Cold acclimation and prospects for cold-resilient crops

Walker

2022

Preprint

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Antifreeze proteins (AFPs) from the model crop, Brachypodium distachyon, allow freeze survival and attenuate pathogen-mediated ice nucleation. Intriguingly, each Brachypodium AFP gene encodes two proteins, an autonomous AFP and a leucine-rich repeat (LRR). We present structural models suggesting that ice-binding motifs on the ~13 kDa AFPs can "spoil" nucleating arrays on the ~120 kDa bacterial ice nucleating proteins that form ice at high sub-zero temperatures, consistent with decreases in ice nucleating activity by lysates from wildtype compared to transgenic Brachypodium lines. Strikingly, the expression of Brachypodium LRRs in transgenic Arabidopsis inhibited an immune response to pathogen flagella peptides (flg22), with structural modelling suggesting this was due to affinity of the LRR domains to flg22. Thus, these genes play distinctive roles in connecting freeze survival and anti-pathogenic systems via their encoded proteins' ability to adsorb to ice as well as attenuating bacterial ice nucleation and the host immune response.

Section: Discussionmentioning

confidence: 99%

Brachypodium antifreeze protein gene products inhibit ice recrystallization, attenuate ice nucleation, and reduce immune response

Walker

2022

Preprint

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“…M2, for protection against P. syringae and other phytopathogens, while at the same time benefiting from other plant growth-promoting characteristics as well as enhancing cold resilience. With the presentation of this first CA Brachypodium microbiome, it is hoped that the insights gained will inspire treatment options to enhance cold tolerance and other intersecting stresses tailored toward specific agriculturally important grain crops [1,9,85,86].…”

Section: Prospects and Conclusionmentioning

confidence: 99%

“…As sessile organisms, plants are at the mercy of an array of abiotic stresses, and, as winter approaches in mid-to high-latitudes and altitudes, one such stress is low temperature. Plants employ various strategies that allow them to recognise and cope with the cold [1]. As autumn progresses, perennials undergo a period of cold acclimation, which in a few days of low temperature exposure allows them to physiologically prepare for freezing conditions.…”

Section: Introductionmentioning

confidence: 99%

“…In turn, these proteins are involved in complex crosstalk networks that prime the Brachypodium defensive response to a variety of abiotic and pathogenic stresses. Studies of cold acclimation have, for the most part, ignored the host-associated microbiota [1,7,8]. Nevertheless, the plant microbiome is emerging as an important factor in stress responses, including symbiont-mediated tolerance [9,10].…”

Section: Introductionmentioning

confidence: 99%

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Cold Acclimation in Brachypodium Is Accompanied by Changes in Above-Ground Bacterial and Fungal Communities

Walker

2021

Plants

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Shifts in microbiota undoubtedly support host plants faced with abiotic stress, including low temperatures. Cold-resistant perennials prepare for freeze stress during a period of cold acclimation that can be mimicked by transfer from growing conditions to a reduced photoperiod and a temperature of 4 °C for 2–6 days. After cold acclimation, the model cereal, Brachypodium distachyon, was characterized using metagenomics supplemented with amplicon sequencing (16S ribosomal RNA gene fragments and an internal transcribed spacer region). The bacterial and fungal rhizosphere remained largely unchanged from that of non-acclimated plants. However, leaf samples representing bacterial and fungal communities of the endo- and phyllospheres significantly changed. For example, a plant-beneficial bacterium, Streptomyces sp. M2, increased more than 200-fold in relative abundance in cold-acclimated leaves, and this increase correlated with a striking decrease in the abundance of Pseudomonas syringae (from 8% to zero). This change is of consequence to the host, since P. syringae is a ubiquitous ice-nucleating phytopathogen responsible for devastating frost events in crops. We posit that a responsive above-ground bacterial and fungal community interacts with Brachypodium’s low temperature and anti-pathogen signalling networks to help ensure survival in subsequent freeze events, underscoring the importance of inter-kingdom partnerships in the response to cold stress.

“…In the field, the formation of ice at high sub-zero temperatures is initiated by ice nucleation active (INA+) bacteria and is a major driver of crop destruction (Snyder and Melo-Abreu, 2005). Since they cannot escape low temperatures, many temperate climate plants have adopted a freeze tolerant strategy with some producing antifreeze proteins (AFPs) to help prevent freeze damage (Bredow and Walker, 2017; Juurakko et al, 2021a). In contrast, other organisms such as polar fish and temperate arthropods, which can escape low temperatures and find hibernacula, frequently adopt a freeze-avoidance strategy that can also employ AFPs, in this case to lower the freezing point relative to the melting point, also known as the thermal hysteresis (TH) gap (Duman, 2001; Bar Dolev et al, 2016; Kim et al, 2017).…”

Section: Introductionmentioning

confidence: 99%

Cold-inducible promoter-driven knockdown of Brachypodium antifreeze proteins confers freeze sensitivity

Bredow

et al. 2022

Preprint

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The model forage crop, Brachypodium distachyon, has a family of ice recrystallization inhibition (BdIRI) genes, which encode antifreeze proteins that function by adsorbing to ice crystals and inhibiting their growth. The genes were previously targeted for knockdown using a constitutive CaMV 35S promoter and the resulting transgenic Brachypodium showed reduced antifreeze activity and a greater susceptibility to freezing. However, the transgenic plants also showed developmental defects with shortened stem lengths and were almost completely sterile, raising the possibility that their reduced freeze tolerance could be attributed to developmental deficits. A cold-induced promoter from rice (prOsMYB1R35) has now been substituted for the constitutive promoter to generate temporal miRNA-mediated Brachypodium antifreeze protein knockdowns. Although transgenic lines showed no apparent pleiotropic developmental defects, they demonstrated reduced antifreeze activity as assessed by assays for ice-recrystallization inhibition, thermal hysteresis, electrolyte leakage, leaf infrared thermography, and leaf damage after infection with an ice nucleating phytopathogen. Strikingly, the number of cold-acclimated transgenic plants that survived freezing at -8 °C was reduced by half or killed entirely, depending on the line, compared to cold-acclimated wild type plants. Although these proteins have been studied for almost 60 years, this is the first unequivocal demonstration in any organism of the utility of antifreeze protein function and their contribution to freeze protection, independent of obvious developmental defects. These proteins are thus of potential interest in a wide range of biotechnological applications from accessible cryopreservation, to frozen product additives, to the engineering of transgenic crops with enhanced freezing tolerance.