Skip to main content
Log in

The transcriptomes of healthy and bitter pit-affected ‘Honeycrisp’ fruit reveal genes associated with disorder development and progression

  • Original Article
  • Published:
Tree Genetics & Genomes Aims and scope Submit manuscript

Abstract

The high-value apple cultivar ‘Honeycrisp’ is prone to bitter pit, a physiological disorder characterized by the development of dark, sunken lesions on fruit particularly towards the calyx-end. Identifying the genetic basis of bitter pit is critical for mitigation of the disorder and development of bitter pit-free varieties. To this end, we performed an RNA sequencing experiment to compare the peel and flesh transcriptomes of healthy and bitter pit-affected ‘Honeycrisp’ fruit, with the objective of identifying bitter pit candidate genes. A similar number of genes were expressed in flesh and peel (~ 27,000), and we detected flesh- and peel-specific expression. Within the flesh and peel datasets, 2.6% and 1% of genes were differentially expressed between healthy and bitter pit apples, respectively. We identified over 1000 differentially expressed genes in both flesh and peel datasets that are distributed throughout the genome, sixteen of which are within a previously identified bitter pit QTL, making them potential candidate genes. Both MdMADS5 and MdIDL1, the orthologs of Arabidopsis APETALA1 and INFLORESCENCE DEFICIENT IN ABSCISSION-LIKE 1, respectively, which are possibly involved in fruit development and organ abscission, stand out as candidate genes for bitter pit, based on their proposed function, differential expression, and location within a bitter pit QTL. The RNA-seq data also revealed that many genes involved with defense response and stress are upregulated in bitter pit-affected tissues. We discuss how these differentially expressed genes could provide important information about the physiology, development, and progression of bitter pit.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2

Similar content being viewed by others

References

  • Anders S, Huber W (2010) Differential expression analysis for sequence count data. Genom Bio 11:R106

    Article  CAS  Google Scholar 

  • Askew HO, Chittenden ET, Monk RJ, Watson J (1960) Chemical investigations on bitter pit of apples. NZ J Ag Res 1:169–178

    Article  Google Scholar 

  • Bai Y, Dougherty L, Xu K (2014) Towards an improved apple reference transcriptome using RNA-seq. Mol Gen and Genom 289:427–438

    Article  CAS  Google Scholar 

  • Bai Y, Dougherty L, Cheng L, Zhong GY, Xu K (2015) Uncovering co-expression gene network modules regulating fruit acidity in diverse apples. BMS Genom 16:612

    Article  CAS  Google Scholar 

  • Ballester AR, Norelli J, Burchard E, Abdelfattah A, Levin E, González-Candelas L, Droby S, Wisniewski M (2017) Transcriptomic response of resistant (PI613981 - Malus sieversii) and susceptible (‘Royal Gala’) genotypes of apple to blue mold (Penicillium expansum) infection. Frontin P Sci 8:1981

    Article  Google Scholar 

  • Baugher TA, Marini R, Schupp JR, Watkins CB (2017) Prediction of bitter pit in ‘Honeycrisp’ apples and best management implications. Hort Sci 52:1368–1374

    CAS  Google Scholar 

  • Botton A, Lezzer P, Dorigoni A, Barcaccia G, Ruperti B, Ramina A (2008) Genetic and environmental factors affecting allergen-related gene expression in apple fruit (Malus domestica L. Borkh). J Ag Food Chem 56:6707–6716

    Article  CAS  Google Scholar 

  • Busatto N, Farneti B, Commisso M, Bianconi M, Iadarola B, Zago E, Ruperti B, Spinelli F, Zanella A, Velasco R, Ferrarini A (2018) Apple fruit superficial scald resistance mediated by ethylene inhibition is associated with diverse metabolic processes. Plant J 93:270–285

    Article  CAS  PubMed  Google Scholar 

  • Butenko MA, Patterson SE, Grini PE, Stenvik GE, Amundsen SS, Mandal A, Aalen RB (2003) INFLORESCENCE DEFICIENT IN ABSCISSION controls floral organ abscission in Arabidopsis and identifies a novel family of putative ligands in plants. Plant Cell 15:2296–2307

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Buti M, Poles L, Caset D, Magnago P, Fernandez Fernandez F, Colgan RJ, Velasco R, Sargent DJ (2015) Identification and validation of a QTL influencing bitter pit symptoms in apple (Malus × domestica). Mol Breed 35:29

    Article  Google Scholar 

  • Buti M, Sargent DJ, Bianco L, Magnago P, Velasco R, Colgan RJ (2018) A study of gene expression changes at the Bp-2 locus associated with bitter pit symptom expression in apple (Malus pumila). Mol Breed 38:85

    Article  CAS  Google Scholar 

  • Carne WM, Pittman HA, Elliot HG (1929) Bitter pit of apple in Australia. Bul CSIRO (austral) 4:1–23

    Google Scholar 

  • Cevik C, Ryder CD, Popovich A, Manning K, King G, Seymour G (2010) A FRUITFULL-like gene is associated with genetic variation for fruit flesh firmness in apple (Malus domestica Borkh.). Tree Gen Genom 6:271–279

    Article  Google Scholar 

  • Cheng L, Raba R (2009) Accumulation of macro- and micronutrients and nitrogen demand-supply relationship of ‘Gala’/’Malling 26’ trees grown in sand culture. J Amer Soc Hort Sci 134:3–13

    Article  Google Scholar 

  • Cheng L, Sazo M (2018) Why Is ‘Honeycrisp’ so susceptible to bitter pit? NY Fruit Quarterly 26:19–23

    Google Scholar 

  • Daccord N, Celton JM, Linsmith G, Becker C, Choisne N, Schijlen E, van de Geest H, Bianco L, Micheletti D, Velasco R, Di Pierro EA (2017) High-quality de novo assembly of the apple genome and methylome dynamics of early fruit development. Nat Gen 49:1099–1108

    Article  CAS  Google Scholar 

  • De Freitas ST, Amarante CVT, Labavitch JM, Mitcham E (2010) Cellular approach to understand bitter pit development in apple fruit. Postharvest Biol Tech 57:6–13

    Article  CAS  Google Scholar 

  • De Freitas ST, Mitcham EJ (2012) Factors involved in fruit calcium deficiency disorders. Hort Rev 40:107–146

    Article  Google Scholar 

  • Di Guardo M, Tadiello A, Farneti B, Lorenz G, Masuero D, Vrhovsek U, Costa G, Velasco R, Costa F (2013) A multidisciplinary approach providing new insight into fruit flesh browning physiology in apple (Malus x domestica Borkh.). PLos one 8:e78004

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Dražeta L, Lang A, Hall AJ, Volz RK, Jameson PE (2004) Causes and effects of changes in xylem functionality in apple fruit. Ann Bot 93:275–282

    Article  PubMed  PubMed Central  Google Scholar 

  • Everaert C, Luypaert M, Maag JL, Cheng QX, Dinger ME, Hellemans J, Mestdagh P (2017) Benchmarking of RNA-sequencing analysis workflows using whole-transcriptome RT-qPCR expression data. Sci Rep 7:1–11

    Article  CAS  Google Scholar 

  • Ferguson IB, Watkins CB (1989) Bitter pit in apple fruit. Hort Rev 11:289–355

    CAS  Google Scholar 

  • Fernandes H, Michalska K, Sikorski M, Jaskolski M (2013) Structural and functional aspects of PR-10 proteins. FEBS J 280:1169–1199

    Article  CAS  PubMed  Google Scholar 

  • Gao ZS, Van de Weg WE, Schaart JG, Schouten HJ, Tran DH, Kodde LP, Van der Meer IM, Van der Geest AH, Kodde J, Breiteneder H, Hoffmann-Sommergruber K (2004) Genomic cloning and linkage mapping of the Mal d 1 (PR-10) gene family in apple (Malus domestica). TAG 111:171–183

    Article  CAS  Google Scholar 

  • Gapper NE, Hertog ML, Lee J, Buchanan DA, Leisso RS, Fei Z, Qu G, Giovannoni JJ, Johnston JW, Schaffer RJ, Nicolaï BM (2017) Delayed response to cold stress is characterized by successive metabolic shifts culminating in apple fruit peel necrosis. BMC P Bio 17:77

    Article  CAS  Google Scholar 

  • Gasic K, Hernandez A, Korban SS (2004) RNA extraction from different apple tissues rich in polyphenols and polysaccharides for cDNA library construction. Plant Mol Biol Rep 22:437a–437g

    Article  Google Scholar 

  • Graether SP, Boddington KF (2014) Disorder and function: a review of the dehydrin protein family. Front Plant Sci 5:576

    Article  PubMed  PubMed Central  Google Scholar 

  • Gu Q, Ferrandiz C, Yanofsky MF, Martienssen R (1998) The FRUITFULL MADS-box gene mediates cell differentiation during Arabidopsis fruit development. Development 125:1509–1517

    Article  CAS  PubMed  Google Scholar 

  • Hocking B, Tyerman SD, Burton RA, Gilliham M (2016) Fruit calcium: transport and physiology. Front Plant Sci 7:569

    Article  PubMed  PubMed Central  Google Scholar 

  • Jemrić T, Fruk I, Fruk M, Radman S, Sinkovic L, Fruk G (2017) Bitter pit in apples: pre-and postharvest factors: a review. Spanish J Ag Res 14:e08R01

    Article  Google Scholar 

  • Kalcsits L, van der Heijden G, Reid M, Mullin K (2017) Calcium absorption during fruit development in ‘Honeycrisp’ apple measured using 44Ca as a stable isotope tracer. Hort Sci 52:1804–1809

    Google Scholar 

  • Kaufmann K, Wellmer F, Muiño JM, Ferrier T, Wuest SE, Kumar V, Serrano-Mislata A, Madueno F, Krajewski P, Meyerowitz EM, Angenent GC (2010) Orchestration of floral initiation by APETALA1. Science 328:85–89

    Article  CAS  PubMed  Google Scholar 

  • Kotoda N, Wada M, Komori S, Kidou S, Abe K, Masuda T, Soejima J (2000) Expression pattern of homologues of floral meristem identity genes LFY and AP1 during flower development in apple. JASHS 125:398–403

    Article  CAS  Google Scholar 

  • Krawitzky M, Orera I, Lopez-Millan AF, Oria R, Val J (2016) Identification of bitter pit protein markers in Malus domestica using differential in-gel electrophoresis (DIGE) and LC–MS/MS. Postharvest Bio Tech 111:224–239

    Article  CAS  Google Scholar 

  • Kumar S, Garrick DJ, Bink M, Whitworth C, Chagné D, Volz V (2013) Novel genomic approaches unravel genetic architecture of complex traits in apple. BMC Genomics 14:393

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Langmead B, Trapnell C, Pop M, Salzberg SL (2009) Ultrafast and memory efficient alignment of short DNA sequences to the human genome. Genome Biol 10:R25

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  • Liang D, Xia H, Wu S, Ma F (2012) Genome-wide identification and expression profiling of dehydrin gene family in Malus domestica. Mol Bio Rep 39:10759–10768

    Article  CAS  Google Scholar 

  • Li L, Steffens JC (2002) Overexpression of polyphenol oxidase in transgenic tomato plants results in enhanced bacterial disease resistance. Planta 215:239–247

    Article  CAS  PubMed  Google Scholar 

  • Liu D, Meng S, Xiang Z, He N, Yang G (2019) Antimicrobial mechanism of reaction products of Morus notabilis (mulberry) polyphenol oxidases and chlorogenic acid. Phytochemistry 163:1–10

    Article  CAS  PubMed  Google Scholar 

  • Liu JJ, Ekramoddoullah A (2006) The family 10 of plant pathogenesis-related proteins: their structure, regulation, and function in response to biotic and abiotic stresses. Phys and Mol Plant Path 68:3–13

    Article  CAS  Google Scholar 

  • Lu Y, Kong J, Li T, Wang Y, Wang A, Han Z (2013) Isolation and characterization of an APETALA1-like gene from pear (Pyrus pyrifolia). Plant Mol Bio Rep 31:1031–1039

    Article  CAS  Google Scholar 

  • Marzban G, Puehringer H, Dey R, Brynda S, Ma Y, Martinelli A, Zaccarini M, Van Der Weg E, Housley Z, Kolarich D, Altmann F, Laimer M (2005) Localisation and distribution of the major allergens in apple fruits. Plant Sci 169:387–394

    Article  CAS  Google Scholar 

  • Matthes A, Schmitz-Eiberger M (2009) Apple (Malus domestica L. Borkh.) allergen Mal d 1: effect of cultivar, cultivation system, and storage conditions. J Ag Food Chem 57:10548–10553

    Article  CAS  Google Scholar 

  • Mellidou I, Buts K, Hatoum D, Ho QT, Johnston JW, Watkins CB, Schaffer RJ, Gapper NE, Giovannoni JJ, Rudell DR, Hertog ML (2014) Transcriptomic events associated with internal browning of apple during postharvest storage. BMC Plant Bio 14:328

    Article  CAS  Google Scholar 

  • Melville F, Hardisty SE (1960) Bitter pit: a progress report on the use of calcium nitrate sprays for its control. J Dept Ag West Aus 1:1017–1019

    CAS  Google Scholar 

  • Miqueloto A, Amarante CVT, Steffens CA, Santos A, Mitcham E (2014) Relationship between xylem functionality, calcium content and the incidence of bitter pit in apple fruit. Sci Hortic 165:319–323

    Article  CAS  Google Scholar 

  • Monroe JD, Storm AR (2018) Review: The Arabidopsis β-amylase (BAM) gene family: diversity of form and function. Plant Sci 276:163–170

    Article  CAS  PubMed  Google Scholar 

  • Muñoz C, Hoffmann T, Escobar NM, Ludemann F, Botella MA, Valpuesta V, Schwab W (2010) The strawberry fruit Fra a allergen functions in flavonoid biosynthesis. Mol Plant 3:113–124

    Article  PubMed  CAS  Google Scholar 

  • Orcheski B, Brown S (2017) High-throughput sequencing reveals that pale green lethal disorder in apple (Malus) stimulates stress responses and affects senescence. Tree Genetics Genomes 13:9

    Article  Google Scholar 

  • Pratt C (1988) Apple flower and fruit: morphology and anatomy. Hort Rev 10:273–307

    Google Scholar 

  • Pühringer H, Moll D, Hoffmann-Sommergruber K, Watillon B, Katinger H, da Câmara Machado ML (2000) The promoter of an apple Ypr10 gene, encoding the major allergen Mal d 1, is stress- and pathogen-inducible. Plant Sci 152:35–50

    Article  Google Scholar 

  • Steidle EA (2010) Investigation of the role of BAM9 in starch metabolism in Arabidopsis thaliana. (Masters theses, James Madison University, Harrisonburg, Virginia, USA). Retrieved from: https://commons.lib.jmu.edu/master201019/384/ 

  • Tabuchi T, Ito S, Arai N (2001) Anatomical studies of the abscission process in the tomato pedicels at flowering stage. J Japan Soc Hort Sci 70:63–65

    Article  Google Scholar 

  • Taranto F, Pasqualone A, Mangini G, Tripodi P, Miazzi M, Pavan S, Montemurro C (2017) Polyphenol oxidases in crops: biochemical, physiological and genetic aspects. Int J Mol Sci 18:377

    Article  PubMed Central  CAS  Google Scholar 

  • Telias A, Hoover E, Rosen C, Bedford D, Cook D (2006) The effect of calcium sprays and fruit thinning on bitter pit incidence and calcium content in ‘Honeycrisp’ apple. J Plant Nut 29:1941–1957

    Article  CAS  Google Scholar 

  • Thipyapong P, Hunt MD, Steffens JC (2004) Antisense downregulation of polyphenol oxidase results in enhanced disease susceptibility. Planta 220:105–117

    Article  CAS  PubMed  Google Scholar 

  • Tian X, Zhang L, Feng S, Zhao Z, Wang X, Gao H (2019) Transcriptome analysis of apple leaves in response to powdery mildew (Podosphaera leucotricha) infection. Int J Mol Sci 20:2326

    Article  CAS  PubMed Central  Google Scholar 

  • Tucker ML, Yang R (2012) IDA-like gene expression in soybean and tomato leaf abscission and requirement for a diffusible stelar abscission signal. AoB Plants 2012:pls035

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  • Serra S, Leisso R, Giordani L, Kalcsits L, Musacchi S (2016) Crop load influences fruit quality, nutritional balance, and return bloom in ‘Honeycrisp’apple. Hort Sci 51:236–244

    Google Scholar 

  • Schonherr J, Bukovac MJ (1973) Ion exchange properties of isolated tomato fruit cuticular membrane: exchange capacity, nature of fixed charges and cation selectivity. Planta 109:73–93

    Article  CAS  PubMed  Google Scholar 

  • Serban C, Kalcsits L, DeEll J, Mattheis JP (2019) Responses of ‘Honeycrisp’ apples to short-term controlled atmosphere storage established during temperature conditioning. HortSci 54:1532–1539

    Article  CAS  Google Scholar 

  • Shi CL, von Wangenheim D, Herrmann U, Wildhagen M, Kulik I, Kopf A, Ishida T, Olsson V, Anker MK, Albert M, Butenko MA, Felix G, Sawa S, Claassen M, Friml J, Aalen RB (2018) The dynamics of root cap sloughing in Arabidopsis is regulated by peptide signaling. Nat Plants 4:596–604

    Article  CAS  PubMed  Google Scholar 

  • Simons RK, Chu MC (1980) Scanning electron microscopy and electron microprobe studies of bitter pit apples. Acta Hort 92:57–69

    Article  Google Scholar 

  • Stenvik GE, Butenko MA, Urbanowicz BR, Rose JCK, Aalen RB (2006) Overexpression of INFLORESCENCE DEFICIENT IN ABSCISSION activates cell separation in vestigial abscission zones in Arabidopsis. Plant Cell 18:1467–1476

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Ubi BE, Saito T, Bai S, Nishitani C, Ban Y, Ikeda K, Ito A, Moriguchi T (2013) Characterization of 10 MADS-box genes from Pyrus pyrifolia and their differential expression during fruit development and ripening. Gene 528:183–194

    Article  CAS  PubMed  Google Scholar 

  • Udrardi MK, Czechowski T, Scheible WR (2008) Eleven golden rules of quantitative RT-PCR. Plant Cell 20:1736–1737

    Article  CAS  Google Scholar 

  • Velasco R, Zharkikh A, Affourtit J, Dhingra A, Cestaro A, Kalyanaraman A, Fontana P, Bhatnagar SK, Troggio M, Pruss D, Salvi S, Pindo M, Baldi P, Castelletti S, Cavaiuolo M, Coppola G, Costa F, Cova V, Dal Ri A, Goremykin V, Komjanc M, Longhi S, Magnago P, Malacarne G, Malnoy M, Micheletti D, Moretto M, Perazzolli M, Si-Ammour A, Vezzulli S, Zini E, Eldredge G, Fitzgerald LM, Gutin N, Lanchbury J, Macalma T, Mitchell JT, Reid J, Wardell B, Kodira C, Chen Z, Desany B, Niazi F, Palmer M, Koepke T, Jiwan D, Schaeffer S, Krishnan V, Wu C, Chu VT, King ST, Vick J, Tao Q, Mraz A, Stormo A, Stormo K, Bogden R, Ederle D, Stella A, Vecchietti A, Kater MM, Masiero S, Lasserre P, Lespinasse Y, Allan AC, Bus V, Chagné D, Crowhurst RN, Gleave AP, Lavezzo E, Fawcett JA, Proost S, Rouzé P, Sterck L, Toppo S, Lazzari B, Hellens RP, Durel CE, Gutin A, Bumgarner RE, Gardiner SE, Skolnick M, Egholm M, Van de Peer Y, Salamini F, Viola R (2010) The genome of the domesticated apple (Malus × domestica Borkh.). Nature Genet 42:833–839

    Article  CAS  PubMed  Google Scholar 

  • Wang X, Hou S, Wu Q, Lin M, Acharya BR, Wu D, Zhang W (2017) IDL6-HAE/HSL2 impacts pectin degradation and resistance to Pseudomonas syringae pv tomato DC3000 in Arabidopsis leaves. Plant J 89:250–263

    Article  CAS  PubMed  Google Scholar 

  • Watkins CB, Nock JF, Weis SA, Jayanty S, Beaudry RM (2004) Storage temperature, diphenylamine, and pre-storage delay effects on soft scald, soggy breakdown and bitter pit of ‘Honeycrisp’ apples. Postharvest Bio Tech 32:213–221

    Article  CAS  Google Scholar 

  • Yao JL, Dong YH, Kvarnheden A, Morris B (1999) Seven MADS-box genes in apple are expressed in different parts of the fruit. JASHS 124:8–13

    Article  CAS  Google Scholar 

  • Yermiyahu U, Nir S, Ben-Hayyim G, Kafkafi U (1994) Quantitative competition of calcium with sodium or magnesium for sorption sites on plasma membrane vesicles of melon (Cucumis melo L.) root cells. J Membr Biol 138:55–63

    Article  CAS  PubMed  Google Scholar 

  • Zhong S, Joung JG, Zheng Y, Chen YR, Liu B, Shao Y, Xiang JZ, Fei Z, Giovannoni JJ (2011) High-throughput Illumina strand-specific RNA sequencing library preparation. CSH Pro 2011:pp.pdb-prot5652

    Google Scholar 

  • Zupan A, Mikulic-Petkovsek M, Cunja V, Stampar F, Veberic R (2013) Comparison of phenolic composition of healthy apple tissues and tissues affected by bitter pit. J Agric Food Chem 61:12066–12071

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgements

We thank Cherry Lawn Fruit Farms in Western New York for their cooperation on fruit sample collection.

Funding

This work is supported in part by USDA National Institute of Food and Agriculture—Specialty Crop Research Initiative project “AppleRoot2Fruit: Accelerating the development, evaluation and adoption of new apple rootstocks” (2016–51181-25406) and New York Apple Research and Development Program.

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Benjamin Orcheski.

Ethics declarations

Conflict of interest

The authors declare no competing interests.

Data archiving statement

Raw transcriptome sequencing reads have been deposited into the NCBI BioProject database under the accession PRJNA725518.

Additional information

Communicated by A.M. Dandekar

Publisher’s note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Supplementary Information

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Orcheski, B., Meng, D., Bai, Y. et al. The transcriptomes of healthy and bitter pit-affected ‘Honeycrisp’ fruit reveal genes associated with disorder development and progression. Tree Genetics & Genomes 17, 37 (2021). https://doi.org/10.1007/s11295-021-01518-7

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • DOI: https://doi.org/10.1007/s11295-021-01518-7

Keywords

Navigation