Excess Pyrophosphate within Guard Cells Delays Stomatal Closure

Plant Cell Physiol. 2019 Apr 1;60(4):875-887. doi: 10.1093/pcp/pcz002.

Abstract

A variety of cellular metabolic reactions generate inorganic pyrophosphate (PPi) as an ATP hydrolysis byproduct. The vacuolar H+-translocating pyrophosphatase (H+-PPase) loss-of-function fugu5 mutant is susceptible to drought and displays pleotropic postgerminative growth defects due to excess PPi. It was recently reported that stomatal closure after abscisic acid (ABA) treatment is delayed in vhp1-1, a fugu5 allele. In contrast, we found that specific removal of PPi rescued all of the above fugu5 developmental and growth defects. Hence, we speculated that excess PPi itself, rather than vacuolar acidification, might delay stomatal closure. To test this hypothesis, we constructed transgenic plants expressing the yeast IPP1 gene (encoding a cytosolic pyrophosphatase) driven by a guard cell-specific promoter (pGC1::IPP1) in the fugu5 background. Our measurements confirmed stomatal closure defects in fugu5, further supporting a role for H+-PPase in stomatal functioning. Importantly, while pGC1::IPP1 transgenics morphologically mimicked fugu5, stomatal closure was restored in response to ABA and darkness. Quantification of water loss revealed that fugu5 stomata were almost completely insensitive to ABA. In addition, growth of pGC1::IPP1 plants was promoted compared to fugu5 throughout their life; however, it did not reach the wild type level. fugu5 also displayed an increased stomatal index, in violation of the one-cell-spacing rule, and phenotypes recovered upon removal of PPi by pAVP1::IPP1 (FUGU5, VHP1 and AVP1 are the same gene encoding H+-PPase), but not in the pGC1::IPP1 line. Taken together, these results clearly support our hypothesis that dysfunction in stomata is triggered by excess PPi within guard cells, probably via perturbed guard cell metabolism.

Keywords: fugu5 mutants; Arabidopsis; H+-translocating pyrophosphatase (H+-PPase); Pyrophosphate; Stomatal closure.

MeSH terms

  • Abscisic Acid / pharmacology
  • Arabidopsis / drug effects
  • Arabidopsis / genetics
  • Arabidopsis / metabolism
  • Diphosphates / metabolism*
  • Inorganic Pyrophosphatase / genetics
  • Inorganic Pyrophosphatase / metabolism
  • Mutation / genetics
  • Plant Stomata / drug effects
  • Plant Stomata / metabolism*
  • Plant Stomata / physiology

Substances

  • Diphosphates
  • diphosphoric acid
  • Abscisic Acid
  • Inorganic Pyrophosphatase