Structural and functional insights into the mechanism of action of plant borate transporters

Savvas Saouros,Yilmaz Alguel,Thotegowdanapalya C Mohan,Alexandra M E Jones,Paul Simpson,Bernadette Byrne,Silke Lehmann,Alexander D Cameron,Joseph D Barrit,Nicola J Scull,Cristina Cecchetti

doi:10.1038/s41598-021-91763-6

Savvas Saouros, Yilmaz Alguel + Show 9 more

Open Access

https://doi.org/10.1038/s41598-021-91763-6

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Abstract

Boron has essential roles in plant growth and development. BOR proteins are key in the active uptake and distribution of boron, and regulation of intracellular boron concentrations. However, their mechanism of action remains poorly studied. BOR proteins are homologues of the human SLC4 family of transporters, which includes well studied mammalian transporters such as the human Anion Exchanger 1 (hAE1). Here we generated Arabidopsis thaliana BOR1 (AtBOR1) variants based (i) on known disease causing mutations of hAE1 (S466R, A500R) and (ii) a loss of function mutation (D311A) identified in the yeast BOR protein, ScBOR1p. The AtBOR1 variants express in yeast and localise to the plasma membrane, although both S466R and A500R exhibit lower expression than the WT AtBOR1 and D311A. The D311A, S466R and A500R mutations result in a loss of borate efflux activity in a yeast bor1p knockout strain. A. thaliana plants containing these three individual mutations exhibit substantially decreased growth phenotypes in soil under conditions of low boron. These data confirm an important role for D311 in the function of the protein and show that mutations equivalent to disease-causing mutations in hAE1 have major effects in AtBOR1. We also obtained a low resolution cryo-EM structure of a BOR protein from Oryza sativa, OsBOR3, lacking the 30 C-terminal amino acid residues. This structure confirms the gate and core domain organisation previously observed for related proteins, and is strongly suggestive of an inward facing conformation.

Highlights

Boron has essential roles in plant growth and development
Mutations on Arabidopsis thaliana BOR1 (AtBOR1) were selected based on homology with human Anion Exchanger 1 (hAE1)
D311 (Fig. 1) is a possible candidate H+ binding residue and mutation of the equivalent residue (D347) in S. cerevisiae has previously been shown to inhibit borate efflux function in a yeast based functional complementation assay[25]

Summary

Introduction

BOR proteins are key in the active uptake and distribution of boron, and regulation of intracellular boron concentrations. A. thaliana plants containing these three individual mutations exhibit substantially decreased growth phenotypes in soil under conditions of low boron. These data confirm an important role for D311 in the function of the protein and show that mutations equivalent to disease-causing mutations in hAE1 have major effects in AtBOR1. Under conditions of B sufficiency OsBOR1 expression is only detectable in the stele[20] In both species BOR1 allows passage of B though the Casparian strips enabling active efflux of B from the root cells to the xylem[20]

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Conclusion