The Mechanosensitive Ion Channel MSL10 Potentiates Responses to Cell Swelling in Arabidopsis Seedlings

Abstract

+1 314 935-9223 14 15 16 Key words: Programmed cell death, cell swelling, hypo-osmotic, mechanosensitive, reactive 17 oxygen species 18osmotic shock, where they are thought to up-regulate rehydration-responsive genes, including 68 several enzymes that reduce levels of the plant hormone ABA (Tsugama et al., 2012; Tsugama et 69 al., 2014; Tsugama et al., 2016). Despite these many observations, it is still not clear how these 70 downstream signaling components are connected, nor how they might lead to adaptive responses. 71 72It is also poorly understood how cell swelling is initially perceived in plants. The many potential 73 signals include molecular crowding, membrane tension, osmotic gradient, turgor, membrane 74 curvature, cell wall damage, and the disruption of the plasma membrane-cell wall connections; 75 (Haswell and Verslues, 2015; Cuevas-Velazquez and Dinneny, 2018; Hýskova and Ryslava, 2018; 76 Gigli-Bisceglia et al., 2019; Le Roux et al., 2019). Several families of receptor-like kinases, 77including the WALL-ASSOCATED KINASES, Catharanthus roseus RLK family, have been 78 implicated in sensing cell wall integrity (Nissen et al., 2016; Gigli-Bisceglia et al., 2019). In 79 particular, the CrRLK FERONIA has been implicated in the perception of mechanical signals, 80including hypo-osmotic shock ((Shih et al., 2014). Here we investigate the role of another class of 81 candidate cell swelling sensors, mechanosensitive ion channels (MS) (Cazalé et al., 1998; Haswell 82 and Verslues, 2015; Kobayashi et al., 2018). 83 84 MS ion channels are multimeric proteins embedded in the membrane that mediate ion flux across 85 the membrane in response to lateral membrane tension, and are found in all kingdoms of life 86 (Martinac, 2012;Ranade et al., 2015; Basu and Haswell, 2017). Rapid cell swelling is likely to 87 lead to an immediate increase in membrane tension, leading to the opening of MS ion channels 88 and thereby to the exit of osmolytes and/or entry of calcium signals that regulate downstream 89 adaptive events. In Escherichia coli, the MS ion channels MscS and MscL serve as osmotic safety 90 valves, opening in response to severe hypo-osmotic shock and mediating the release of osmolytes 91 (Levina et al., 1999;Sukharev, 2002; Belyy et al., 2010). The subsequent departure of water from 92 the cell presumably reduces swelling and preserves cell integrity. It is also possible that EcMscS 93 mediates the swelling-induced entry of calcium, leading to intracellular signaling events (Cox et 94 al., 2013). In plants, the overexpression of members of the MCA family leads to increased Ca 2+ 95 uptake in response to hypoosmotic shock in Arabidopsis, tobacco, and rice; but are not strictly 96 required for swelling-induced Ca 2+ uptake (Nakagawa et al., 2007; Kurusu et al., 2012b; Kurusu 97 et al., 2012a). Members of the MscS-Like (MSL) family of MS ion channels have been implicated 98 in osmotic homeostasis in chloroplasts (Haswell and Meyerowitz, 2006; Veley et al., 2012), and 99 pollen (Hamilton et al., 2015), bu...

Unbiased proteomic and forward genetic screens reveal that mechanosensitive ion channel MSL10 functions at ER-plasma membrane contact sites in Arabidopsis thaliana

“…Salinity-induced EPCS expansion is reversible when seedlings are moved to media lacking NaCl, triggering a hypo-osmotic shock (Lee et al, 2019). As MSL10 plays a role in the cellular response to hypo-osmotic cell swelling (Basu and Haswell, 2020), we asked if MSL10 was also responsible for EPCS shrinking under these conditions. We found that MAPPER-GFP signal decreased in cotyledon epidermal cells 24 hr after hypo-osmotic shock ( Figure 3-figure supplement 1b ) but that this phenomenon was unaffected by the msl10-1 or msl10-3G alleles.…”

“…The mechanosensitive ion channel MSL10 has been well studied using electrophysiological approaches (Haswell et al, 2008; Maksaev and Haswell, 2012; Maksaev et al, 2018). Genetic analyses have attributed a variety of roles to MSL10, like the induction of Ca 2+ transients, reactive oxygen species accumulation, enhanced immune responses, and programmed cell death (Basu and Haswell, 2020; Moe-Lange et al, 2021; Basu et al, 2021), but we lack a clear understanding of how MSL10 activation leads to these downstream signaling outcomes. Studies using multiple gain-of-function MSL10 alleles found that MSL10 signaling can trigger cell death independently of ion flux (Veley et al, 2014; Zou et al, 2016; Maksaev et al, 2018; Basu et al, 2020), though it remains unknown how this occurs.…”

“…However, the fact that phosphomimetic (MSL10 7D ), phosphodead (MSL10 7A ), and gain-of-function msl10-3G (MSL10 S640L) versions all interacted with VAP27-1 and VAP27-3 ( Figure 2-supplemental figure 1 ) implies that MSL10 signaling activation is independent of VAP binding. Rather, MSL10 and VAP27s are likely to interact constitutively, as they interacted both in adult leaves, a tissue type in which MSL10-GFP overexpression promotes cell death signaling (Veley et al, 2014) and in seedlings, a stage where MSL10-GFP overexpression has no effect under normal conditions (Basu and Haswell, 2020).…”

“…MSL10 is plasma membrane-localized (Haswell et al, 2008; Veley et al, 2014), and genetic studies have implicated it in a range of physiological roles. In response to hypo-osmotic cell swelling, MSL10 promotes a cytosolic Ca 2+ transient, the accumulation of reactive oxygen species, the induction of TOUCH-INDUCIBLE gene expression, and programmed cell death (Basu and Haswell, 2020). MSL10 also contributes to systemic electrical and Ca 2+ signaling in response to wounding (Moe-Lange et al, 2021).…”

Phosphate Deprivation Can Impair Mechano-Stimulated Cytosolic Free Calcium Elevation in Arabidopsis Roots

“…Plasma membrane (PM) mechano-sensitive channels or components of channel complexes may be involved in the [Ca 2+ ] cyt response, mediating Ca 2+ influx. Candidates include MCA1 (Mid1-Complementing Activity1, [ 1 ]), MCA2 [ 8 ], OSCA1 (reduced hyperosmolality-induced [CA 2+ ] cyt increase1, [ 9 , 10 ]), MSL10 (MscS-like10, [ 11 ]) and DEK1 (Defective Kernel 1, [ 12 , 13 ]). Alternatively, PM Ca 2+ channels governed by PM receptor-like kinases (that may or may not be wall-associated) would be competent to elevate [Ca 2+ ] cyt [ 3 , 14 , 15 ].…”