Expression ofTaTAR2.3-1B,TaYUC9-1andTaYUC10correlates with auxin and starch content of developing wheat grains

Abstract

The role of auxin in developing grains of wheat (Triticum aestivum) is contentious with contradictory reports indicating either positive or negative effects of IAA (indole-3-acetic acid) on grain size. In addition, the contributions to the IAA pool from de novo synthesis via tryptophan, and from hydrolysis of IAA-glucose are unclear. Here we describe the first comprehensive study of tryptophan aminotransferase and indole-3-pyruvate mono-oxygenase expression during wheat grain development from 5 to 20 days after anthesis. A comparison of expression data with measurements of endogenous IAA via combined liquid chromatography-tandem mass spectrometry with heavy isotope labelled internal standards indicates that TaTAR2.3-1B, TaYUC9-A1, TaYUC9-B, TaYUC9-D1, TaYUC10-A and TaYUC10-D are primarily responsible for IAA production in developing grains. Furthermore, we show that IAA synthesis is controlled by genes expressed specifically in developing wheat grains as has already been reported in rice (Oryza sativa) and maize (Zea mays). Our results cast doubt on the proposed role of THOUSAND-GRAIN WEIGHT gene, TaTGW6, in promoting larger grain size via negative effects on grain IAA content. The work on TaTGW6 has overlooked the contribution of the dominant IAA biosynthesis pathway. Although IAA synthesis occurs primarily in the endosperm of wheat grains, we show that the TaYUC9-1 group is also strongly expressed in the embryo. Within the endosperm, TaYUC9-1 expression is highest in aleurone and transfer cells, supporting data from other cereals suggesting that IAA has a key role in differentiation of these tissues.

Reinvestigation of grain weight genesTaTGW6andOsTGW6casts doubt on their role in auxin regulation in developing grains

“…In addition, an exhaustive search for expression of the wheat orthologues of IAGLU was unable to detect gene activity in developing grains and other tissues. In contrast, our recent work on the TAR/YUCCA pathway showed this was highly active in developing wheat grains from 10 to 20 DAA (Kabir et al 2020) similar to previously published work on rice (Abu-Zaitoon et al 2012). Expression of TaTAR2.3-1B, TaYUC9-1 and TaYUC10 was strongly correlated with the free IAA content of developing wheat grains.…”

“…IAA was eluted with 1 mL ACN and transferred to vials and stored at -20ºC. The remaining half samples were prepared for free IAA analysis as described by Kabir et al (2020). The analysis of C: 13 C IAA was conducted using a triple quadrupole Liquid Chromatograph Mass Spectrometer (LCMS)-8050, (Shimadzu) with XBridge TM C18 3.5 µm, 2.1×50 mm column (Phenomenex).…”

“…) and from 10 DAA in wheat (Kabir et al 2020). The TaTGW6 work has further problems in that the authors did not assess the activity of the gene product and the IAA measurements used inappropriate, low specificity methods (High performance liquid chromatography with UV absorbance detector).…”

Reinvestigation of THOUSAND-GRAIN WEIGHT 6 grain weight genes in wheat and rice indicates a role in pollen development rather than regulation of auxin content in grains

“…IAA was eluted with 1 mL ACN and transferred to vials and stored at -20ºC. The remaining half samples were prepared for free IAA analysis as described by Kabir et al (2020). The analysis of 12 C: C IAA was conducted using a triple quadrupole Liquid Chromatograph Mass Spectrometer (LCMS)-8050, (Shimadzu) with XBridge TM C18 3.5 µm, 2.1×50 mm column (Phenomenex).…”

“…Neither Ishimaru et al (2013) nor Hu et al (2016a) investigated the availability of IAA-Glc as a substrate for TGW6. Furthermore, the TAR/YUCCA pathway of IAA production is strongly upregulated in rice from 4 DAA (Abu-Zaitoon et al 2012) and from 10 DAA in wheat (Kabir et al 2020). The TaTGW6 work has further problems in that the authors did not assess the activity of the gene product and the IAA measurements used inappropriate, low specificity methods (High performance liquid chromatography with UV absorbance detector).…”