Establishment of genetically encoded biosensors for cytosolic boric acid in plant cells

Abstract

Boron (B) is an essential micronutrient for plants. To maintain B concentration in tissues at appropriate levels, plants use boric acid channels belonging to the NIP subfamily of aquaporins and BOR borate exporters. To regulate B transport, these transporters exhibit different cell-type specific expression, polar localization, and B-dependent post-transcriptional regulation. Here, we describe the development of genetically encoded biosensors for cytosolic boric acid to visualize the spatial distribution and temporal dynamics of B in plant tissues. The biosensors were designed based on the function of the NIP5;1 5'-untranslated region (UTR), which promotes mRNA degradation in response to an elevated cytosolic boric acid concentration. The signal intensities of the biosensor coupled with Venus fluorescent protein and a nuclear localization signal (uNIP5;1-Venus) showed negative correlation with intracellular B concentrations in cultured tobacco BY-2 cells. When expressed in Arabidopsis thaliana, uNIP5;1-Venus enabled the quantification of B distribution in roots at single-cell resolution. In mature roots, cytosolic B levels in stele were maintained under low B supply, while those in epidermal, cortical, and endodermal cells were influenced by external B concentrations. Another biosensor coupled with a luciferase protein fused to a destabilization PEST sequence (uNIP5;1-Luc) was used to visualize changes in cytosolic boric acid concentrations. Thus, uNIP5;1-Venus/Luc enables visualization of B transport in various plant cells/tissues.

Defects in meristem maintenance, cell division, and cytokinin signaling are early responses in the boron deficient maize mutant tassel‐less1

“…Boron functions to crosslink two pectin subunits of Rhamnogalacturonan‐II (RG‐II) in the primary cell wall (Kobayashi et al, 1996; Matoh et al, 1996; O'Neill et al, 1996), yet cell wall defects are not sufficient to explain all boron deficiency induced developmental defects (Brown et al, 2002) suggesting potential cell wall‐independent boron functions. Recent research has shed light on additional functions of boron in the plasma membrane (Matthes & Torres‐Ruiz, 2016; Voxeur & Fry, 2014; Wimmer et al, 2009), interactions of boron with auxin and cytokinins (Camacho‐Cristóbal et al, 2015; Gómez‐Soto et al, 2019; Li et al, 2015; Martín‐Rejano et al, 2011; Matthes & Torres‐Ruiz, 2016; Poza‐Viejo et al, 2018), roles of cytosolic boron (Fukuda et al, 2018), boron‐dependent transcript‐level changes (Abreu et al, 2014; Camacho‐Cristóbal et al, 2011; Feng et al, 2020; Kasajima et al, 2010; Kasajima & Fujiwara, 2007; Reguera et al, 2009; Takano et al, 2006), and boron‐dependent post‐transcriptional regulation (Sotta et al, 2021; Tanaka et al, 2016). The molecular mechanisms underlying boron deficiency‐induced phenotypic defects have been studied in the Arabidopsis root apical meristem (RAM) (Matthes et al, 2020 and references therein).…”

Assessment of a 18F-Phenylboronic Acid Radiotracer for Imaging Boron in Maize

“…B itself regulates uptake by controlling mRNA degradation and protein stability of the transporters [20]. The B responsivity of AtNIP5;1 was recently used to develop a B sensor that responds to cytosolic B levels [23]. BOR1 and NIP5 homologs have been identified in several species including maize.…”

What Can Boron Deficiency Symptoms Tell Us about Its Function and Regulation?

“…Regarding B deprivation sensing mechanisms, a computational model for B distribution in roots localized the highest concentration of soluble B around the QC, which might be likely used to keep RAM activity [ 148 ]. Also, the fact that the 5′-untranslated region (UTR) of NIP5;1 responds to the increase in cytosolic B promoting mRNA degradation [ 149 ] led to the assumption of a sensor mechanism acting in the cytosol resulting in the development of biosensors of cytosolic B [ 150 ]. Interestingly, this 5′-UTR response to B seems to function also in animal cells [ 149 ].…”