Pineapple SWEET10 is a glucose transporter

Abstract

SWEET transporters are a unique class of sugar transporters that play vital roles in various developmental and physiological processes in plants. While the functions of SWEETs have been well established in model plants such as Arabidopsis, their functions in economically important fruit crops like pineapple have not been well studied. Here we aimed to investigate the substrate specificity of pineapple SWEETs by comparing the protein sequences of known glucose and sucrose transporters in Arabidopsis with those in pineapple. Our genome-wide approach and 3D structure comparison showed that the Arabidopsis SWEET8 homolog in pineapple, AcSWEET10, shares similar sequences and protein properties responsible for glucose transport. To determine the functional conservation of AcSWEET10, we tested its ability to complement glucose transport mutants in yeast and analyzed its expression in stamens and impact on the microspore phenotype and seed set in transgenic Arabidopsis. The results showed that AcSWEET10 is functionally equivalent to AtSWEET8 and plays a critical role in regulating microspore formation through the regulation of the Callose synthase5 ( CalS5), which highlights the importance of SWEET transporters in pineapple. This information could have important implications for improving fruit crop yield and quality by manipulating SWEET transporter activity.

Acknowledgements

We especially thank Dr Binghua Wu (Fujian Agriculture and Forestry University, China) for kindly providing the yeast mutant strain EBY.VW4000 and Prof. Zhong-Nan Yang (Shanghai Normal University, China) for sharing Atsweet8 seeds. This work was supported by the Science and Technology Major Project of Guangxi (Gui Ke AA22068096), the Science and Technology Innovation Project of Pingtan Science and Technology Research Institute (PT2021007, PT2021003), the General Project of Fujian Natural Science Foundation (2020 J01594), the Project of Guangxi Featured Fruit Innovation Team on Pineapple Breeding and Cultivation Post under the National Modern Agricultural Industry Technology System (nycytxgxcxtd-17-05), and the Guangxi Academy of Agricultural Sciences Basic Research Project (Gui Nong Ke 2021YT046). The funding bodies played no role in the design of the study and collection, analysis and interpretation of the data, and writing the manuscript.

Author contributions

M.A. and Y. Q. designed the study. B.F. performed all the experiments except RNA-seq. L.W., X.W., and P.Z. performed RNA-seq. B.F, M.A.A., M.A., and Y.Q. analyzed the data and wrote the paper. All authors have read and approved the manuscript.

Data availability

The datasets analyzed in this study are publicly available with the accession numbers PRJNA331052, PRJNA305042, and PRJEB38680.

Conflict of interest

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

References

[1] Hedrich R, Sauer N, Neuhaus HE . Sugar transport across the plant vacuolar membrane: nature and regulation of carrier proteins. Curr Opin Plant Biol. 2015; 25: 63-70

[2] Johnson DA, Thomas MA . The monosaccharide transporter gene family in Arabidopsis and rice: a history of duplications, adaptive evolution, and functional divergence . Mol Biol Evol. 2007; 24: 2412-23

[3] Büttner M, Sauer N . Monosaccharide transporters in plants: structure, function and physiology. Biochim Biophys Acta. 2000; 1465: 263-74

[4] Li X, Si W, Qin Q et al. Deciphering evolutionary dynamics of SWEET genes in diverse plant lineages. Sci Rep. 2018; 8: 13440

[5] Xue X, Wang J, Shukla D et al. When SWEETs turn tweens: updates and perspectives. Annu Rev Plant Biol. 2022; 73: 379-403

[6] Chen L-Q, Qu XQ, Hou BH et al. Sucrose efflux mediated by SWEET proteins as a key step for phloem transport. Science. 2012; 335: 207-11

[7] Chen LQ, Hou BH, Lalonde S et al. Sugar transporters for intercellular exchange and nutrition of pathogens. Nature. 2010; 468: 527-32

[8] Chen L-Q, Lin IW, Qu XQ et al. A cascade of sequentially expressed sucrose transporters in the seed coat and endosperm provides nutrition for the Arabidopsis embryo . Plant Cell. 2015; 27: 607-19

[9] Zhang X, Feng C, Wang M et al. Plasma membrane-localized SlSWEET7a and SlSWEET14 regulate sugar transport and storage in tomato fruits. Hortic Res. 2021; 8: 186

[10] Klemens PAW, Patzke K, Deitmer J et al. Overexpression of the vacuolar sugar carrier AtSWEET16 modifies germination, growth, and stress tolerance in Arabidopsis . Plant Physiol. 2013; 163: 1338-52

[11] Chardon F, Bedu M, Calenge F et al. Leaf fructose content is controlled by the vacuolar transporter SWEET17 in Arabidopsis . Curr Biol. 2013; 23: 697-702

[12] Keller R, Ziegler C, Schneider D . When two turn into one: evolution of membrane transporters from half modules. Biol Chem. 2014; 395: 1379-88

[13] Xu Y, Tao Y, Cheung LS et al. Structures of bacterial homologues of SWEET transporters in two distinct conformations. Nature. 2014; 515: 448-52

[14] Lee Y, Nishizawa T, Yamashita K et al. Structural basis for the facilitative diffusion mechanism by SemiSWEET transporter. Nat Commun. 2015; 6: 6112

[15] Xuan YH, Hu YB, Chen LQ et al. Functional role of oligomerization for bacterial and plant SWEET sugar transporter family. Proc Natl Acad Sci USA. 2013; 110: E3685-94

[16] Feng L, Frommer WB . Structure and function of SemiSWEET and SWEET sugar transporters. Trends Biochem Sci. 2015; 40: 480-6

[17] Guo WJ, Nagy R, Chen HY et al. SWEET17, a facilitative transporter, mediates fructose transport across the tonoplast of Arabidopsis roots and leaves . Plant Physiol. 2014; 164: 777-89

[18] Selvam B, Yu Y-C, Chen L-Q et al. Molecular basis of the glucose transport mechanism in plants. ACS Cent Sci. 2019; 5: 1085-96

[19] Wang J, Yan C, Li Y et al. Crystal structure of a bacterial homologue of SWEET transporters. Cell Res. 2014; 24: 1486-9

[20] Lin IW, Sosso D, Chen LQ et al. Nectar secretion requires sucrose phosphate synthases and the sugar transporter SWEET9. Nature. 2014; 508: 546-9

[21] Wang J, Xue X, Zeng H et al. Sucrose rather than GA transported by AtSWEET13 and AtSWEET14 supports pollen fitness at late anther development stages. New Phytol. 2022; 236: 525-37

[22] Guan YF, Huang XY, Zhu J et al. RUPTURED POLLEN GRAIN1, a member of the MtN3/saliva gene family, is crucial for exine pattern formation and cell integrity of microspores in Arabidopsis . Plant Physiol. 2008; 147: 852-63

[23] Han L, Zhu Y, Liu M et al. Molecular mechanism of substrate recognition and transport by the AtSWEET13 sugar transporter. Proc Natl Acad Sci USA. 2017; 114: 10089-94

[24] Sun MX, Huang XY, Yang J et al. Arabidopsis RPG1 is important for primexine deposition and functions redundantly with RPG2 for plant fertility at the late reproductive stage. Plant Reprod. 2013; 26: 83-91

[25] Gautam T, Dutta M, Jaiswal V . et al. Emerging roles of SWEET sugar transporters in plant development and abiotic stress responses . Cell. 2022; 11: 1303

[26] Braun DM . Phloem loading and unloading of sucrose: what a long, strange trip from source to sink. Annu Rev Plant Biol. 2022; 73: 553-84

[27] Isoda R, Palmai Z, Yoshinari A et al. SWEET13 transport of sucrose, but not gibberellin, restores male fertility in Arabidopsis sweet13;14 . Proc Natl Acad Sci USA. 2022; 119: e2207558119

[28] Fakher B, Jakada BH, Greaves JG . et al. Identification and expression analysis of pineapple sugar transporters reveal their role in the development and environmental response. Front Plant Sci. 2022; 13: 964897

[29] Wu LB, Eom JS, Isoda R et al. OsSWEET11b, a potential sixth leaf blight susceptibility gene involved in sugar transport-dependent male fertility. New Phytol. 2022; 234: 975-89

[30] Breia R, Conde A, Badim H et al. Plant SWEETs: from sugar transport to plant-pathogen interaction and more unexpected physiological roles. Plant Physiol. 2021; 186: 836-52

[31] Eom JS, Chen LQ, Sosso D et al. SWEETs, transporters for intracellular and intercellular sugar translocation. Curr Opin Plant Biol. 2015; 25: 53-62

[32] Ji J, Yang L, Fang Z . et al. Plant SWEET family of sugar transporters: structure, evolution and biological functions. Biomol Ther. 2022; 12: 205

[33] Chen LQ, Cheung LS, Feng L et al. Transport of sugars. Annu Rev Biochem. 2015; 84: 865-94

[34] Tao Y, Cheung LS, Li S et al. Structure of a eukaryotic SWEET transporter in a homotrimeric complex. Nature. 2015; 527: 259-63

[35] Yamada K, Osakabe Y, Yamaguchi-Shinozaki K . A C-terminal motif contributes to the plasma membrane localization of Arabidopsis STP transporters . PLoS One. 2017; 12: e0186326

[36] Nobukuni M, Mochizuki H, Okada S et al. The C-terminal region of serotonin transporter is important for its trafficking and glycosylation. J Pharmacol Sci. 2009; 111: 392-404

[37] Fujita S, Sato D, Kasai H et al. The C-terminal region of the yeast monocarboxylate transporter Jen1 acts as a glucose signal-responding degron recognized by the α-arrestin Rod1 . J Biol Chem. 2018; 293: 10926-36

[38] Tamura K, Stecher G, Peterson D et al. MEGA6: Molecular Evolutionary Genetics Analysis version 6.0. Mol Biol Evol. 2013; 30: 2725-9

[39] Kumar S, Stecher G, Tamura K . MEGA7: Molecular Evolutionary Genetics Analysis version 7.0 for bigger datasets. Mol Biol Evol. 2016; 33: 1870-4

[40] Omasits U, Ahrens CH, Müller S et al. Protter: interactive protein feature visualization and integration with experimental proteomic data. Bioinformatics. 2014; 30: 884-6

[41] Jumper J, Evans R, Pritzel A et al. Highly accurate protein structure prediction with AlphaFold. Nature. 2021; 596: 583-9

[42] Clough SJ, Bent AF . Floral dip: a simplified method for Agrobacterium-mediated transformation of Arabidopsis thaliana . Plant J. 1998; 16: 735-43

[43] Aslam M, Fakher B, Jakada BH . et al. Genome-wide identification and expression profiling of CBL-CIPK gene family in pineapple (Ananas comosus) and the role of AcCBL1 in abiotic and biotic stress response . Biomol Ther. 2019; 9: 293

[44] Wang L, Li Y, Jin X et al. Floral transcriptomes reveal gene networks in pineapple floral growth and fruit development. Commun Biol. 2020; 3: 500

[45] Livak KJ, Schmittgen TD . Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) method. Methods. 2001; 25: 402-8

[46] Bashandy H, Jalkanen S, Teeri TH . Within leaf variation is the largest source of variation in agroinfiltration of Nicotiana benthamiana . Plant Methods. 2015; 11: 47

[47] Peterson. R, Slovin J, Chen C . A simplified method for differential staining of aborted and non-aborted pollen grains. Int J Plant Biol. 2010; 1: e13

[48] Karimi M, Inzé D, Depicker A . GATEWAY™vectors for Agrobacterium-mediated plant transformation . Trends Plant Sci. 2002; 7: 193-5

[49] Wieczorke R, Krampe S, Weierstall T et al. Concurrent knockout of at least 20 transporter genes is required to block uptake of hexoses in Saccharomyces cerevisiae . FEBS Lett. 1999; 464: 123-8

[50] Guo C, Li H, Xia X et al. Functional and evolution characterization of SWEET sugar transporters in Ananas comosus. Biochem Biophys Res Commun. 2018; 496: 407-14