Interactions of Thellungiella salsuginea dehydrins TsDHN-1 and TsDHN-2 with membranes at cold and ambient temperatures—Surface morphology and single-molecule force measurements show phase separation, and reveal tertiary and quaternary associations

Abstract

Dehydrins (group 2 late embryogenesis abundant proteins) are intrinsically-disordered proteins that are expressed in plants experiencing extreme environmental conditions such as drought or low temperature. Their roles include stabilizing cellular proteins and membranes, and sequestering metal ions. Here, we investigate the membrane interactions of the acidic dehydrin TsDHN-1 and the basic dehydrin TsDHN-2 derived from the crucifer Thellungiella salsuginea that thrives in the Canadian sub-Arctic. We show using compression studies with a Langmuir-Blodgett trough that both dehydrins can stabilize lipid monolayers with a lipid composition mimicking the composition of the plant outer mitochondrial membrane, which had previously been shown to induce ordered secondary structures (disorder-to-order transitions) in the proteins. Ellipsometry of the monolayers during compression showed an increase in monolayer thickness upon introducing TsDHN-1 (acidic) at 4°C and TsDHN-2 (basic) at room temperature. Atomic force microscopy of supported lipid bilayers showed temperature-dependent phase transitions and domain formation induced by the proteins. These results support the conjecture that acidic dehydrins interact with and potentially stabilize plant outer mitochondrial membranes in conditions of cold stress. Single-molecule force spectroscopy of both proteins pulled from supported lipid bilayers indicated the induced formation of tertiary conformations in both proteins, and potentially a dimeric association for TsDHN-2.

Characterization of dehydrin AhDHN from Mediterranean saltbush (Atriplex halimus)

“…A transmembrane pore-lining stretch in AhDHN was assigned for the residues I127-Y142 (Figures 5 and 6). In fact, DHNs were recorded to be localized to plasma membranes during cold acclimation in wheat (Danyluk et al, 1998) and in Thellungiella salsuginea (Rahman et al, 2013). Borovskii et al (2002) found that DHNs can accumulate in mitochondria in response to cold, freezing, and drought in cereals.…”

Disorder and function: a review of the dehydrin protein family

“…However, examination of the CD spectra of both proteins shows that there is no gain of α-helicity, suggesting a different mode of binding compared to ZmDHN1 (Koag et al, 2003). Using Fourier-transformed infrared spectroscopy (FTIR), Rahman et al (2013) showed that for Thellungiella salsuginea dehydrin 1 (TsDHN-1) the type and amount of structural change was dependent on lipid type, where they detected the gain of some β-strand structure for this protein when bound to lipid compositions mimicking the plasma membrane, mitochondrial membrane and chloroplast membrane (Rahman et al, 2010, 2013). It is challenging to reconcile the gain of β-strand structure compared to the α-helical structure seen in other studies for dehydrins bound to membranes, since the plasma membrane mimicking vesicles contain phospholipids.…”

The Ubiquitous Distribution of Late Embryogenesis Abundant Proteins across Cell Compartments in Arabidopsis Offers Tailored Protection against Abiotic Stress

“…Five proteins (LEA4, 5, 8, 10, and 33) belonging to the dehydrin family could interact with the plasma membrane since LEA5, LEA10, and LEA33 have been shown to bind anionic phospholipid vesicles (Kovacs et al, 2008;Eriksson et al, 2011). Moreover, dehydrins from maize (Zea mays) and the salt-tolerant Thellungiella salsuginea have also been shown to bind phospholipid vesicles (Koag et al, 2003(Koag et al, , 2009Rahman et al, 2010Rahman et al, , 2013. Interestingly, four dehydrins (LEA4, 5, 8, and 10) with a clear cytosolic localization in our experiments were also identified in chloroplast proteome (Kleffmann et al, 2004;Zybailov et al, 2008;Ferro et al, 2010;Kong et al, 2011) or in the tonoplast in the case of LEA10 (Whiteman et al, 2008).…”