Loss Of GSNOR1Function Leads To Compromised Auxin Signaling And Polar Auxin Transport

Opposite effects of nitric oxide (NO) on auxin transport and signaling have been reported recently. However, genetic evidence for the effect of S-nitrosylation on auxin physiology is still lacking. To provide genetic evidence, both DR5-GUS or DR5-GFP and HS:AXR3NT-GUS (HS, heat shock promoter), a useful reporter in investigating auxin mediated degradation of AUX/IAA by auxin receptors, were introgressed into gsnorl-3, a loss of function GSNOR1mutant defective in protein de-nitrosylation. The accumulations of these three reporters were monitored both in WT and gsnor1-3in the presence or absence of auxin and/or GSNO treatments. DR5-GUS or DR5-GFP accumulation was significantly reduced in gsnor1-3compared to WT regardless of auxin presence, consistent with the results that exogenous GSNO treatment significantly reduced the DR5-GUS accumulation in WT. To our surprise, we found that degradation of AXR3NT-GUS was delayed not only by GSNO treatment but also in gsnorl-3mutant relative to WT, providing genetic evidence that auxin signaling was impaired in gsnorl-3mutant. In addition, we showed that the the overall protein S-nitrosylation was enhanced in gsnor1-3mutant relative to Col-0and the impairment of AXR3-GUS degradation by GSNO treatment is on the steps beyond assembly of SCFTIR1complex. Furthermore, we provided evidence that polar auxin transport was compromised in gsnorl-3mutant and the levels of both endogenous PIN1and PIN2as well as GFP-PIN1, GFP-PIN2, GFP-PIN3, GFP-PIN4and GFP-PIN7driven under their native promoters in the roots were universally reduced in gsnorl-3mutant compared to WT. qRT-PCR results for various PIN genes suggested that the regulation of PIN accumulation in gsnorl-3is not at transcriptional level. Together, our results indicated that impaired auxin signaling and compromised auxin transport are responsible for the auxin related phenotypes and impaired gravitropism displayed by gsmprl-3mutant.