Indole-3-acetic acid is a physiological inhibitor of TORC1 in yeast

Abstract

Indole-3-acetic acid (IAA) is the most common, naturally occurring phytohormone that regulates cell division, differentiation, and senescence in plants. The capacity to synthesize IAA is also widespread among plant-associated bacterial and fungal species, which may use IAA as an effector molecule to define their relationships with plants or to coordinate their physiological behavior through cell-cell communication. Fungi, including many species that do not entertain a plant-associated life style, are also able to synthesize IAA, but the physiological role of IAA in these fungi has largely remained enigmatic. Interestingly, in this context, growth of the budding yeast Saccharomyces cerevisiae is sensitive to extracellular IAA. Here, we use a combination of various genetic approaches including chemical-genetic profiling, SAturated Transposon Analysis in Yeast (SATAY), and genetic epistasis analyses to identify the mode-of-action by which IAA inhibits growth in yeast. Surprisingly, these analyses pinpointed the target of rapamycin complex 1 (TORC1), a central regulator of eukaryotic cell growth, as the major growth-limiting target of IAA. Our biochemical analyses further demonstrate that IAA inhibits TORC1 both in vivo and in vitro. Intriguingly, we also show that yeast cells are able to synthesize IAA and specifically accumulate IAA upon entry into stationary phase. Our data therefore suggest that IAA contributes to proper entry of yeast cells into a quiescent state by acting as a metabolic inhibitor of TORC1.

An improved auxin-inducible degron system for fission yeast

“…In particular, the model organism Saccharomyces cerevisiae is able to synthesize and secrete auxin ( Rao et al 2010 ). Furthermore, in S. cerevisiae , IAA promotes the morphological transition to a filamentous form, retards growth, and inhibits the activity of TORC1 ( Prusty et al 2004 ; Liu et al 2016 ; Snyder et al 2019 ; Nicastro et al 2021 ), and NAA exhibits a stronger TORC1 inhibition effect than IAA ( Prusty et al 2004 ; Liu et al 2016 ; Snyder et al 2019 ; Nicastro et al 2021 ). In the other model yeast, the fission yeast Schizosaccharomyces pombe , growth inhibition was observed in the presence of high concentrations of NAA ( Kanke et al 2011 ).…”

Manganese is a Physiologically Relevant TORC1 Activator in Yeast and Mammals

“…A similar preference for Mn 2+ over Mg 2+ has also been observed in mTOR kinase autophosphorylation assays (31, 32). Based on these and our observations, we decided to assess whether Mn 2+ may act as a metal cofactor for TORC1 activity in vitro using TORC1 purified from yeast and a truncated form of Lst4 (Lst4 Loop ; (47)) as a substrate. In control experiments without divalent ions, TORC1 activity remained undetectable (Figure 3A and B).…”

“…TORC1 was purified and kinase assays were performed as previously described 47 with minor modifications. For the kinase assays in the presence of various concentrations of magnesium or manganese, reactions were performed in a total volume of 30 μl with kinase buffer (50 mM HEPES/NaOH [pH 7.5], 150 mM NaCl), 400 ng of purified His 6 -Lst4 Loop , 60 ng TORC1 (quantified with respect to the Tor1 subunit) and 640, 320, 160, 80, 40 or 20 μM MgCl2/MnCl2.…”

A toolbox for systematic discovery of stable and transient protein interactors in baker’s yeast

“…In addition, despite the clear benefits of the ABOLISH system, we generated a TurboID ‘only’ library for instances where the addition of auxin may interfere with the proteins being studied, such as for TORC1 and its associated signalling pathways (Nicastro et al , 2021). Similarly, a BioID2 library was also included as part of our toolkit since it has already been adopted by the yeast community (Opitz et al , 2017; Singh et al , 2020).…”