A cross population between D. kaki and D. virginiana shows high variability for saline tolerance and improved salt stress tolerance

[EN] Persimmon (Diospyros kaki Thunb.) production is facing important problems related to climate change in the Mediterranean areas. One of them is soil salinization caused by the decrease and change of the rainfall distribution. In this context, there is a need to develop cultivars adapted to the increasingly challenging soil conditions. In this study, a backcross between (D. kaki x D. virginiana) x D. kaki was conducted, to unravel the mechanism involved in salinity tolerance of persimmon. The backcross involved the two species most used as rootstock for persimmon production. Both species are clearly distinct in their level of tolerance to salinity. Variables related to growth, leaf gas exchange, leaf water relations and content of nutrients were significantly affected by saline stress in the backcross population. Water flow regulation appears as a mechanism of salt tolerance in persimmon via differences in water potential and transpiration rate, which reduces ion entrance in the plant. Genetic expression of eight putative orthologous genes involved in different mechanisms leading to salt tolerance was analyzed. Differences in expression levels among populations under saline or control treatment were found. The 'High affinity potassium transporter' (HKT1-like) reduced its expression levels in the roots in all studied populations. Results obtained allowed selection of tolerant rootstocks genotypes and describe the hypothesis about the mechanisms involved in salt tolerance in persimmon that will be useful for breeding salinity tolerant rootstocks.This study was funded by the IVIA and the European Funds for Regional Development. F. G.M.was funded by a PhD fellowship from the European Social Fund and the Generalitat Valenciana (ACIF/2016/115). The funders had no role in study design, data collection and analysis, decision to publish or preparation of the manuscript.Gil Muñoz, F.; Pérez-Pérez, JG.; Quiñones, A.; Primo-Capella, A.; Cebolla Cornejo, J.; Forner Giner, MA.; Badenes Catala, M.... (2020). A cross population between D. kaki and D. virginiana shows high variability for saline tolerance and improved salt stress tolerance. PLoS ONE. 15(2):1-27. https://doi.org/10.1371/journal.pone.0229023S127152Visconti, F., de Paz, J. M., Bonet, L., Jordà, M., Quiñones, A., & Intrigliolo, D. S. (2015). Effects of a commercial calcium protein hydrolysate on the salt tolerance of Diospyros kaki L. cv. «Rojo Brillante» grafted on Diospyros lotus L. Scientia Horticulturae, 185, 129-138. doi:10.1016/j.scienta.2015.01.028Forner-Giner, M. A., & Ancillo, G. (2013). Breeding Salinity Tolerance in Citrus Using Rootstocks. Salt Stress in Plants, 355-376. doi:10.1007/978-1-4614-6108-1_14Visconti, F., Intrigliolo, D. S., Quiñones, A., Tudela, L., Bonet, L., & de Paz, J. M. (2017). Differences in specific chloride toxicity to Diospyros kaki cv. «Rojo Brillante» grafted on D. lotus and D. virginiana. Scientia Horticulturae, 214, 83-90. doi:10.1016/j.scienta.2016.11.025INCESU, M., CIMEN, B., YESILOGLU, T., & YILMAZ, B. (2014). Growth and Photosynthetic Response of Two Persimmon Rootstocks (Diospyros kaki and D. virginiana) under Different Salinity Levels. Notulae Botanicae Horti Agrobotanici Cluj-Napoca, 42(2), 386-391. doi:10.15835/nbha4229471De Paz, J. M., Visconti, F., Chiaravalle, M., & Quiñones, A. (2016). Determination of persimmon leaf chloride contents using near-infrared spectroscopy (NIRS). Analytical and Bioanalytical Chemistry, 408(13), 3537-3545. doi:10.1007/s00216-016-9430-2Gil-Muñoz, F., Peche, P. M., Climent, J., Forner, M. A., Naval, M. M., & Badenes, M. L. (2018). Breeding and screening persimmon rootstocks for saline stress tolerance. Acta Horticulturae, (1195), 105-110. doi:10.17660/actahortic.2018.1195.18Besada, C., Gil, R., Bonet, L., Quiñones, A., Intrigliolo, D., & Salvador, A. (2016). Chloride stress triggers maturation and negatively affects the postharvest quality of persimmon fruit. Involvement of calyx ethylene production. Plant Physiology and Biochemistry, 100, 105-112. doi:10.1016/j.plaphy.2016.01.006Acosta-Motos, J., Ortuño, M., Bernal-Vicente, A., Diaz-Vivancos, P., Sanchez-Blanco, M., & Hernandez, J. (2017). Plant Responses to Salt Stress: Adaptive Mechanisms. Agronomy, 7(1), 18. doi:10.3390/agronomy7010018Munns, R., & Tester, M. (2008). Mechanisms of Salinity Tolerance. Annual Review of Plant Biology, 59(1), 651-681. doi:10.1146/annurev.arplant.59.032607.092911Sibole, J. V., Cabot, C., Poschenrieder, C., & Barceló, J. (2003). Ion allocation in two different salt-tolerant MediterraneanMedicagospecies. Journal of Plant Physiology, 160(11), 1361-1365. doi:10.1078/0176-1617-00811CRAIG PLETT, D., & MØLLER, I. S. (2010). Na+transport in glycophytic plants: what we know and would like to know. Plant, Cell & Environment, 33(4), 612-626. doi:10.1111/j.1365-3040.2009.02086.xSHAPIRA, O., KHADKA, S., ISRAELI, Y., SHANI, U., & SCHWARTZ, A. (2009). Functional anatomy controls ion distribution in banana leaves: significance of Na+seclusion at the leaf margins. Plant, Cell & Environment, 32(5), 476-485. doi:10.1111/j.1365-3040.2009.01941.xHuang, C. X., & Van Steveninck, R. F. M. (1989). Maintenance of Low Cl− Concentrations in Mesophyll Cells of Leaf Blades of Barley Seedlings Exposed to Salt Stress. Plant Physiology, 90(4), 1440-1443. doi:10.1104/pp.90.4.1440Karley, A. J., Leigh, R. A., & Sanders, D. (2000). Differential Ion Accumulation and Ion Fluxes in the Mesophyll and Epidermis of Barley. Plant Physiology, 122(3), 835-844. doi:10.1104/pp.122.3.835Karley, A. J., Leigh, R. A., & Sanders, D. (2000). Where do all the ions go? The cellular basis of differential ion accumulation in leaf cells. Trends in Plant Science, 5(11), 465-470. doi:10.1016/s1360-1385(00)01758-1JAMES, R. A., MUNNS, R., VON CAEMMERER, S., TREJO, C., MILLER, C., & CONDON, T. (A. G. . (2006). Photosynthetic capacity is related to the cellular and subcellular partitioning of Na+, K+and Cl-in salt-affected barley and durum wheat. Plant, Cell and Environment, 29(12), 2185-2197. doi:10.1111/j.1365-3040.2006.01592.xZekri, M., & Parsons, L. R. (1989). Growth and root hydraulic conductivity of several citrus rootstocks under salt and polyethylene glycol stresses. Physiologia Plantarum, 77(1), 99-106. doi:10.1111/j.1399-3054.1989.tb05984.xJoly, R. J. (1989). Effects of Sodium Chloride on the Hydraulic Conductivity of Soybean Root Systems. Plant Physiology, 91(4), 1262-1265. doi:10.1104/pp.91.4.1262Maurel, C., Verdoucq, L., Luu, D.-T., & Santoni, V. (2008). Plant Aquaporins: Membrane Channels with Multiple Integrated Functions. Annual Review of Plant Biology, 59(1), 595-624. doi:10.1146/annurev.arplant.59.032607.092734Johanson, U., Karlsson, M., Johansson, I., Gustavsson, S., Sjövall, S., Fraysse, L., … Kjellbom, P. (2001). The Complete Set of Genes Encoding Major Intrinsic Proteins in Arabidopsis Provides a Framework for a New Nomenclature for Major Intrinsic Proteins in Plants. Plant Physiology, 126(4), 1358-1369. doi:10.1104/pp.126.4.1358Carmen Martínez-Ballesta, M., Aparicio, F., Pallás, V., Martínez, V., & Carvajal, M. (2003). Influence of saline stress on root hydraulic conductance and PIP expression inArabidopsis. Journal of Plant Physiology, 160(6), 689-697. doi:10.1078/0176-1617-00861Boursiac, Y., Chen, S., Luu, D.-T., Sorieul, M., van den Dries, N., & Maurel, C. (2005). Early Effects of Salinity on Water Transport in Arabidopsis Roots. Molecular and Cellular Features of Aquaporin Expression. Plant Physiology, 139(2), 790-805. doi:10.1104/pp.105.065029López-Pérez, L., Martínez-Ballesta, M. del C., Maurel, C., & Carvajal, M. (2009). Changes in plasma membrane lipids, aquaporins and proton pump of broccoli roots, as an adaptation mechanism to salinity. Phytochemistry, 70(4), 492-500. doi:10.1016/j.phytochem.2009.01.014Rodríguez-Gamir, J., Ancillo, G., Legaz, F., Primo-Millo, E., & Forner-Giner, M. A. (2012). Influence of salinity on pip gene expression in citrus roots and its relationship with root hydraulic conductance, transpiration and chloride exclusion from leaves. Environmental and Experimental Botany, 78, 163-166. doi:10.1016/j.envexpbot.2011.12.027Chaumont, F., & Tyerman, S. D. (2014). Aquaporins: Highly Regulated Channels Controlling Plant Water Relations. Plant Physiology, 164(4), 1600-1618. doi:10.1104/pp.113.233791Amtmann, A., & Sanders, D. (1998). Mechanisms of Na+ Uptake by Plant Cells. Advances in Botanical Research, 75-112. doi:10.1016/s0065-2296(08)60310-9TESTER, M. (2003). Na+ Tolerance and Na+ Transport in Higher Plants. Annals of Botany, 91(5), 503-527. doi:10.1093/aob/mcg058Qiu, Q.-S., Barkla, B. J., Vera-Estrella, R., Zhu, J.-K., & Schumaker, K. S. (2003). Na+/H+ Exchange Activity in the Plasma Membrane of Arabidopsis. Plant Physiology, 132(2), 1041-1052. doi:10.1104/pp.102.010421Shi, H., Quintero, F. J., Pardo, J. M., & Zhu, J.-K. (2002). The Putative Plasma Membrane Na+/H+ Antiporter SOS1 Controls Long-Distance Na+ Transport in Plants. The Plant Cell, 14(2), 465-477. doi:10.1105/tpc.010371Zhu, J.-K., Liu, J., & Xiong, L. (1998). Genetic Analysis of Salt Tolerance in Arabidopsis: Evidence for a Critical Role of Potassium Nutrition. The Plant Cell, 10(7), 1181-1191. doi:10.1105/tpc.10.7.1181Liu, J., & Zhu, J.-K. (1998). A Calcium Sensor Homolog Required for Plant Salt Tolerance. Science, 280(5371), 1943-1945. doi:10.1126/science.280.5371.1943Halfter, U., Ishitani, M., & Zhu, J.-K. (2000). The Arabidopsis SOS2 protein kinase physically interacts with and is activated by the calcium-binding protein SOS3. Proceedings of the National Academy of Sciences, 97(7), 3735-3740. doi:10.1073/pnas.97.7.3735Liu, J., Ishitani, M., Halfter, U., Kim, C.-S., & Zhu, J.-K. (2000). The Arabidopsis thaliana SOS2 gene encodes a protein kinase that is required for salt tolerance. Proceedings of the National Academy of Sciences, 97(7), 3730-3734. doi:10.1073/pnas.97.7.3730Hrabak, E. M., Chan, C. W. M., Gribskov, M., Harper, J. F., Choi, J. H., Halford, N., … Harmon, A. C. (2003). The Arabidopsis CDPK-SnRK Superfamily of Protein Kinases. Plant Physiology, 132(2), 666-680. doi:10.1104/pp.102.011999Shi, H., Ishitani, M., Kim, C., & Zhu, J.-K. (2000). The Arabidopsis thaliana salt tolerance gene SOS1 encodes a putative Na+/H+ antiporter. Proceedings of the National Academy of Sciences, 97(12), 6896-6901. doi:10.1073/pnas.120170197Qiu, Q.-S., Guo, Y., Dietrich, M. A., Schumaker, K. S., & Zhu, J.-K. (2002). Regulation of SOS1, a plasma membrane Na+/H+ exchanger in Arabidopsis thaliana, by SOS2 and SOS3. Proceedings of the National Academy of Sciences, 99(12), 8436-8441. doi:10.1073/pnas.122224699Quintero, F. J., Ohta, M., Shi, H., Zhu, J.-K., & Pardo, J. M. (2002). Reconstitution in yeast of the Arabidopsis SOS signaling pathway for Na+ homeostasis. Proceedings of the National Academy of Sciences, 99(13), 9061-9066. doi:10.1073/pnas.132092099Quan, R., Lin, H., Mendoza, I., Zhang, Y., Cao, W., Yang, Y., … Guo, Y. (2007). SCABP8/CBL10, a Putative Calcium Sensor, Interacts with the Protein Kinase SOS2 to Protect Arabidopsis Shoots from Salt Stress. The Plant Cell, 19(4), 1415-1431. doi:10.1105/tpc.106.042291Quintero, F. J., Martinez-Atienza, J., Villalta, I., Jiang, X., Kim, W.-Y., Ali, Z., … Pardo, J. M. (2011). Activation of the plasma membrane Na/H antiporter Salt-Overly-Sensitive 1 (SOS1) by phosphorylation of an auto-inhibitory C-terminal domain. Proceedings of the National Academy of Sciences, 108(6), 2611-2616. doi:10.1073/pnas.1018921108Ji, H., Pardo, J. M., Batelli, G., Van Oosten, M. J., Bressan, R. A., & Li, X. (2013). The Salt Overly Sensitive (SOS) Pathway: Established and Emerging Roles. Molecular Plant, 6(2), 275-286. doi:10.1093/mp/sst017Isayenkov, S. V., & Maathuis, F. J. M. (2019). Plant Salinity Stress: Many Unanswered Questions Remain. Frontiers in Plant Science, 10. doi:10.3389/fpls.2019.00080Evans, A. R., Hall, D., Pritchard, J., & Newbury, H. J. (2011). The roles of the cation transporters CHX21 and CHX23 in the development of Arabidopsis thaliana. Journal of Experimental Botany, 63(1), 59-67. doi:10.1093/jxb/err271Uozumi, N., Kim, E. J., Rubio, F., Yamaguchi, T., Muto, S., Tsuboi, A., … Schroeder, J. I. (2000). The Arabidopsis HKT1 Gene Homolog Mediates Inward Na+ Currents in Xenopus laevis Oocytes and Na+ Uptake in Saccharomyces cerevisiae  . Plant Physiology, 122(4), 1249-1260. doi:10.1104/pp.122.4.1249Mäser, P., Eckelman, B., Vaidyanathan, R., Horie, T., Fairbairn, D. J., Kubo, M., … Schroeder, J. I. (2002). Altered shoot/root Na+ distribution and bifurcating salt sensitivity in Arabidopsis by genetic disruption of the Na+ transporter AtHKT1. FEBS Letters, 531(2), 157-161. doi:10.1016/s0014-5793(02)03488-9Berthomieu, P. (2003). Functional analysis of AtHKT1 in Arabidopsis shows that Na+ recirculation by the phloem is crucial for salt tolerance. The EMBO Journal, 22(9), 2004-2014. doi:10.1093/emboj/cdg207Rus, A., Lee, B., Muñoz-Mayor, A., Sharkhuu, A., Miura, K., Zhu, J.-K., … Hasegawa, P. M. (2004). AtHKT1 Facilitates Na+ Homeostasis and K+ Nutrition in Planta. Plant Physiology, 136(1), 2500-2511. doi:10.1104/pp.104.042234Sunarpi, Horie, T., Motoda, J., Kubo, M., Yang, H., Yoda, K., … Uozumi, N. (2005). Enhanced salt tolerance mediated by AtHKT1 transporter-induced Na+ unloading from xylem vessels to xylem parenchyma cells. The Plant Journal, 44(6), 928-938. doi:10.1111/j.1365-313x.2005.02595.xHuang, S., Spielmeyer, W., Lagudah, E. S., James, R. A., Platten, J. D., Dennis, E. S., & Munns, R. (2006). A Sodium Transporter (HKT7) Is a Candidate forNax1, a Gene for Salt Tolerance in Durum Wheat. Plant Physiology, 142(4), 1718-1727. doi:10.1104/pp.106.088864Byrt, C. S., Platten, J. D., Spielmeyer, W., James, R. A., Lagudah, E. S., Dennis, E. S., … Munns, R. (2007). HKT1;5-Like Cation Transporters Linked to Na+ Exclusion Loci in Wheat, Nax2 and Kna1. Plant Physiology, 143(4), 1918-1928. doi:10.1104/pp.106.093476Garciadeblás, B., Senn, M. E., Bañuelos, M. A., & Rodríguez-Navarro, A. (2003). Sodium transport and HKT transporters: the rice model. The Plant Journal, 34(6), 788-801. doi:10.1046/j.1365-313x.2003.01764.xHuang, S., Spielmeyer, W., Lagudah, E. S., & Munns, R. (2008). Comparative mapping of HKT genes in wheat, barley, and rice, key determinants of Na+ transport, and salt tolerance. Journal of Experimental Botany, 59(4), 927-937. doi:10.1093/jxb/ern033Horie, T., Costa, A., Kim, T. H., Han, M. J., Horie, R., Leung, H.-Y., … Schroeder, J. I. (2007). Rice OsHKT2;1 transporter mediates large Na+ influx component into K+-starved roots for growth. The EMBO Journal, 26(12), 3003-3014. doi:10.1038/sj.emboj.7601732Almeida, P., Katschnig, D., & de Boer, A. (2013). HKT Transporters—State of the Art. International Journal of Molecular Sciences, 14(10), 20359-20385. doi:10.3390/ijms141020359Cellier, F., Conéjéro, G., Ricaud, L., Luu, D. T., Lepetit, M., Gosti, F., & Casse, F. (2004). Characterization ofAtCHX17, a member of the cation/H+exchangers, CHX family, fromArabidopsis thalianasuggests a role in K+homeostasis. The Plant Journal, 39(6), 834-846. doi:10.1111/j.1365-313x.2004.02177.xSong, C.-P., Guo, Y., Qiu, Q., Lambert, G., Galbraith, D. W., Jagendorf, A., & Zhu, J.-K. (2004). A probable Na+(K+)/H+ exchanger on the chloroplast envelope functions in pH homeostasis and chloroplast development in Arabidopsis thaliana. Proceedings of the National Academy of Sciences, 101(27), 10211-10216. doi:10.1073/pnas.0403709101Padmanaban, S., Chanroj, S., Kwak, J. M., Li, X., Ward, J. M., & Sze, H. (2007). Participation of Endomembrane Cation/H+ Exchanger AtCHX20 in Osmoregulation of Guard Cells. Plant Physiology, 144(1), 82-93. doi:10.1104/pp.106.092155Szczerba, M. W., Britto, D. T., & Kronzucker, H. J. (2009). K+ transport in plants: Physiology and molecular biology. Journal of Plant Physiology, 166(5), 447-466. doi:10.1016/j.jplph.2008.12.009Brini, F., Gaxiola, R. A., Berkowitz, G. A., & Masmoudi, K. (2005). Cloning and characterization of a wheat vacuolar cation/proton antiporter and pyrophosphatase proton pump. Plant Physiology and Biochemistry, 43(4), 347-354. doi:10.1016/j.plaphy.2005.02.010Barragán, V., Leidi, E. O., Andrés, Z., Rubio, L., De Luca, A., Fernández, J. A., … Pardo, J. M. (2012). Ion Exchangers NHX1 and NHX2 Mediate Active Potassium Uptake into Vacuoles to Regulate Cell Turgor and Stomatal Function in Arabidopsis. The Plant Cell, 24(3), 1127-1142. doi:10.1105/tpc.111.095273Barbier-Brygoo, H., De Angeli, A., Filleur, S., Frachisse, J.-M., Gambale, F., Thomine, S., & Wege, S. (2011). Anion Channels/Transporters in Plants: From Molecular Bases to Regulatory Networks. Annual Review of Plant Biology, 62(1), 25-51. doi:10.1146/annurev-arplant-042110-103741Apse, M. P., Aharon, G. S., Snedden, W. A., & Blumwald, E. (1999). Salt Tolerance Conferred by Overexpression of a Vacuolar Na + /H + Antiport in Arabidopsis. Science, 285(5431), 1256-1258. doi:10.1126/science.285.5431.1256Gaxiola, R. A., Li, J., Undurraga, S., Dang, L. M., Allen, G. J., Alper, S. L., & Fink, G. R. (2001). Drought- and salt-tolerant plants result from overexpression of the AVP1 H+-pump. Proceedings of the National Academy of Sciences, 98(20), 11444-11449. doi:10.1073/pnas.191389398Callister, A. N., Arndt, S. K., & Adams, M. A. (2006). Comparison of four methods for measuring osmotic potential of tree leaves. Physiologia Plantarum, 127(3), 383-392. doi:10.1111/j.1399-3054.2006.00652.xBates, L. S., Waldren, R. P., & Teare, I. D. (1973). Rapid determination of free proline for water-stress studies. Plant and Soil, 39(1), 205-207. doi:10.1007/bf00018060Gilliam, J. W. (1971). Rapid Measurement of Chlorine in Plant Materials. Soil Science Society of America Journal, 35(3), 512-513. doi:10.2136/sssaj1971.03615995003500030051xGambino, G., Perrone, I., & Gribaudo, I. (2008). A Rapid and effective method for RNA extraction from different tissues of grapevine and other woody plants. Phytochemical Analysis, 19(6), 520-525. doi:10.1002/pca.1078Akagi, T., Henry, I. M., Kawai, T., Comai, L., & Tao, R. (2016). Epigenetic Regulation of the Sex Determination Gene MeGI in Polyploid Persimmon. The Plant Cell, 28(12), 2905-2915. doi:10.1105/tpc.16.00532Andersen, C. L., Jensen, J. L., & Ørntoft, T. F. (2004). Normalization of Real-Time Quantitative Reverse Transcription-PCR Data: A Model-Based Variance Estimation Approach to Identify Genes Suited for Normalization, Applied to Bladder and Colon Cancer Data Sets. Cancer Research, 64(15), 5245-5250. doi:10.1158/0008-5472.can-04-0496Akagi, T., Ikegami, A., Tsujimoto, T., Kobayashi, S., Sato, A., Kono, A., & Yonemori, K. (2009). DkMyb4 Is a Myb Transcription Factor Involved in Proanthocyanidin Biosynthesis in Persimmon Fruit. Plant Physiology, 151(4), 2028-2045. doi:10.1104/pp.109.146985Chambers, J. M., Cleveland, W. S., Kleiner, B., & Tukey, P. A. (2018). Graphical Methods for Data Analysis. doi:10.1201/9781351072304Flowers, T. J., & Colmer, T. D. (2008). Salinity tolerance in halophytes*. New Phytologist, 179(4), 945-963. doi:10.1111/j.1469-8137.2008.02531.xMunns, R. (2002). Comparative physiology of salt and water stress. Plant, Cell & Environment, 25(2), 239-250. doi:10.1046/j.0016-8025.2001.00808.xBrugnoli, E., & Lauteri, M. (1991). Effects of Salinity on Stomatal Conductance, Photosynthetic Capacity, and Carbon Isotope Discrimination of Salt-Tolerant (Gossypium hirsutum L.) and Salt-Sensitive (Phaseolus vulgaris L.) C3 Non-Halophytes. Plant Physiology, 95(2), 628-635. doi:10.1104/pp.95.2.628Koyro, H.-W. (2006). Effect of salinity on growth, photosynthesis, water relations and solute composition of the potential cash crop halophyte Plantago coronopus (L.). Environmental and Experimental Botany, 56(2), 136-146. doi:10.1016/j.envexpbot.2005.02.001Rahnama, A., James, R. A., Poustini, K., & Munns, R. (2010). Stomatal conductance as a screen for osmotic stress tolerance in durum wheat growing in saline soil. Functional Plant Biology, 37(3), 255. doi:10.1071/fp09148Zhu, X., Cao, Q., Sun, L., Yang, X., Yang, W., & Zhang, H. (2018). Stomatal Conductance and Morphology of Arbuscular Mycorrhizal Wheat Plants Response to Elevated CO2 and NaCl Stress. Frontiers in Plant Science, 9. doi:10.3389/fpls.2018.01363Horie, T., Sugawara, M., Okunou, K., Nakayama, H., Schroeder, J. I., Shinmyo, A., & Yoshida, K. (2008). Functions of HKT transporters in sodium transport in roots and in protecting leaves from salinity stress. Plant Biotechnology, 25(3), 233-239. doi:10.5511/plantbiotechnology.25.233Hazzouri, K. M., Khraiwesh, B., Amiri, K. M. A., Pauli, D., Blake, T., Shahid, M., … Masmoudi, K. (2018). Mapping of HKT1;5 Gene in Barley Using GWAS Approach and Its Implication in Salt Tolerance Mechanism. Frontiers in Plant Science, 9. doi:10.3389/fpls.2018.00156Han, Y., Yin, S., Huang, L., Wu, X., Zeng, J., Liu, X., … Zhang, G. (2018). A Sodium Transporter HvHKT1;1 Confers Salt Tolerance in Barley via Regulating Tissue and Cell Ion Homeostasis. Plant and Cell Physiology, 59(10), 1976-1989. doi:10.1093/pcp/pcy116Henderson, S. W., Baumann, U., Blackmore, D. H., Walker, A. R., Walker, R. R., & Gilliham, M. (2014). Shoot chloride exclusion and salt tolerance in grapevine is associated with differential ion transporter expression in roots. BMC Plant Biology, 14(1). doi:10.1186/s12870-014-0273-8Vitali, V., Bellati, J., Soto, G., Ayub, N. D., & Amodeo, G. (2015). Root hydraulic conductivity and adjustments in stomatal conductance: hydraulic strategy in response to salt stress in a halotolerant species. AoB Plants, 7, plv136. doi:10.1093/aobpla/plv13

MLH1-methylated endometrial cancer under 60 years of age as the “sentinel” cancer in female carriers of high-risk constitutional MLH1 epimutation

Objective. Universal screening of endometrial carcinoma (EC) for mismatch repair deficiency (MMRd) and Lynch syndrome uses presence of MLH1 methylation to omit common sporadic cases from follow-up germline testing. However, this overlooks rare cases with high-risk constitutional MLH1 methylation (epimutation), a poorly-recognized mechanism that predisposes to Lynch-type cancers with MLH1 methylation. We aimed to de-termine the role and frequency of constitutional MLH1 methylation among EC cases with MMRd, MLH1- methylated tumors.Methods. We screened blood for constitutional MLH1 methylation using pyrosequencing and real-time methylation-specific PCR in patients with MMRd, MLH1-methylated EC ascertained from (i) cancer clinics (n = 4, <60 years), and (ii) two population-based cohorts; Columbus-area (n = 68, all ages) and Ohio Colo-rectal Cancer Prevention Initiative (OCCPI) (n = 24, <60 years).Results. Constitutional MLH1 methylation was identified in three out of four patients diagnosed between 36 and 59 years from cancer clinics. Two had mono-/hemiallelic epimutation (similar to 50% alleles methylated). One with multiple primaries had low-level mosaicism in normal tissues and somatic second-hits affecting the unmethylated allele in all tumors, demonstrating causation. In the population-based cohorts, all 68 cases from the Columbus-area cohort were negative and low-level mosaic constitutional MLH1 methylation was identified in one patient aged 36 years out of 24 from the OCCPI cohort, representing one of six (similar to 17%) patients <50 years and one of 45 patients (similar to 2%) <60 years in the combined cohorts. EC was the first/dual-first cancer in three pa-tients with underlying constitutional MLH1 methylation.Conclusions. A correct diagnosis at first presentation of cancer is important as it will significantly alter clinical management. Screening for constitutional MLH1 methylation is warranted in patients with early-onset EC or syn-chronous/metachronous tumors (any age) displaying MLH1 methylation.(c) 2023 The Authors. Published by Elsevier Inc. This is an open access article under the CC BY-NC-ND license (http:// creativecommons.org/licenses/by-nc-nd/4.0/)

Phase I, multicenter, open-label study of intravenous VCN-01 oncolytic adenovirus with or without nab-paclitaxel plus gemcitabine in patients with advanced solid tumors

Background VCN-01 is an oncolytic adenovirus (Ad5 based) designed to replicate in cancer cells with dysfunctional RB1 pathway, express hyaluronidase to enhance virus intratumoral spread and facilitate chemotherapy and immune cells extravasation into the tumor. This phase I clinical trial was aimed to find the maximum tolerated dose/recommended phase II dose (RP2D) and dose-limiting toxicity (DLT) of the intravenous delivery of the replication-competent VCN-01 adenovirus in patients with advanced cancer. Methods Part I: patients with advanced refractory solid tumors received one single dose of VCN-01. Parts II and III: patients with pancreatic adenocarcinoma received VCN-01 (only in cycle 1) and nab-paclitaxel plus gemcitabine (VCN-concurrent on day 1 in Part II, and 7days before chemotherapy in Part III). Patients were required to have anti-Ad5 neutralizing antibody (NAbs) titers lower than 1/350 dilution. Pharmacokinetic and pharmacodynamic analyses were performed. Results 26% of the patients initially screened were excluded based on high NAbs levels. Sixteen and 12 patients were enrolled in Part I and II, respectively: RP2D were 1 x10(13) viral particles (vp)/patient (Part I), and 3.3x10(12) vp/patient (Part II). Fourteen patients were included in Part Ill: there were no DLTs and the RP2D was 1 x10(13) vp/patient. Observed DLTs were grade 4 aspartate aminotransferase increase in one patient (Part I, 1x10(13) vp), grade 4 febrile neutropenia in one patient and grade 5 thrombocytopenia plus enterocolitis in another patient (Part II, 1 x10(13) vp). In patients with pancreatic adenocarcinoma overall response rate were 50% (Part II) and 50% (Part III). VCN-01 viral genomes were detected in tumor tissue in five out of six biopsies (day 8). A second viral plasmatic peak and increased hyaluronidase serum levels suggested replication after intravenous injection in all patients. Increased levels of immune biomarkers (interferon- r,soluble lymphocyte activation ne-3, interleukin (IL)-6, IL-10) were found after VCN-01 administration. Conclusions Treatment with VCN-01 is feasible and has an acceptable safety. Encouraging biological and clinical activity was observed when administered in combination with nab-paditaxel plus gemcitabine to patients with pancreatic adenocarcinoma

Polymorphisms within inflammatory genes and colorectal cancer

BACKGROUND: Chronic inflammation is a risk factor for colorectal cancer and polymorphisms in the inflammatory genes could modulate the levels of inflammation. We have investigated ten single nucleotide polymorphisms (SNPs) in the following inflammation-related genes: TLR4 (Asp299Gly), CD14 (-260 T>C), MCP1 (-2518 A>G), IL12A (+7506 A>T, +8707 A>G, +9177 T>A, +9508 G>A), NOS2A (+524T>C), TNF (-857C>T), and PTGS1 (V444I) in 377 colorectal (CRC) cancer cases and 326 controls from Barcelona (Spain). RESULTS: There was no statistically significant association between the SNPs investigated and colorectal cancer risk. CONCLUSION: The lack of association may show that the inflammatory genes selected for this study are not involved in the carcinogenic process of colorectum. Alternatively, the negative results may derive from no particular biological effect of the analysed polymorphisms in relation to CRC. Otherwise, the eventual biological effect is so little to go undetected, unless analysing a much larger sample size

Cancer incidence and survival in Lynch syndrome patients receiving colonoscopic and gynaecological surveillance : first report from the prospective Lynch syndrome database

Objective Estimates of cancer risk and the effects of surveillance in Lynch syndrome have been subject to bias, partly through reliance on retrospective studies. We sought to establish more robust estimates in patients undergoing prospective cancer surveillance. Design We undertook a multicentre study of patients carrying Lynch syndrome-associated mutations affecting MLH1, MSH2, MSH6 or PMS2. Standardised information on surveillance, cancers and outcomes were collated in an Oracle relational database and analysed by age, sex and mutated gene. Results 1942 mutation carriers without previous cancer had follow-up including colonoscopic surveillance for 13 782 observation years. 314 patients developed cancer, mostly colorectal (n=151), endometrial (n=72) and ovarian (n=19). Cancers were detected from 25 years onwards in MLH1 and MSH2 mutation carriers, and from about 40 years in MSH6 and PMS2 carriers. Among first cancer detected in each patient the colorectal cancer cumulative incidences at 70 years by gene were 46%, 35%, 20% and 10% for MLH1, MSH2, MSH6 and PMS2 mutation carriers, respectively. The equivalent cumulative incidences for endometrial cancer were 34%, 51%, 49% and 24%; and for ovarian cancer 11%, 15%, 0% and 0%. Ten-year crude survival was 87% after any cancer, 91% if the first cancer was colorectal, 98% if endometrial and 89% if ovarian. Conclusions The four Lynch syndrome-associated genes had different penetrance and expression. Colorectal cancer occurred frequently despite colonoscopic surveillance but resulted in few deaths. Using our data, a website has been established at http://LScarisk.org enabling calculation of cumulative cancer risks as an aid to genetic counselling in Lynch syndrome.Peer reviewe

No difference in penetrance between truncating and missense/aberrant splicing pathogenic variants in mlh1 and msh2: A prospective lynch syndrome database study

Background. Lynch syndrome is the most common genetic predisposition for hereditary cancer. Carriers of pathogenic changes in mismatch repair (MMR) genes have an increased risk of developing colorectal (CRC), endometrial, ovarian, urinary tract, prostate, and other cancers, depending on which gene is malfunctioning. In Lynch syndrome, differences in cancer incidence (penetrance) according to the gene involved have led to the stratification of cancer surveillance. By contrast, any differences in penetrance determined by the type of pathogenic variant remain unknown. Objective. To determine cumulative incidences of cancer in carriers of truncating and missense or aberrant splicing pathogenic variants of the MLH1 and MSH2 genes. Methods. Carriers of pathogenic variants of MLH1 (path_MLH1) and MSH2 (path_MSH2) genes filed in the Prospective Lynch Syndrome Database (PLSD) were categorized as truncating or missense/aberrant splicing according to the InSiGHT criteria for pathogenicity. Results. Among 5199 carriers, 1045 had missense or aberrant splicing variants, and 3930 had truncating variants. Prospective observation years for the two groups were 8205 and 34,141 years, respectively, after which there were no significant differences in incidences for cancer overall or for colorectal cancer or endometrial cancers separately. Conclusion. Truncating and missense or aberrant splicing pathogenic variants were associated with similar average cumulative incidences of cancer in carriers of path MLH1 and path_MSH2.Fil: Dominguez Valentin, Mev. St Mark’s Hospital; Reino Unido. The Norwegian Radium Hospital; Noruega. European Hereditary Tumour Group; Reino UnidoFil: Plazzer, John Paul. St Mark’s Hospital; Reino Unido. The Royal Melbourne Hospital; AustraliaFil: Sampson, Julian R.. European Hereditary Tumour Group; Reino Unido. Cardiff University; Reino UnidoFil: Engel, Christoph. European Hereditary Tumour Group; Reino Unido. Universitat Leipzig; AlemaniaFil: Aretz, Stefan. Universitat Bonn; AlemaniaFil: Jenkins, Mark A.. University of Melbourne; AustraliaFil: Sunde, Lone. Aalborg University; DinamarcaFil: Bernstein, Inge. Aalborg University; DinamarcaFil: Capella, Gabriel. European Hereditary Tumour Group; Reino Unido. St Mark’s Hospital; Reino Unido. Institut Català d’Oncologia; EspañaFil: Balaguer Prunés, Francesc. Universidad de Barcelona; EspañaFil: Macrae, Finlay. European Hereditary Tumour Group; Reino Unido. The Royal Melbourne Hospital; AustraliaFil: Winship, Ingrid M.. University of Melbourne; AustraliaFil: Thomas, Huw. Imperial College London; Reino UnidoFil: Evans, Dafydd Gareth. University of Manchester; Reino UnidoFil: Burn, John. Universidad de Newcastle; Australia. The Royal Melbourne Hospital; Australia. St Mark’s Hospital; Reino UnidoFil: Greenblatt, Marc. University of Vermont; Estados UnidosFil: de Vos tot Nederveen Cappel, Wouter H.. Isala Clinics; Países BajosFil: Sijmons, Rolf H.. University of Groningen; Países Bajos. St Mark’s Hospital; Reino Unido. European Hereditary Tumour Group; Reino UnidoFil: Nielsen, Maartje. Leids Universitair Medisch Centrum; Países BajosFil: Bertario, Lucio. Fondazione IRCCS Istituto Nazionale dei Tumori; ItaliaFil: Bonanni, Bernardo. Fondazione IRCCS Istituto Nazionale dei Tumori; ItaliaFil: Tibiletti, Maria Grazia. Università dell’Insubria; ItaliaFil: Cavestro, Giulia Martina. Vita-Salute San Raffaele University; ItaliaFil: Lindblom, Annika. Karolinska Huddinge Hospital; SueciaFil: Della Valle, Adriana. Hospital Fuerzas Armadas; UruguayFil: Lopez Kostner, Francisco. Clínica Universidad de los Andes; ChileFil: Alvarez, Karin. Clínica Universidad de los Andes; ChileFil: Gluck, Nathan. Universitat Tel Aviv; IsraelFil: Katz, Lior. Sheba Medical Center; IsraelFil: Heinimann, Karl. University Hospital Basel; SuizaFil: Piñero, Tamara Alejandra. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Medicina Traslacional e Ingeniería Biomédica - Hospital Italiano. Instituto de Medicina Traslacional e Ingeniería Biomédica.- Instituto Universitario Hospital Italiano de Buenos Aires. Instituto de Medicina Traslacional e Ingeniería Biomédica; ArgentinaFil: Pavicic, Walter Hernan. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Medicina Traslacional e Ingeniería Biomédica - Hospital Italiano. Instituto de Medicina Traslacional e Ingeniería Biomédica.- Instituto Universitario Hospital Italiano de Buenos Aires. Instituto de Medicina Traslacional e Ingeniería Biomédica; Argentin

Cancer risk and survival in path _ MMR carriers by gene and gender up to 75 years of age: a report from the prospective Lynch Syndrome database

Background Most patients with path_MMR gene variants (Lynch syndrome (LS)) now survive both their first and subsequent cancers, resulting in a growing number of older patients with LS for whom limited information exists with respect to cancer risk and survival. Objective and design This observational, international, multicentre study aimed to determine prospectively observed incidences of cancers and survival in path_MMR carriers up to 75 years of age. Results 3119 patients were followed for a total of 24 475 years. Cumulative incidences at 75 years (risks) for colorectal cancer were 46%, 43% and 15% in path_MLH1, path_MSH2 and path_MSH6 carriers; for endometrial cancer 43%, 57% and 46%; for ovarian cancer 10%, 17% and 13%; for upper gastrointestinal (gastric, duodenal, bile duct or pancreatic) cancers 21%, 10% and 7%; for urinary tract cancers 8%, 25% and 11%; for prostate cancer 17%, 32% and 18%; and for brain tumours 1%, 5% and 1%, respectively. Ovarian cancer occurred mainly premenopausally. By contrast, upper gastrointestinal, urinary tract and prostate cancers occurred predominantly at older ages. Overall 5-year survival for prostate cancer was 100%, urinary bladder 93%, ureter 85%, duodenum 67%, stomach 61%, bile duct 29%, brain 22% and pancreas 0%. Path_PMS2 carriers had lower risk for cancer. Conclusion Carriers of different path_MMR variants exhibit distinct patterns of cancer risk and survival as they age. Risk estimates for counselling and planning of surveillance and treatment should be tailored to each patient's age, gender and path_MMR variant. We have updated our open-access website www.lscarisk.org to facilitate this

Incidence of and survival after subsequent cancers in carriers of pathogenic MMR variants with previous cancer : a report from the prospective Lynch syndrome database

Objective Today most patients with Lynch syndrome (LS) survive their first cancer. There is limited information on the incidences and outcome of subsequent cancers. The present study addresses three questions: (i) what is the cumulative incidence of a subsequent cancer; (ii) in which organs do subsequent cancers occur; and (iii) what is the survival following these cancers? Design Information was collated on prospectively organised surveillance and prospectively observed outcomes in patients with LS who had cancer prior to inclusion and analysed by age, gender and genetic variants. Results 1273 patients with LS from 10 countries were followed up for 7753 observation years. 318 patients (25.7%) developed 341 first subsequent cancers, including colorectal (n=147, 43%), upper GI, pancreas or bile duct (n=37, 11%) and urinary tract (n=32, 10%). The cumulative incidences for any subsequent cancer from age 40 to age 70 years were 73% for pathogenic MLH1 (path_MLH1), 76% for path_MSH2 carriers and 52% for path_MSH6 carriers, and for colorectal cancer (CRC) the cumulative incidences were 46%, 48% and 23%, respectively. Crude survival after any subsequent cancer was 82% (95% CI 76% to 87%) and 10-year crude survival after CRC was 91% (95% CI 83% to 95%). Conclusions Relative incidence of subsequent cancer compared with incidence of first cancer was slightly but insignificantly higher than cancer incidence in patients with LS without previous cancer (range 0.94-1.49). The favourable survival after subsequent cancers validated continued follow-up to prevent death from cancer. The interactive website http://lscarisk.org was expanded to calculate the risks by gender, genetic variant and age for subsequent cancer for any patient with LS with previous cancer.Peer reviewe

Cancer risk and survival in path_MMR carriers by gene and gender up to 75 years of age : a report from the Prospective Lynch Syndrome Database

Background Most patients with path_MMR gene variants (Lynch syndrome (LS)) now survive both their first and subsequent cancers, resulting in a growing number of older patients with LS for whom limited information exists with respect to cancer risk and survival. Objective and design This observational, international, multicentre study aimed to determine prospectively observed incidences of cancers and survival in path_MMR carriers up to 75 years of age. Results 3119 patients were followed for a total of 24 475 years. Cumulative incidences at 75 years (risks) for colorectal cancer were 46%, 43% and 15% in path_MLH1, path_MSH2 and path_MSH6 carriers; for endometrial cancer 43%, 57% and 46%; for ovarian cancer 10%, 17% and 13%; for upper gastrointestinal (gastric, duodenal, bile duct or pancreatic) cancers 21%, 10% and 7%; for urinary tract cancers 8%, 25% and 11%; for prostate cancer 17%, 32% and 18%; and for brain tumours 1%, 5% and 1%, respectively. Ovarian cancer occurred mainly premenopausally. By contrast, upper gastrointestinal, urinary tract and prostate cancers occurred predominantly at older ages. Overall 5-year survival for prostate cancer was 100%, urinary bladder 93%, ureter 85%, duodenum 67%, stomach 61%, bile duct 29%, brain 22% and pancreas 0%. Path_PMS2 carriers had lower risk for cancer. Conclusion C arriers of different path_MMR variants exhibit distinct patterns of cancer risk and survival as they age. Risk estimates for counselling and planning of surveillance and treatment should be tailored to each patient's age, gender and path_MMR variant. We have updated our open-access website www. lscarisk. org to facilitate this.Peer reviewe