TRH1/KUP4 is a coupler of auxin-mediated root system architecture responses in Arabidopsis

TRH1, also classified as KUP4, belongs in the family of plant HAK/KUP/KT (High-Affinity K+/ K+ UPtake/ K+ Transporter) transporters, homologous to the bacterial KUP and fungal HAK transporters, which most likely as K+/H+ exchangers function as electrochemical potential-driven transporters. Arabidopsis contains 13 genes, classified into five major clades (Clades I-V). Members of Clade II, including TRH1, are associated with developmental processes. Targeted expression experiments demonstrated that K+ transporter TRH1 is a coupler of two auxin-mediated developmental processes that determine Root System Architecture, root hair morphogenesis and root gravitropism. Targeting TRH1 expression in central cylinder cells that coincide with the acropetal route of auxin transport, rescued trh1 root agravitropism, whereas positional TRH1 expression in peripheral cell layers that define the basipetal route of auxin transport, restored trh1 root hair defects. To dissect the two auxin-mediated developmental responses, a transcriptome analysis was performed identifying the differentially expressed genes involved in either root bending or root hair development. For the transcriptome studies, ~1 mm sections of root tips from trh1, pPIN1::TRH1-YFP and pPIN2::TRH1-YFP transgenic lines were collected for RNA-Seq analysis. Total RNA was extracted using the Direct-zol RNA Miniprep (Zymo research, Irvine CA, USA) kit with an on-column DNase treatment according to the manufacturer’s instructions. The quantity and integrity of RNA were assessed using a NanoDrop ND-1000 spectrophotometer (Thermo Fischer Scientific, Waltham, MA, USA) and agarose gel electrophoresis. RNA-seq libraries were generated using the TruSeq Low Input kit according to the manufacturer’s instructions (Illumina). Sequencing was performed on BGISEQ-500 platform instrument at BGI (Beijing Genomics Institute) in three biological replicates for each sample.