The aromatic/arginine selectivity filter of NIP aquaporins plays a critical role in substrate selectivity for silicon, boron, and arsenic

Abstract

Nodulin-26-like intrinsic proteins (NIPs) of the aquaporin family are involved in the transport of diverse solutes, but the mechanisms controlling the selectivity of transport substrates are poorly understood. The purpose of this study was to investigate how the aromatic/arginine (ar/R) selectivity filter influences the substrate selectivity of two NIP aquaporins; the silicic acid (Si) transporter OsLsi1 (OsNIP2;1) from rice and the boric acid (B) transporter AtNIP5;1 from Arabidopsis; both proteins are also permeable to arsenite. Native and site-directed mutagenized variants of the two genes were expressed in Xenopus oocytes and the transport activities for Si, B, arsenite, and water were assayed. Substitution of the amino acid at the ar/R second helix (H2) position of OsLsi1 did not affect the transport activities for Si, B, and arsenite, but that at the H5 position resulted in a total loss of Si and B transport activities and a partial loss of arsenite transport activity. Conversely, changes of the AtNIP5;1 ar/R selectivity filter and the NPA motifs to the OsLsi1 type did not result in a gain of Si transport activity. B transport activity was partially lost in the H5 mutant but unaffected in the H2 mutant of AtNIP5;1. In contrast, both the single and double mutations at the H2 and/or H5 positions of AtNIP5;1 did not affect arsenite transport activity. The results reveal that the residue at the H5 position of the ar/R filter of both OsLsi1 and AtNIP5;1 plays a key role in the permeability to Si and B, but there is a relatively low selectivity for arsenite.

Genome-wide identification and characterization of aquaporin gene family in common bean (Phaseolus vulgaris L.)

“…Conserved residues of the NPA domain and ar/R selectivity filter (Murata et al 2000;Hub and de Groot 2008;Mitani-Ueno et al 2011) were identified by multiple sequence alignment with MUSCLE of the different sub-families of P. vulgaris aquaporins together with the tomato aquaporins identified by Reuscher et al (2013). Prediction of transmembrane domains was performed using TMHMM server v.2.0, using default parameters (http://www.cbs.dtu.dk/services/TMHMM/) (Krogh et al 2001).…”

“…This aquaporin is also able to transport arsenite and boric acid when expressed in Xenopus oocytes. Mutation in the H5 position of the ar/R selectivity filter resulted in a loss of transport activity for these molecules (Mitani-Ueno et al 2011). On the other hand, a G to A mutation in the H2 position of the ar/R selectivity filter does not affect the transport 15 capability of this protein (Mitani-Ueno et al 2011).…”

“…Mutation in the H5 position of the ar/R selectivity filter resulted in a loss of transport activity for these molecules (Mitani-Ueno et al 2011). On the other hand, a G to A mutation in the H2 position of the ar/R selectivity filter does not affect the transport 15 capability of this protein (Mitani-Ueno et al 2011). Since PvNIP2;1 showed sequence similarities with OsNIP2;1, with a G to A mutation in the H2 position of the ar/R selectivity filter, we suggest that this protein could be involved in both silicon or boric acid homeostasis in common bean.…”

“…This region is characterized by four residues, one in helix 2 (H2), one in helix 5 (H5), and two in loop E (LE1 and LE2). The ar/R selectivity filter acts as a size-exclusion barrier and regulates the transport specificity of these proteins (Hub and de Groot 2008;Mitani-Ueno et al 2011). …”

Effects of exogenous 5-aminolevulinic acid on PIP1 and NIP aquaporin gene expression in seedlings of cucumber cultivars subjected to salinity stress

“…HvPIP2 has been shown to transport water and have the ability to mediate water loss from plant cells under salt stress (Horie et al, 2011). NIPs only exist in plants and can transport different substrates, not only water, glycerol (Dean et al, 1999), and lactic acid (Choi and Roberts, 2007) but also large solutes such as urea (Wallace and Roberts, 2005), boron, silicon, and arsenic (Mitani-Ueno et al, 2011). TaNIP (Triticum asetivum L. nodulin 26-like intrinsic protein) has been reported to be associated with salt tolerance pathways in plants (Gao et al, 2010).…”