Functional analysis of OsPUT1, a rice polyamine uptake transporter

Abstract

Polyamines are nitrogenous compounds found in all eukaryotic and prokaryotic cells and absolutely essential for cell viability. In plants, they regulate several growth and developmental processes and the levels of polyamines are also correlated with the plant responses to various biotic and abiotic stresses. In plant cells, polyamines are synthesized in plastids and cytosol. This biosynthetic compartmentation indicates that the specific transporters are essential to transport polyamines between the cellular compartments. In the present study, a phylogenetic analysis was used to identify candidate polyamine transporters in rice. A full-length cDNA rice clone AK068055 was heterologously expressed in the Saccharomyces cerevisiae spermidine uptake mutant, agp2∆. Radiological uptake and competitive inhibition studies with putrescine indicated that rice gene encodes a protein that functioned as a spermidine-preferential transporter. In competition experiments with several amino acids at 25-fold higher levels than spermidine, only methionine, asparagine, and glutamine were effective in reducing uptake of spermidine to 60% of control rates. Based on those observations, this rice gene was named polyamine uptake transporter 1 (OsPUT1). Tissue-specific expression of OsPUT1 by semiquantitative RT-PCR showed that the gene was expressed in all tissues except seeds and roots. Transient expression assays in onion epidermal cells and rice protoplasts failed to localize to a cellular compartment. The characterization of the first plant polyamine transporter sets the stage for a systems approach that can be used to build a model to fully define how the biosynthesis, degradation, and transport of polyamines in plants mediate developmental and biotic responses.

Natural variation in a polyamine transporter determines paraquat tolerance in Arabidopsis

“…Collectively, these results provide evidence that the LAT family transporter RMV1 is engaged in the proton-dependent transport of PA and PQ (MV) in Arabidopsis, consistent with the recent report of a rice gene mediating spermidine uptake (25). Moreover, we demonstrated that natural variation in RMV1 defines PQ uptake in A. thaliana.…”

Phloem-Specific Methionine Recycling Fuels Polyamine Biosynthesis in a Sulfur-Dependent Manner and Promotes Flower and Seed Development

“…Root application of PA has been previously shown to complement PA-associated growth defects (Waduwara-Jayabahu et al, 2012), and recently PA transporters have been identified from rice and Arabidopsis (Fujita et al, 2012;Mulangi et al, 2012aMulangi et al, , 2012b, suggesting uptake of polyamines by the root system. We added varying amounts of the non-Scontaining PA putrescine, spermidine, or spermine directly to the 10 mM sulfate-containing medium and observed that neither putrescine nor spermidine improved the growth of the mti1 and dep1 mutant plants (Supplemental Figs.…”

Polyamine Transport Systems in Plants

“…The study identifi ed a rice gene designated as PA uptake transporter 1 ( OsPUT1 ), which partially complemented the PA-insensitive agp2 mutant phenotype. Radiological uptake assays and competitive inhibition analyses in the yeast transformant revealed that OsPUT1 preferentially transported Spd with K m = 15.2 μM (Mulangi et al 2012b ). A similar experimental approach identifi ed fi ve additional PA transporters in rice and Arabidopsis , including OsPUT2 , OsPUT3 , AtPUT1 , AtPUT2 , and AtPUT3 (Mulangi et al 2012a ).…”

“…Mulangi et al ( 2012b ) searched for rice and Arabidopsis genes that show sequence similarity to PA transporters of Leishmania major ( LmPOT1 ) (Hasne and Ullman 2005 ) and Trypanosoma cruzi ( TcPAT12 ) (Carrillo et al 2006 ). Full-length cDNAs of candidate genes were transformed into the Saccharomyces cerevisiae agp2∆ mutant, which is defi cient in Spd transport (Aouida et al 2005 ).…”