3.4. Heat sensing is mediated by lipid signals
Lipids are the primary structural components of membranes, but also have signaling and regulatory functions, coupling perception of environmental cues to cellular responses (Hou et al., 2015). Lipids such as PA and phosphatidylinositol 4,5-bisphosphate (PIP2), and their metabolic enzymes, i.e. phospholipases C and D (PLC/PLD), and lipid kinases, diacylglycerol (DAG) kinase (DGK) and phosphatidylinositol-4-phosphate 5-kinase (PIP5K) have a wide range of cellular regulatory functions in environmental stress responses. In Arabidopsis seedlings, heat stress (40°C) triggers increases in PA and PIP2 abundance within 2 min (Mishkind et al., 2009).
This extremely rapid response suggests that the synthesis of these signaling lipids is closely tied to thermosensing, but as of yet it is unknown how increases in temperature activate these lipid modifying enzymes (Fig. 4). High-temperature induction of PA is largely dependent on membrane lipid hydrolysis by PLD (Mishkind et al., 2009; Shiva et al., 2020) (Fig. 4). The PLD enzyme is localized at the plasma membrane and associated with microtubules, where it regulates their membrane-anchorage (Andreeva et al., 2009). In heat-stressed stomatal cells, apoplastic H2O2 enters the cytosol through aquaporins. H2O2oxidizes cysteine residues in the C2 domain of PLDδ. The modified cysteine residues promoted Ca2+ binding to PLDδ, which resulted in depolymerization of microtubules (Song et al., 2020; S.-S. Zhang et al., 2017). Blocking microtubule depolymerization by chemical stabilizers inhibited the upregulation of HSP70 and the induction of MAPK activity under heat stress (Sangwan et al., 2002; Suri & Dhindsa, 2007), which suggests that this pathway acts to promote thermal acclimation. Curiously though, mutants lacking PLDδ were more tolerant to heat stress, which suggests the opposite. The fact that PLDδ requires Ca2+ and H2O2for its activation hints that, despite its rapid activation, phospholipid signaling occurs downstream of primary thermosensing.
Glyceraldehyde-3-phosphate dehydrogenase (GAPC) may also play a role in heat stress signaling. GAPC was shown to translocate to the nucleus under heat stress, where it activates the transcription factor NF-YC10. Activated NF-YC10 then promotes the expression of genes that confer thermotolerance (S.-C. Kim et al., 2020). The mechanism the promotes GAPC nuclear translocation is as yet unresolved. When in the cytosol, GAPC can directly bind PLDδ and positively promote its activity (Guo et al., 2012). It has also been shown to bind PA (McLoughlin et al., 2013) but it is unclear how or if these attributes contribute to temperature regulation of GAPC. The involvement of GAPC in PLD/PA signaling raises the possibility that the heat stress response is coordinated with basal cell metabolism.
PLC also appears to have a function in the heat stress response, because PLC9 and PLC3 knock-out seedlings show severely impaired basal and/or acquired heat tolerance, while overexpression improved this (K. Gao et al., 2014; Zheng et al., 2012) (Fig. 4). PLC hydrolyzes PIP and PIP2 to generate DAG and inositolphosphates. The latter could eventually result in the activation of a Ca2+channel (Munnik, 2014). PLC9 and PLC3, both localized at the plasma membrane, were required for the induction of cytosolic Ca2+ and enhanced expression of sHSPs under heat stress.
The heat stress-induced accumulation of PIP2 displayed interesting dynamics. During heat exposure, PIP2accumulated first at the plasma membrane, after which it appeared in cytoplasmic punctate structures, followed by accumulation at the nuclear envelope (Mishkind et al., 2009). PIP2 can function in endocytosis and associates with membrane microdomains (Furt et al., 2010). Microdomains are considered critical in the regulation of early stress signaling, as they contain signaling proteins such as RbohD, HSPs, and CNGCs (Dietrich et al., 2020; Horvath et al., 1998; Niu & Xiang, 2018). Acting very early in the response pathway, PLC3 and PLC9 may be physically close to the thermosensor. Regulation of PLCs is complex, involving calcium, G-proteins and post-translational modifications (Munnik, 2014). Potential protein interactors of PLC3 and PLC9, including two receptor-like kinases, could provide interesting clues as to their heat-responsive mode of activation (Pokotylo et al., 2013).