leader vsRNAs from an attenuated geminivirus led to increased accumulation of this chimeric virus. Thus, massive production of leader-derived vsRNAs does not restrict viral replication but may serve as a decoy diverting the silencing machinery from viral promoter and coding region

Short ORF-Dependent Ribosome Shunting Operates in an RNA Picorna-Like Virus and a DNA Pararetrovirus that Cause Rice Tungro Disease

Rice tungro disease is caused by synergistic interaction of an RNA picorna-like virus Rice tungro spherical virus (RTSV) and a DNA pararetrovirus Rice tungro bacilliform virus (RTBV). It is spread by insects owing to an RTSV-encoded transmission factor. RTBV has evolved a ribosome shunt mechanism to initiate translation of its pregenomic RNA having a long and highly structured leader. We found that a long leader of RTSV genomic RNA remarkably resembles the RTBV leader: both contain several short ORFs (sORFs) and potentially fold into a large stem-loop structure with the first sORF terminating in front of the stem basal helix. Using translation assays in rice protoplasts and wheat germ extracts, we show that, like in RTBV, both initiation and proper termination of the first sORF translation in front of the stem are required for shunt-mediated translation of a reporter ORF placed downstream of the RTSV leader. The base pairing that forms the basal helix is required for shunting, but its sequence can be varied. Shunt efficiency in RTSV is lower than in RTBV. But in addition to shunting the RTSV leader sequence allows relatively efficient linear ribosome migration, which also contributes to translation initiation downstream of the leader. We conclude that RTSV and RTBV have developed a similar, sORF-dependent shunt mechanism possibly to adapt to the host translation system and/or coordinate their life cycles. Given that sORF-dependent shunting also operates in a pararetrovirus Cauliflower mosaic virus and likely in other pararetroviruses that possess a conserved shunt configuration in their leaders it is tempting to propose that RTSV may have acquired shunt cis-elements from RTBV during their co-existence

Multilayer regulatory mechanisms control cleavage factor I proteins in filamentous fungi

Cleavage factor I (CFI) proteins are core components of the polyadenylation machinery that can regulate several steps of mRNA life cycle, including alternative polyadenylation, splicing, export and decay. Here, we describe the regulatory mechanisms that control two fungal CFI protein classes in Magnaporthe oryzae: Rbp35/CfI25 complex and Hrp1. Using mutational, genetic and biochemical studies we demonstrate that cellular concentration of CFI mRNAs is a limited indicator of their protein abundance. Our results suggest that several post-transcriptional mechanisms regulate Rbp35/CfI25 complex and Hrp1 in the rice blast fungus, some of which are also conserved in other ascomycetes. With respect to Rbp35, these include C-terminal processing, RGG-dependent localization and cleavage, C-terminal autoregulatory domain and regulation by an upstream open reading frame of Rbp35-dependent TOR signalling pathway. Our proteomic analyses suggest that Rbp35 regulates the levels of proteins involved in melanin and phenylpropanoids synthesis, among others. The drastic reduction of fungal CFI proteins in carbon-starved cells suggests that the pre-mRNA processing pathway is altered. Our findings uncover broad and multilayer regulatory mechanisms controlling fungal polyadenylation factors, which have profound implications in pre-mRNA maturation. This area of research offers new avenues for fungicide design by targeting fungal-specific proteins that globally affect thousands of mRNAs

A Genome-Wide RNAi Screen Identifies Regulators of Cholesterol-Modified Hedgehog Secretion in Drosophila

Hedgehog (Hh) proteins are secreted molecules that function as organizers in animal development. In addition to being palmitoylated, Hh is the only metazoan protein known to possess a covalently-linked cholesterol moiety. The absence of either modification severely disrupts the organization of numerous tissues during development. It is currently not known how lipid-modified Hh is secreted and released from producing cells. We have performed a genome-wide RNAi screen in Drosophila melanogaster cells to identify regulators of Hh secretion. We found that cholesterol-modified Hh secretion is strongly dependent on coat protein complex I (COPI) but not COPII vesicles, suggesting that cholesterol modification alters the movement of Hh through the early secretory pathway. We provide evidence that both proteolysis and cholesterol modification are necessary for the efficient trafficking of Hh through the ER and Golgi. Finally, we identified several putative regulators of protein secretion and demonstrate a role for some of these genes in Hh and Wingless (Wg) morphogen secretion in vivo. These data open new perspectives for studying how morphogen secretion is regulated, as well as provide insight into regulation of lipid-modified protein secretion

Genomic analysis of codon, sequence and structural conservation with selective biochemical-structure mapping reveals highly conserved and dynamic structures in rotavirus RNAs with potential cis-acting functions

Rotaviruses are a major cause of acute, often fatal, gastroenteritis in infants and young children world-wide. Virions contain an 11 segment double-stranded RNA genome. Little is known about the cis-acting sequences and structural elements of the viral RNAs. Using a database of 1621 full-length sequences of mammalian group A rotavirus RNA segments, we evaluated the codon, sequence and RNA structural conservation of the complete genome. Codon conservation regions were found in eight ORFs, suggesting the presence of functional RNA elements. Using ConStruct and RNAz programmes, we identified conserved secondary structures in the positive-sense RNAs including long-range interactions (LRIs) at the 5′ and 3′ terminal regions of all segments. In RNA9, two mutually exclusive structures were observed suggesting a switch mechanism between a conserved terminal LRI and an independent 3′ stem–loop structure. In RNA6, a conserved stem–loop was found in a region previously reported to have translation enhancement activity. Biochemical structural analysis of RNA11 confirmed the presence of terminal LRIs and two internal helices with high codon and sequence conservation. These extensive in silico and in vitro analyses provide evidence of the conservation, complexity, multi-functionality and dynamics of rotavirus RNA structures which likely influence RNA replication, translation and genome packaging

Regulation of translation initiation under biotic and abiotic stresses

[EN] Plants have developed versatile strategies to deal with the great variety of challenging conditions they are exposed to. Among them, the regulation of translation is a common target to finely modulate gene expression both under biotic and abiotic stress situations. Upon environmental challenges, translation is regulated to reduce the consumption of energy and to selectively synthesize proteins involved in the proper establishment of the tolerance response. In the case of viral infections, the situation is more complex, as viruses have evolved unconventional mechanisms to regulate translation in order to ensure the production of the viral encoded proteins using the plant machinery. Although the final purpose is different, in some cases, both plants and viruses share common mechanisms to modulate translation. In others, the mechanisms leading to the control of translation are viral- or stress-specific. In this paper, we review the different mechanisms involved in the regulation of translation initiation under virus infection and under environmental stress in plants. In addition, we describe the main features within the viral RNAs and the cellular mRNAs that promote their selective translation in plants undergoing biotic and abiotic stress situations.This work was supported by the ERC Starting Grant 260468 to M. Mar Castellano.Echevarria-Zomeno, S.; Yanguez, E.; Fernandez-Bautista, N.; Castro-Sanz, AB.; Ferrando Monleón, AR.; Castellano, MM. (2013). Regulation of translation initiation under biotic and abiotic stresses. International Journal of Molecular Sciences. 14(3):4670-4683. https://doi.org/10.3390/ijms14034670S46704683143Dever, T. E., & Green, R. (2012). The Elongation, Termination, and Recycling Phases of Translation in Eukaryotes. Cold Spring Harbor Perspectives in Biology, 4(7), a013706-a013706. doi:10.1101/cshperspect.a013706Sonenberg, N., & Hinnebusch, A. G. (2009). Regulation of Translation Initiation in Eukaryotes: Mechanisms and Biological Targets. Cell, 136(4), 731-745. doi:10.1016/j.cell.2009.01.042Graber, T. E., & Holcik, M. (2007). Cap-independent regulation of gene expression in apoptosis. Molecular BioSystems, 3(12), 825. doi:10.1039/b708867aAl-Fageeh, M. B., & Smales, C. M. (2006). Control and regulation of the cellular responses to cold shock: the responses in yeast and mammalian systems. Biochemical Journal, 397(2), 247-259. doi:10.1042/bj20060166Braunstein, S., Karpisheva, K., Pola, C., Goldberg, J., Hochman, T., Yee, H., … Schneider, R. J. (2007). A Hypoxia-Controlled Cap-Dependent to Cap-Independent Translation Switch in Breast Cancer. Molecular Cell, 28(3), 501-512. doi:10.1016/j.molcel.2007.10.019Castelli, L. M., Lui, J., Campbell, S. G., Rowe, W., Zeef, L. A. H., Holmes, L. E. A., … Ashe, M. P. (2011). Glucose depletion inhibits translation initiation via eIF4A loss and subsequent 48S preinitiation complex accumulation, while the pentose phosphate pathway is coordinately up-regulated. Molecular Biology of the Cell, 22(18), 3379-3393. doi:10.1091/mbc.e11-02-0153Gilbert, W. V., Zhou, K., Butler, T. K., & Doudna, J. A. (2007). Cap-Independent Translation Is Required for Starvation-Induced Differentiation in Yeast. Science, 317(5842), 1224-1227. doi:10.1126/science.1144467Liu, L., & Simon, M. C. (2004). Regulation of Transcription and Translation by Hypoxia. Cancer Biology & Therapy, 3(6), 492-497. doi:10.4161/cbt.3.6.1010Sun, J., Conn, C. S., Han, Y., Yeung, V., & Qian, S.-B. (2010). PI3K-mTORC1 Attenuates Stress Response by Inhibiting Cap-independent Hsp70 Translation. Journal of Biological Chemistry, 286(8), 6791-6800. doi:10.1074/jbc.m110.172882Walsh, D., Mathews, M. B., & Mohr, I. (2012). Tinkering with Translation: Protein Synthesis in Virus-Infected Cells. Cold Spring Harbor Perspectives in Biology, 5(1), a012351-a012351. doi:10.1101/cshperspect.a012351Floris, M., Mahgoub, H., Lanet, E., Robaglia, C., & Menand, B. (2009). Post-transcriptional Regulation of Gene Expression in Plants during Abiotic Stress. International Journal of Molecular Sciences, 10(7), 3168-3185. doi:10.3390/ijms10073168Jackson, R. J., Hellen, C. U. T., & Pestova, T. V. (2010). The mechanism of eukaryotic translation initiation and principles of its regulation. Nature Reviews Molecular Cell Biology, 11(2), 113-127. doi:10.1038/nrm2838Clemens, M. J. (2001). Translational regulation in cell stress and apoptosis. Roles of the eIF4E binding proteins. Journal of Cellular and Molecular Medicine, 5(3), 221-239. doi:10.1111/j.1582-4934.2001.tb00157.xWek, R. C., Jiang, H.-Y., & Anthony, T. G. (2006). Coping with stress: eIF2 kinases and translational control. Biochemical Society Transactions, 34(1), 7-11. doi:10.1042/bst0340007Holcik, M., & Sonenberg, N. (2005). Translational control in stress and apoptosis. Nature Reviews Molecular Cell Biology, 6(4), 318-327. doi:10.1038/nrm1618Muñoz, A., & Castellano, M. M. (2012). Regulation of Translation Initiation under Abiotic Stress Conditions in Plants: Is It a Conserved or Not so Conserved Process among Eukaryotes? Comparative and Functional Genomics, 2012, 1-8. doi:10.1155/2012/406357Hinnebusch, A. G. (2005). TRANSLATIONAL REGULATION OFGCN4AND THE GENERAL AMINO ACID CONTROL OF YEAST. Annual Review of Microbiology, 59(1), 407-450. doi:10.1146/annurev.micro.59.031805.133833Harding, H. P., Novoa, I., Zhang, Y., Zeng, H., Wek, R., Schapira, M., & Ron, D. (2000). Regulated Translation Initiation Controls Stress-Induced Gene Expression in Mammalian Cells. Molecular Cell, 6(5), 1099-1108. doi:10.1016/s1097-2765(00)00108-8Ventoso, I., Kochetov, A., Montaner, D., Dopazo, J., & Santoyo, J. (2012). Extensive Translatome Remodeling during ER Stress Response in Mammalian Cells. PLoS ONE, 7(5), e35915. doi:10.1371/journal.pone.0035915Sudhakar, A., Ramachandran, A., Ghosh, S., Hasnain, S. E., Kaufman, R. J., & Ramaiah, K. V. A. (2000). Phosphorylation of Serine 51 in Initiation Factor 2α (eIF2α) Promotes Complex Formation between eIF2α(P) and eIF2B and Causes Inhibition in the Guanine Nucleotide Exchange Activity of eIF2B†. Biochemistry, 39(42), 12929-12938. doi:10.1021/bi0008682García, M. A., Meurs, E. F., & Esteban, M. (2007). The dsRNA protein kinase PKR: Virus and cell control. Biochimie, 89(6-7), 799-811. doi:10.1016/j.biochi.2007.03.001Katze, M. G., He, Y., & Gale, M. (2002). Viruses and interferon: a fight for supremacy. Nature Reviews Immunology, 2(9), 675-687. doi:10.1038/nri888Mohr, I. (2006). Phosphorylation and dephosphorylation events that regulate viral mRNA translation. Virus Research, 119(1), 89-99. doi:10.1016/j.virusres.2005.10.009Zhang, Y., Wang, Y., Kanyuka, K., Parry, M. A. J., Powers, S. J., & Halford, N. G. (2008). GCN2-dependent phosphorylation of eukaryotic translation initiation factor-2α in Arabidopsis. Journal of Experimental Botany, 59(11), 3131-3141. doi:10.1093/jxb/ern169Lageix, S., Lanet, E., Pouch-Pélissier, M.-N., Espagnol, M.-C., Robaglia, C., Deragon, J.-M., & Pélissier, T. (2008). Arabidopsis eIF2α kinase GCN2 is essential for growth in stress conditions and is activated by wounding. BMC Plant Biology, 8(1), 134. doi:10.1186/1471-2229-8-134Bilgin, D. D., Liu, Y., Schiff, M., & Dinesh-Kumar, S. . (2003). P58IPK, a Plant Ortholog of Double-Stranded RNA-Dependent Protein Kinase PKR Inhibitor, Functions in Viral Pathogenesis. Developmental Cell, 4(5), 651-661. doi:10.1016/s1534-5807(03)00125-4Gallie, D. R., Le, H., Caldwell, C., Tanguay, R. L., Hoang, N. X., & Browning, K. S. (1997). The Phosphorylation State of Translation Initiation Factors Is Regulated Developmentally and following Heat Shock in Wheat. Journal of Biological Chemistry, 272(2), 1046-1053. doi:10.1074/jbc.272.2.1046Gingras, A. C., Svitkin, Y., Belsham, G. J., Pause, A., & Sonenberg, N. (1996). Activation of the translational suppressor 4E-BP1 following infection with encephalomyocarditis virus and poliovirus. Proceedings of the National Academy of Sciences, 93(11), 5578-5583. doi:10.1073/pnas.93.11.5578Gingras, A.-C., & Sonenberg, N. (1997). Adenovirus Infection Inactivates the Translational Inhibitors 4E-BP1 and 4E-BP2. Virology, 237(1), 182-186. doi:10.1006/viro.1997.8757Freire, M. A. (2005). Translation initiation factor (iso) 4E interacts with BTF3, the β subunit of the nascent polypeptide-associated complex. Gene, 345(2), 271-277. doi:10.1016/j.gene.2004.11.030Freire, M. A., Tourneur, C., Granier, F., Camonis, J., El Amrani, A., Browning, K. S., & Robaglia, C. (2000). Plant Molecular Biology, 44(2), 129-140. doi:10.1023/a:1006494628892Dreher, T. W., & Miller, W. A. (2006). Translational control in positive strand RNA plant viruses. Virology, 344(1), 185-197. doi:10.1016/j.virol.2005.09.031Thivierge, K., Nicaise, V., Dufresne, P. J., Cotton, S., Laliberté, J.-F., Le Gall, O., & Fortin, M. G. (2005). Plant Virus RNAs. Coordinated Recruitment of Conserved Host Functions by (+) ssRNA Viruses during Early Infection Events: Figure 1. Plant Physiology, 138(4), 1822-1827. doi:10.1104/pp.105.064105Deprost, D., Yao, L., Sormani, R., Moreau, M., Leterreux, G., Nicolaï, M., … Meyer, C. (2007). The Arabidopsis TOR kinase links plant growth, yield, stress resistance and mRNA translation. EMBO reports, 8(9), 864-870. doi:10.1038/sj.embor.7401043Manjunath, S., Williams, A. J., & Bailey-Serres, J. (1999). Oxygen deprivation stimulates Ca2+-mediated phosphorylation of mRNA cap-binding protein eIF4E in maize roots. The Plant Journal, 19(1), 21-30. doi:10.1046/j.1365-313x.1999.00489.xRausell, A., Kanhonou, R., Yenush, L., Serrano, R., & Ros, R. (2003). The translation initiation factor eIF1A is an important determinant in the tolerance to NaCl stress in yeast and plants. The Plant Journal, 34(3), 257-267. doi:10.1046/j.1365-313x.2003.01719.xSanan-Mishra, N., Pham, X. H., Sopory, S. K., & Tuteja, N. (2005). Pea DNA helicase 45 overexpression in tobacco confers high salinity tolerance without affecting yield. Proceedings of the National Academy of Sciences, 102(2), 509-514. doi:10.1073/pnas.0406485102Kim, T.-H., Kim, B.-H., Yahalom, A., Chamovitz, D. A., & von Arnim, A. G. (2004). Translational Regulation via 5′ mRNA Leader Sequences Revealed by Mutational Analysis of the Arabidopsis Translation Initiation Factor Subunit eIF3h. The Plant Cell, 16(12), 3341-3356. doi:10.1105/tpc.104.026880Schepetilnikov, M., Kobayashi, K., Geldreich, A., Caranta, C., Robaglia, C., Keller, M., & Ryabova, L. A. (2011). Viral factor TAV recruits TOR/S6K1 signalling to activate reinitiation after long ORF translation. The EMBO Journal, 30(7), 1343-1356. doi:10.1038/emboj.2011.39Mayberry, L. K., Allen, M. L., Nitka, K. R., Campbell, L., Murphy, P. A., & Browning, K. S. (2011). Plant Cap-binding Complexes Eukaryotic Initiation Factors eIF4F and eIFISO4F. Journal of Biological Chemistry, 286(49), 42566-42574. doi:10.1074/jbc.m111.280099Carberry, S. E., Goss, D. J., & Darzynkiewicz, E. (1991). A comparison of the binding of methylated cap analogs to wheat germ protein synthesis initiation factors 4F and (iso) 4F. Biochemistry, 30(6), 1624-1627. doi:10.1021/bi00220a026Lellis, A. D., Allen, M. L., Aertker, A. W., Tran, J. K., Hillis, D. M., Harbin, C. R., … Browning, K. S. (2010). Deletion of the eIFiso4G subunit of the Arabidopsis eIFiso4F translation initiation complex impairs health and viability. Plant Molecular Biology, 74(3), 249-263. doi:10.1007/s11103-010-9670-zDinkova, T. D., Zepeda, H., Martínez-Salas, E., Martínez, L. M., Nieto-Sotelo, J., & Jiménez, E. S. (2005). Cap-independent translation of maize Hsp101. The Plant Journal, 41(5), 722-731. doi:10.1111/j.1365-313x.2005.02333.xHutvagner, G. (2002). A microRNA in a Multiple-Turnover RNAi Enzyme Complex. Science, 297(5589), 2056-2060. doi:10.1126/science.1073827Voinnet, O. (2009). Origin, Biogenesis, and Activity of Plant MicroRNAs. Cell, 136(4), 669-687. doi:10.1016/j.cell.2009.01.046Brodersen, P., Sakvarelidze-Achard, L., Bruun-Rasmussen, M., Dunoyer, P., Yamamoto, Y. Y., Sieburth, L., & Voinnet, O. (2008). Widespread Translational Inhibition by Plant miRNAs and siRNAs. Science, 320(5880), 1185-1190. doi:10.1126/science.1159151Sunkar, R., Li, Y.-F., & Jagadeeswaran, G. (2012). Functions of microRNAs in plant stress responses. Trends in Plant Science, 17(4), 196-203. doi:10.1016/j.tplants.2012.01.010Dong, Z., Shi, L., Wang, Y., Chen, L., Cai, Z., Wang, Y., … Li, X. (2013). Identification and Dynamic Regulation of microRNAs Involved in Salt Stress Responses in Functional Soybean Nodules by High-Throughput Sequencing. International Journal of Molecular Sciences, 14(2), 2717-2738. doi:10.3390/ijms14022717Srivastava, S., Srivastava, A. K., Suprasanna, P., & D’Souza, S. F. (2012). Identification and profiling of arsenic stress-induced microRNAs inBrassica juncea. Journal of Experimental Botany, 64(1), 303-315. doi:10.1093/jxb/ers333Dugas, D. V., & Bartel, B. (2008). Sucrose induction of Arabidopsis miR398 represses two Cu/Zn superoxide dismutases. Plant Molecular Biology, 67(4), 403-417. doi:10.1007/s11103-008-9329-1Aukerman, M. J., & Sakai, H. (2003). Regulation of Flowering Time and Floral Organ Identity by a MicroRNA and Its APETALA2-Like Target Genes. The Plant Cell, 15(11), 2730-2741. doi:10.1105/tpc.016238Chen, X. (2004). A MicroRNA as a Translational Repressor of APETALA2 in Arabidopsis Flower Development. Science, 303(5666), 2022-2025. doi:10.1126/science.1088060Park, W., Li, J., Song, R., Messing, J., & Chen, X. (2002). CARPEL FACTORY, a Dicer Homolog, and HEN1, a Novel Protein, Act in microRNA Metabolism in Arabidopsis thaliana. Current Biology, 12(17), 1484-1495. doi:10.1016/s0960-9822(02)01017-5Gu, S., & Kay, M. A. (2010). How do miRNAs mediate translational repression? Silence, 1(1), 11. doi:10.1186/1758-907x-1-11Lanet, E., Delannoy, E., Sormani, R., Floris, M., Brodersen, P., Crété, P., … Robaglia, C. (2009). Biochemical Evidence for Translational Repression by Arabidopsis MicroRNAs. The Plant Cell, 21(6), 1762-1768. doi:10.1105/tpc.108.063412Olsthoorn, R. C. L. (1999). A conformational switch at the 3’ end of a plant virus RNA regulates viral replication. The EMBO Journal, 18(17), 4856-4864. doi:10.1093/emboj/18.17.4856Smirnyagina, E. V., Morozov, S. Y., Rodionova, N. P., Miroshnichenko, N. A., Solovyev, A. G., Fedorkin, O. N., & Atabekov, J. G. (1991). Translational efficiency and competitive ability of mRNAs with 5′-untranslated αβ-leader of potato virus X RNA. Biochimie, 73(5), 587-598. doi:10.1016/0300-9084(91)90027-xThanaraj, T. A., & Pandit, M. W. (1990). Translation-Initiation Promoting Site on Transcripts of Highly Expressed Genes FromSaccharomyces cerevisiaeand the Role of Hairpin Stems to Position the Site Near the Initiation Codon. Journal of Biomolecular Structure and Dynamics, 7(6), 1279-1289. doi:10.1080/07391102.1990.10508565Tomashevskaya, O. L., Solovyev, A. G., Karpova, O. V., Fedorkin, O. N., Rodionova, N. P., Morozov, S. Y., & Atabekov, J. G. (1993). Effects of sequence elements in the potato virus X RNA 5’ non-translated  beta-leader on its translation enhancing activity. Journal of General Virology, 74(12), 2717-2724. doi:10.1099/0022-1317-74-12-2717Belkum, A. van, Abrahams, J. P., Pleij, C. W. A., & Bosch, L. (1985). Five pseudoknots are present at the 204 nucleotides long 3’ noncoding region of tobacco mosak virus RNA. Nucleic Acids Research, 13(21), 7673-7686. doi:10.1093/nar/13.21.7673Gallie, D. R. (2002). The 5’-leader of tobacco mosaic virus promotes translation through enhanced recruitment of eIF4F. Nucleic Acids Research, 30(15), 3401-3411. doi:10.1093/nar/gkf457Wells, D. R., Tanguay, R. L., Le, H., & Gallie, D. R. (1998). HSP101 functions as a specific translational regulatory protein whose activity is regulated by nutrient status. Genes & Development, 12(20), 3236-3251. doi:10.1101/gad.12.20.3236Leonard, S., Plante, D., Wittmann, S., Daigneault, N., Fortin, M. G., & Laliberte, J.-F. (2000). Complex Formation between Potyvirus VPg and Translation Eukaryotic Initiation Factor 4E Correlates with Virus Infectivity. Journal of Virology, 74(17), 7730-7737. doi:10.1128/jvi.74.17.7730-7737.2000Wittmann, S., Chatel, H., Fortin, M. G., & Laliberté, J.-F. (1997). Interaction of the Viral Protein Genome Linked of Turnip Mosaic Potyvirus with the Translational Eukaryotic Initiation Factor (iso) 4E ofArabidopsis thalianaUsing the Yeast Two-Hybrid System. Virology, 234(1), 84-92. doi:10.1006/viro.1997.8634Robaglia, C., & Caranta, C. (2006). Translation initiation factors: a weak link in plant RNA virus infection. Trends in Plant Science, 11(1), 40-45. doi:10.1016/j.tplants.2005.11.004WANG, A., & KRISHNASWAMY, S. (2012). Eukaryotic translation initiation factor 4E-mediated recessive resistance to plant viruses and its utility in crop improvement. Molecular Plant Pathology, 13(7), 795-803. doi:10.1111/j.1364-3703.2012.00791.xLellis, A. D., Kasschau, K. D., Whitham, S. A., & Carrington, J. C. (2002). Loss-of-Susceptibility Mutants of Arabidopsis thaliana Reveal an Essential Role for eIF(iso)4E during Potyvirus Infection. Current Biology, 12(12), 1046-1051. doi:10.1016/s0960-9822(02)00898-9Duprat, A., Caranta, C., Revers, F., Menand, B., Browning, K. S., & Robaglia, C. (2002). The Arabidopsis eukaryotic initiation factor (iso)4E is dispensable for plant growth but required for susceptibility to potyviruses. The Plant Journal, 32(6), 927-934. doi:10.1046/j.1365-313x.2002.01481.xSato, M., Nakahara, K., Yoshii, M., Ishikawa, M., & Uyeda, I. (2005). Selective involvement of members of the eukaryotic initiation factor 4E family in the infection ofArabidopsis thalianaby potyviruses. FEBS Letters, 579(5), 1167-1171. doi:10.1016/j.febslet.2004.12.086Ruffel, S., Dussault, M.-H., Palloix, A., Moury, B., Bendahmane, A., Robaglia, C., & Caranta, C. (2002). A natural recessive resistance gene against potato virus Y in pepper corresponds to the eukaryotic initiation factor 4E (eIF4E). The Plant Journal, 32(6), 1067-1075. doi:10.1046/j.1365-313x.2002.01499.xNicaise, V., German-Retana, S., Sanjuán, R., Dubrana, M.-P., Mazier, M., Maisonneuve, B., … LeGall, O. (2003). The Eukaryotic Translation Initiation Factor 4E Controls Lettuce Susceptibility to the Potyvirus Lettuce mosaic virus. Plant Physiology, 132(3), 1272-1282. doi:10.1104/pp.102.017855Ruffel, S., Gallois, J. L., Lesage, M. L., & Caranta, C. (2005). The recessive potyvirus resistance gene pot-1 is the tomato orthologue of the pepper pvr2-eIF4E gene. Molecular Genetics and Genomics, 274(4), 346-353. doi:10.1007/s00438-005-0003-xKhan, M. A., Miyoshi, H., Gallie, D. R., & Goss, D. J. (2007). Potyvirus Genome-linked Protein, VPg, Directly Affects Wheat Germin VitroTranslation. Journal of Biological Chemistry, 283(3), 1340-1349. doi:10.1074/jbc.m703356200Cotton, S., Dufresne, P. J., Thivierge, K., Ide, C., & Fortin, M. G. (2006). The VPgPro protein of Turnip mosaic virus: In vitro inhibition of translation from a ribonuclease activity. Virology, 351(1), 92-100. doi:10.1016/j.virol.2006.03.019Grzela, R., Strokovska, L., Andrieu, J.-P., Dublet, B., Zagorski, W., & Chroboczek, J. (2006). Potyvirus terminal protein VPg, effector of host eukaryotic initiation factor eIF4E. Biochimie, 88(7), 887-896. doi:10.1016/j.biochi.2006.02.012Kneller, E. L. P., Rakotondrafara, A. M., & Miller, W. A. (2006). Cap-independent translation of plant viral RNAs. Virus Research, 119(1), 63-75. doi:10.1016/j.virusres.2005.10.010Zeenko, V., & Gallie, D. R. (2005). Cap-independent Translation of Tobacco Etch Virus Is Conferred by an RNA Pseudoknot in the 5′-Leader. Journal of Biological Chemistry, 280(29), 26813-26824. doi:10.1074/jbc.m503576200Miller, W. A., & White, K. A. (2006). Long-Distance RNA-RNA Interactions in Plant Virus Gene Expression and Replication. Annual Review of Phytopathology, 44(1), 447-467. doi:10.1146/annurev.phyto.44.070505.143353Wang, S., Browning, K. S., & Miller, W. A. (1997). A viral sequence in the 3′-untranslated region mimics a 5′ cap in facilitating translation of uncapped mRNA. The EMBO Journal, 16(13), 4107-4116. doi:10.1093/emboj/16.13.4107Gao, F., Kasprzak, W., Stupina, V. A., Shapiro, B. A., & Simon, A. E. (2012). A Ribosome-Binding, 3′ Translational Enhancer Has a T-Shaped Structure and Engages in a Long-Distance RNA-RNA Interaction. Journal of Virology, 86(18), 9828-9842. doi:10.1128/jvi.00677-12Wang, Z., Treder, K., & Miller, W. A. (2009). Structure of a Viral Cap-independent Translation Element That Functions via High Affinity Binding to the eIF4E Subunit of eIF4F. Journal of Biological Chemistry, 284(21), 14189-14202. doi:10.1074/jbc.m808841200Gazo, B. M., Murphy, P., Gatchel, J. R., & Browning, K. S. (2004). A Novel Interaction of Cap-binding Protein Complexes Eukaryotic Initiation Factor (eIF) 4F and eIF(iso)4F with a Region in the 3′-Untranslated Region of Satellite Tobacco Necrosis Virus. Journal of Biological Chemistry, 279(14), 13584-13592. doi:10.1074/jbc.m311361200Mardanova, E. S., Zamchuk, L. A., Skulachev, M. V., & Ravin, N. V. (2008). The 5′ untranslated region of the maize alcohol dehydrogenase gene contains an internal ribosome entry site. Gene, 420(1), 11-16. doi:10.1016/j.gene.2008.04.00

Pre-symptomatic transcriptome changes during cold storage of chilling sensitive and resistant peach cultivars to elucidate chilling injury mechanisms

Background: Cold storage induces chilling injury (CI) disorders in peach fruit (woolliness/mealiness, flesh browning and reddening/bleeding) manifested when ripened at shelf life. To gain insight into the mechanisms underlying CI, we analyzed the transcriptome of 'Oded' (high tolerant) and 'Hermoza' (relatively tolerant to woolliness, but sensitive to browning and bleeding) peach cultivars at pre-symptomatic stages. The expression profiles were compared and validated with two previously analyzed pools (high and low sensitive to woolliness) from the Pop-DG population. The four fruit types cover a wide range of sensitivity to CI. The four fruit types were also investigated with the ROSMETER that provides information on the specificity of the transcriptomic response to oxidative stress. Results: We identified quantitative differences in a subset of core cold responsive genes that correlated with sensitivity or tolerance to CI at harvest and during cold storage, and also subsets of genes correlating specifically with high sensitivity to woolliness and browning. Functional analysis indicated that elevated levels, at harvest and during cold storage, of genes related to antioxidant systems and the biosynthesis of metabolites with antioxidant activity correlates with tolerance. Consistent with these results, ROSMETER analysis revealed oxidative stress in 'Hermoza' and the progeny pools, but not in the cold resistant 'Oded'. By contrast, cold storage induced, in sensitivity to woolliness dependant manner, a gene expression program involving the biosynthesis of secondary cell wall and pectins. Furthermore, our results indicated that while ethylene is related to CI tolerance, differential auxin subcellular accumulation and signaling may play a role in determining chilling sensitivity/tolerance. In addition, sugar partitioning and demand during cold storage may also play a role in the tolerance/sensitive mechanism. The analysis also indicates that vesicle trafficking, membrane dynamics and cytoskeleton organization could have a role in the tolerance/sensitive mechanism. In the case of browning, our results suggest that elevated acetaldehyde related genes together with the core cold responses may increase sensitivity to browning in shelf life. Conclusions: Our data suggest that in sensitive fruit a cold response program is activated and regulated by auxin distribution and ethylene and these hormones have a role in sensitivity to CI even before fruit are cold stored.This research was funded by US-Israel Binational Agriculture Research and Development Fund (BARD) Grant no. US-4027-07. We thank the European Science Foundation for Short Term Scientific Mission grants to A. Dagar (COST Action 924, reference codes COST-STSM-924-04254 and Quality Fruit COST FA1106 for networking.Pons Puig, C.; Dagar, A.; Martí Ibáñez, MC.; Singh, V.; Crisosto, CH.; Friedman, H.; Lurie, S.... (2015). Pre-symptomatic transcriptome changes during cold storage of chilling sensitive and resistant peach cultivars to elucidate chilling injury mechanisms. BMC Genomics. 16:1-35. https://doi.org/10.1186/s12864-015-1395-6S13516Crisosto C, Mitchell F, Ju Z. Susceptibility to chilling injury of peach, nectarine, and plum cultivars grown in California. Hort Sci. 1999;34:1116–8.Lurie S, Crisosto C. Chilling injury in peach and nectarine. Postharvest Biol Technol. 2005;37:195–208.Crisosto C, Mitchell F, Johnson S. Factors in fresh market stone fruit quality. Postharvest News Inform. 1995;6(2):17–21.Dawson DM, Melton LD, Watkins CB. Cell wall changes in nectarines (Prunus persica): solubilization and depolymerization of pectic and neutral polymers during ripening and in mealy fruit. Plant Physiol. 1992;100(3):1203–10.Zhou H, Sonego L, Khalchitski A, Ben Arie R, Lers A, Lurie A. Cell wall enzymes and cell wall changes in ‘Flavortop’ nectarines: mRNA abundance, enzyme activity, and changes in pectic and neutral polymers during ripening and in woolly fruit. J Am Soc Hort Sci. 2000;125:630–7.Jarvis MC, Briggs SPH, Knox JP. Intercellular adhesion and cell separation in plants. Plant Cell Environ. 2003;26(7):977–89.Zhou H, Ben-Arie R, Lurie S. Pectin esterase, polygalacturonase and gel formation in peach pectin fractions. Phytochemistry. 2000;55(3):191–5.Brummell DA, Dal Cin V, Lurie S, Crisosto CH, Labavitch JM. Cell wall metabolism during the development of chilling injury in cold-stored peach fruit: association of mealiness with arrested disassembly of cell wall pectins. J Exp Bot. 2004;55(405):2041–52.Kader AA, Chordas A. Evaluating the browning potential of peaches. Calif Agric. 1984;38:14–5.Cevallos-Casals BA, Byrne D, Okie WR, Cisneros-Zevallos L. Selecting new peach and plum genotypes rich in phenolic compounds and enhanced functional properties. Food Chem. 2006;96(2):273–80.Rojas G, Méndez MA, Muñoz C, Lemus G, Hinrichsen P. Identification of a minimal microsatellite marker panel for the fingerprinting of peach and nectarine cultivars. Electron J Biotechnol. 2008;11:4–5.Scorza R, Sherman WB, Lightner GW. Inbreeding and co-ancestry of low chill short fruit development period freestone peaches and nectarines produced by the University of Florida breeding program. Fruit Varieties J. 1988;43:79–85.Brooks R, Olmo HP. Register of New Fruit and Nut Varieties, 2nd edition edn: Univ of California Press; 1972.Okie WR, Service USAR. Handbook of peach and nectarine varieties: performance in the southeastern United States and index of names: U.S. Dept. of Agriculture, Agricultural Research Service; 1998Martínez-García P, Peace C, Parfitt D, Ogundiwin E, Fresnedo-Ramírez J, Dandekar A, et al. Influence of year and genetic factors on chilling injury susceptibility in peach (Prunus persica (L.) Batsch). Euphytica. 2012;185(2):267–80.Ogundiwin E, Martí C, Forment J, Pons C, Granell A, Gradziel T, et al. Development of ChillPeach genomic tools and identification of cold-responsive genes in peach fruit. Plant Mol Biol. 2008;68(4–5):379–97.Pons C, Martí C, Forment J, Crisosto CH, Dandekar AM, Granell A. A Bulk Segregant Gene Expression Analysis of a Peach Population Reveals Components of the Underlying Mechanism of the Fruit Cold Response. PLoS ONE. 2014;9(3):e90706.Rosenwasser S, Fluhr R, Joshi JR, Leviatan N, Sela N, Hetzroni A, et al. ROSMETER: A Bioinformatic Tool for the Identification of Transcriptomic Imprints Related to Reactive Oxygen Species Type and Origin Provides New Insights into Stress Responses. Plant Physiol. 2013;163(2):1071–83.Kader AA, Mitchell FG. Maturity and quality. In: James H. LaRue RSJ, vol. Publication No. 3331, editor. Peaches, Plums, and Nectarines: Growing and Handling for Fresh Market. Oakland, Calif: Cooperative Extension, University of California, Division of Agriculture and Natural Resources; 1989. p. 191–6.Dagar A, Friedman H, Lurie S. Thaumatin-like proteins and their possible role in protection against chilling injury in peach fruit. Postharvest Biol Technol. 2010;57(2):77–85.Lill RE, Van Der Mespel GJ. A method for measuring the juice content of mealy nectarines. Sci Hortic. 1988;36(3–4):267–71.Tusher VG, Tibshirani R, Chu G. Significance analysis of microarrays applied to the ionizing radiation response. Proc Natl Acad Sci. 2001;98(9):5116–21.Pavlidis P, Noble WS. Matrix2png: a utility for visualizing matrix data. Bioinformatics. 2003;1-9(2):295–6.Dagar A, Pons Puig C, Marti Ibanez C, Ziliotto F, Bonghi CH, Crisosto C, et al. Comparative transcript profiling of a peach and its nectarine mutant at harvest reveals differences in gene expression related to storability. Tree Genet Genomes. 2013;9(1):223–35.Doherty CJ, Van Buskirk HA, Myers SJ, Thomashow MF. Roles for Arabidopsis CAMTA transcription factors in cold-regulated gene expression and freezing tolerance. Plant Cell. 2009;21(3):972–84.Gilmour S, Zarka D, Stockinger E, Salazar M, Houghton J, Thomashow M. Low temperature regulation of the Arabidopsis CBF family of AP2 transcriptional activators as an early step in cold-induced COR gene expression. Plant J. 1998;16(4):433–42.Dong L, Zhou H-W, Sonego L, Lers A, Lurie S. Ethylene involvement in the cold storage disorder of ‘Flavortop’ nectarine. Postharvest Biol Technol. 2001;23(2):105–15.Zhou H-W, Dong L, Ben-Arie R, Lurie S. The role of ethylene in the prevention of chilling injury in nectarines. J Plant Physiol. 2001;158(1):55–61.Kilian J, Whitehead D, Horak J, Wanke D, Weinl S, Batistic O, et al. The AtGenExpress global stress expression data set: protocols, evaluation and model data analysis of UV-B light, drought and cold stress responses. Plant J. 2007;50(2):347–63.Giraldo E, Díaz A, Corral JM, García A. Applicability of 2-DE to assess differences in the protein profile between cold storage and not cold storage in nectarine fruits. J Proteome. 2012;75(18):5774–82.Obenland D, Vensel W, Hurkman Ii W. Alterations in protein expression associated with the development of mealiness in peaches. J Hortic Sci Biotechnol. 2008;83(1):85–93.Vizoso P, Meisel L, Tittarelli A, Latorre M, Saba J, Caroca R, et al. Comparative EST transcript profiling of peach fruits under different post-harvest conditions reveals candidate genes associated with peach fruit quality. BMC Genomics. 2009;10(1):423.Hannah M, Heyer A, Hincha D. A Global Survey of Gene Regulation during Cold Acclimation in Arabidopsis thaliana. PLoS Genet. 2005;1(2):e26.Walia H, Wilson C, Condamine P, Liu X, Ismail AM, Zeng L, et al. Comparative Transcriptional Profiling of Two Contrasting Rice Genotypes under Salinity Stress during the Vegetative Growth Stage. Plant Physiol. 2005;139(2):822–35.Lurie S, Zhou H-W, Lers A, Sonego L, Alexandrov S, Shomer I. Study of pectin esterase and changes in pectin methylation during normal and abnormal peach ripening. Physiol Plant. 2003;119(2):287–94.Peace C, Crisosto C, Gradziel T. Endopolygalacturonase: A candidate gene for Freestone and Melting flesh in peach. Mol Breed. 2005;16(1):21–31.Luza JG, van Gorsel R, Polito VS, Kader AA. Chilling Injury in Peaches: A Cytochemical and Ultrastructural Cell Wall Study. J Am Soc Hortic Sci. 1992;117(1):114–8.Masia A, Zanchin A, Rascio N, Ramina A. Some Biochemical and Ultrastructural Aspects of Peach Fruit Development. J Am Soc Hortic Sci. 1992;117(5):808–15.Dean GH, Zheng H, Tewari J, Huang J, Young DS, Hwang YT, et al. The Arabidopsis MUM2 Gene Encodes a β-Galactosidase Required for the Production of Seed Coat Mucilage with Correct Hydration Properties. Plant Cell Online. 2007;19(12):4007–21.Johnson CS, Kolevski B, Smyth DR. TRANSPARENT TESTA GLABRA2, a Trichome and Seed Coat Development Gene of Arabidopsis, Encodes a WRKY Transcription Factor. Plant Cell Online. 2002;14(6):1359–75.Karssen CM, der Swan DLC B-v, Breekland AE, Koornneef M. Induction of dormancy during seed development by endogenous abscisic acid: studies on abscisic acid deficient genotypes of Arabidopsis thaliana (L.) Heynh. Planta. 1983;157(2):158–65.Bui M, Lim N, Sijacic P, Liu Z. LEUNIG_HOMOLOG and LEUNIG Regulate Seed Mucilage Extrusion in ArabidopsisF. J Integr Plant Biol. 2011;53(5):399–408.Hussey S, Mizrachi E, Spokevicius A, Bossinger G, Berger D, Myburg A. SND2, a NAC transcription factor gene, regulates genes involved in secondary cell wall development in Arabidopsis fibres and increases fibre cell area in Eucalyptus. BMC Plant Biol. 2011;11(1):173.Jin H, Cominelli E, Bailey P, Parr A, Mehrtens F, Jones J, et al. Transcriptional repression by AtMYB4 controls production of UV‐protecting sunscreens in Arabidopsis. EMBO J. 2000;19(22):6150–61.Romera-Branchat M, Ripoll JJ, Yanofsky MF, Pelaz S. The WOX13 homeobox gene promotes replum formation in the Arabidopsis thaliana fruit. Plant J. 2013;73(1):37–49.Itkin M, Seybold H, Breitel D, Rogachev I, Meir S, Aharoni A. TOMATO AGAMOUS-LIKE 1 is a component of the fruit ripening regulatory network. Plant J. 2009;60(6):1081–95.Bemer M, Karlova R, Ballester AR, Tikunov YM, Bovy AG, Wolters-Arts M, et al. The Tomato FRUITFULL Homologs TDR4/FUL1 and MBP7/FUL2 Regulate Ethylene-Independent Aspects of Fruit Ripening. Plant Cell Online. 2012;24(11):4437–51.Jaakola L, Poole M, Jones MO, Kämäräinen-Karppinen T, Koskimäki JJ, Hohtola A, et al. A SQUAMOSA MADS Box Gene Involved in the Regulation of Anthocyanin Accumulation in Bilberry Fruits. Plant Physiol. 2010;153(4):1619–29.Ogundiwin EA, Peace CP, Nicolet CM, Rashbrook VK, Gradziel TM, Bliss FA, et al. Leucoanthocyanidin dioxygenase gene (PpLDOX): a potential functional marker for cold storage browning in peach. Tree Genetics Genomes. 2008;4(3):543–54.Baxter IR, Young JC, Armstrong G, Foster N, Bogenschutz N, Cordova T, et al. A plasma membrane H + −ATPase is required for the formation of proanthocyanidins in the seed coat endothelium of Arabidopsis thaliana. Proc Natl Acad Sci U S A. 2005;102(7):2649–54.Cheng GW, Crisosto CH. Browning Potential, Phenolic Composition, and Polyphenoloxidase Activity of Buffer Extracts of Peach and Nectarine Skin Tissue. J Am Soc Hortic Sci. 1995;120(5):835–8.Wang Y-S, Tian S-P, Xu Y. Effects of high oxygen concentration on pro- and anti-oxidant enzymes in peach fruits during postharvest periods. Food Chem. 2005;91(1):99–104.Sevillano L, Sanchez-Ballesta MT, Romojaro F, Flores FB. Physiological, hormonal and molecular mechanisms regulating chilling injury in horticultural species. Postharvest technologies applied to reduce its impact. J Sci Food Agric. 2009;89(4):555–73.Provart NJ, Gil P, Chen W, Han B, Chang HS, Wang X, et al. Gene expression phenotypes of Arabidopsis associated with sensitivity to low temperatures. Plant Physiol. 2003;132(2):893–906.Prasad T, Anderson M, Stewart C. Acclimation, Hydrogen Peroxide, and Abscisic Acid Protect Mitochondria against Irreversible Chilling Injury in Maize Seedlings. Plant Physiol. 1994;105(2):619–27.Mhamdi A, Queval G, Chaouch S, Vanderauwera S, Van Breusegem F, Noctor G. Catalase function in plants: a focus on Arabidopsis mutants as stress-mimic models. J Exp Bot. 2010;61(15):4197–220.Ulmasov T, Murfett J, Hagen G, Guilfoyle TJ. Aux/IAA proteins repress expression of reporter genes containing natural and highly active synthetic auxin response elements. Plant Cell Online. 1997;9(11):1963–71.Schepetilnikov M, Dimitrova M, Mancera E, Martínez AG, Keller M, Ryabova LA. TOR and S6K1 promote translation reinitiation of uORF containing mRNAs via phosphorylation of eIF3h. EMBO J. 2013;32(8):1087–102.Xiong Y, Sheen J. The Role of Target of Rapamycin Signaling Networks in Plant Growth and Metabolism. Plant Physiol. 2014;164(2):499–512.Murray JAH, Jones A, Godin C, Traas J. Systems Analysis of Shoot Apical Meristem Growth and Development: Integrating Hormonal and Mechanical Signaling. Plant Cell Online. 2012;24(10):3907–19.Leiber R-M, John F, Verhertbruggen Y, Diet A, Knox JP, Ringli C. The TOR Pathway Modulates the Structure of Cell Walls in Arabidopsis. Plant Cell Online. 2010;22(6):1898–908.Garcia-Hernandez M, Davies E, Baskin TI, Staswick PE. Association of Plant p40 Protein with Ribosomes Is Enhanced When Polyribosomes Form during Periods of Active Tissue Growth. Plant Physiol. 1996;111(2):559–68.Cunningham JT, Rodgers JT, Arlow DH, Vazquez F, Mootha VK, Puigserver P. mTOR controls mitochondrial oxidative function through a YY1-PGC-1[agr] transcriptional complex. Nature. 2007;450(7170):736–40.Baur AH, Yang SF. Methionine metabolism in apple tissue in relation to ethylene biosynthesis. Phytochemistry. 1972;11(11):3207–14.Peiser GD, Wang T-T, Hoffman NE, Yang SF, Liu H-w, Walsh CT. Formation of cyanide from carbon 1 of 1-aminocyclopropane-1-carboxylic acid during its conversion to ethylene. Proc Natl Acad Sci. 1984;81(10):3059–63.Begheldo M, Manganaris GA, Bonghi C, Tonutti P. Different postharvest conditions modulate ripening and ethylene biosynthetic and signal transduction pathways in Stony Hard peaches. Postharvest Biol Technol. 2008;48(1):8–8.Shi Y, Tian S, Hou L, Huang X, Zhang X, Guo H, et al. Ethylene signaling negatively regulates freezing tolerance by repressing expression of CBF and type-A ARR genes in Arabidopsis. Plant Cell. 2012;24(6):2578–95.Thain SC, Vandenbussche F, Laarhoven LJJ, Dowson-Day MJ, Wang Z-Y, Tobin EM, et al. Circadian Rhythms of Ethylene Emission in Arabidopsis. Plant Physiol. 2004;136(3):3751–61.Wang KLC, Yoshida H, Lurin C, Ecker JR. Regulation of ethylene gas biosynthesis by the Arabidopsis ETO1 protein. Nature. 2004;428(6986):945–50.Zheng Z, Guo Y, Novák O, Dai X, Zhao Y, Ljung K, et al. Coordination of auxin and ethylene biosynthesis by the aminotransferase VAS1. Nat Chem Biol. 2013;9(4):244–6.Poschet G, Hannich B, Raab S, Jungkunz I, Klemens PAW, Krueger S, et al. A Novel Arabidopsis Vacuolar Glucose Exporter is involved in cellular Sugar Homeostasis and affects Composition of Seed Storage Compounds. Plant Physiol. 2011;157(4):1664–76.Wang K, Shao X, Gong Y, Zhu Y, Wang H, Zhang X, et al. The metabolism of soluble carbohydrates related to chilling injury in peach fruit exposed to cold stress. Postharvest Biol Technol. 2013;86:53–61.Liu Y-H, Offler CE, Ruan Y-L. Regulation of fruit and seed response to heat and drought by sugars as nutrients and signals. Frontiers Plant Sci. 2013;4:282.Coello P, Hey SJ, Halford NG. The sucrose non-fermenting-1-related (SnRK) family of protein kinases: potential for manipulation to improve stress tolerance and increase yield. J Exp Bot. 2011;62(3):883–93.Baena-González E, Sheen J. Convergent energy and stress signaling. Trends Plant Sci. 2008;13(9):474–82.Baena-González E. Energy Signaling in the Regulation of Gene Expression during Stress. Mol Plant. 2010;3(2):300–13.Robaglia C, Thomas M, Meyer C. Sensing nutrient and energy status by SnRK1 and TOR kinases. Curr Opin Plant Biol. 2012;15(3):301–7.Uemura M, Joseph RA, Steponkus PL. Cold Acclimation of Arabidopsis thaliana (Effect on Plasma Membrane Lipid Composition and Freeze-Induced Lesions). Plant Physiol. 1995;109(1):15–30.Zhang C, Tian S. Crucial contribution of membrane lipids’ unsaturation to acquisition of chilling-tolerance in peach fruit stored at 0°c. Food Chem. 2009;115(2):405–11.Abdrakhamanova A, Wang QY, Khokhlova L, Nick P. Is Microtubule Disassembly a Trigger for Cold Acclimation? Plant Cell Physiol. 2003;44(7):676–86.Baluška F, Hlavacka A, Šamaj J, Palme K, Robinson DG, Matoh T, et al. F-Actin-Dependent Endocytosis of Cell Wall Pectins in Meristematic Root Cells. Insights from Brefeldin A-Induced Compartments. Plant Physiology. 2002;130(1):422–31.Baluška F, Liners F, Hlavačka A, Schlicht M, Van Cutsem P, McCurdy DW, et al. Cell wall pectins and xyloglucans are internalized into dividing root cells and accumulate within cell plates during cytokinesis. Protoplasma. 2005;225(3–4):141–55.Gonzalez-Aguero M, Pavez L, Ibanez F, Pacheco I, Campos-Vargas R, Meisel L, et al. Identification of woolliness response genes in peach fruit after post-harvest treatments. J Exp Bot. 2008;59(8):1973–86.Bashline L, Lei L, Li S, Gu Y. Cell Wall, Cytoskeleton, and Cell Expansion in Higher Plants. Mol Plant. 2014;4:586–600.Dhonukshe P, Grigoriev I, Fischer R, Tominaga M, Robinson DG, Hašek J, et al. Auxin transport inhibitors impair vesicle motility and actin cytoskeleton dynamics in diverse eukaryotes. Proc Natl Acad Sci. 2008;105(11):4489–94.Fischer U, Men S, Grebe M. Lipid function in plant cell polarity. Curr Opin Plant Biol. 2004;7(6):670–6.Schrick K, Fujioka S, Takatsuto S, Stierhof Y-D, Stransky H, Yoshida S, et al. A link between sterol biosynthesis, the cell wall, and cellulose in Arabidopsis. Plant J. 2004;38(2):227–43.Cheng GW, Crisosto CH. Iron—Polyphenol Complex Formation and Skin Discoloration in Peaches and Nectarines. J Am Soc Hortic Sci. 1997;122(1):95–9.Bouché N, Fait A, Zik M, Fromm H. The root-specific glutamate decarboxylase (GAD1) is essential for sustaining GABA levels in Arabidopsis. Plant Mol Biol. 2004;55(3):315–25.Pedreschi R, Franck C, Lammertyn J, Erban A, Kopka J, Hertog M, et al. Metabolic profiling of ‘Conference’ pears under low oxygen stress. Postharvest Biol Technol. 2009;51(2):123–30