Steffen P. Graether

Plants undergo a wide range of deleterious effects as a result of exposure to abiotic stress, such as cold, salinity, and drought conditions. Some of these effects include protein denaturation and aggregation, membrane leakage and phase transitions, and DNA damage. One protective measure includes the expression of dehydrins, which are members of the group 2 LEA protein family. Several studies have shown that the S-segment, a serine-rich tract found in many dehydrins, can be phosphorylated in vivo . The main functional role of this phosphorylation appears to recruit the dehydrin to the nucleus, though this is not an absolute requirement. Here, we investigate whether pseudophosphorylation (i.e. the substitution of serine residues with glutamate) had any effect on the protection of proteins, membranes and DNA using VrDHN1 as a model YSK ₂ dehydrin. The changes resulted in improved protection of lactate dehydrogenase from freeze-thaw damage and caused an increase in α-helicity after binding to SDS micelles. Pseudophosphorylation was found to reduce DNA binding but had no effect on the protection of DNA from ROS damage. In addition to a possible role in nuclear localization, phosphorylation may result in a tuning of dehydrin function toward improved protective ability.

Antifreeze proteins (AFPs) are natural biomolecules found in cold-adapted organisms that lower the freezing point of water, allowing survival in icy conditions. These proteins have the potential to improve cryopreservation techniques by enhancing the quality of genetic material postthaw. Deschampsia antarctica, a freezing-tolerant plant, possesses AFPs and is a promising candidate for cryopreservation applications. In this study, we investigated the cryoprotective properties of AFPs from D. antarctica extracts on Atlantic salmon spermatozoa. Apoplastic extracts were used to determine ice recrystallization inhibition (IRI), thermal hysteresis (TH) activities and ice crystal morphology. Spermatozoa were cryopreserved using a standard cryoprotectant medium (C+) and three alternative media supplemented with apoplastic extracts. Flow cytometry was employed to measure plasma membrane integrity (PMI) and mitochondrial membrane potential (MMP) postthaw. Results showed that a low concentration of AFPs (0.05 mg/mL) provided significant IRI activity. Apoplastic extracts from D. antarctica demonstrated a cryoprotective effect on salmon spermatozoa, with PMI comparable to the standard medium. Moreover, samples treated with apoplastic extracts exhibited a higher percentage of cells with high MMP. These findings represent the first and preliminary report that suggests that AFPs derived from apoplastic extracts of D. antarctica have the potential to serve as cryoprotectants and could allow the development of novel freezing media.

Intrinsically disordered proteins (IDPs) are proteins that lack a stable 3D structure but maintain a biological function. It has been frequently suggested that IDPs are difficult to align because they tend to have fewer conserved residues compared to ordered proteins, but to our knowledge this has never been directly tested. To compare the alignments of ordered proteins to IDPs, their multiple sequence alignments (MSAs) were assessed using two different methods. The first compared the similarity between MSAs produced using the same sequences but created with Clustal Omega, MAFFT, and MUSCLE. The second assessed MSAs based on how well they recapitulated the species tree. These two methods measure the “correctness” of an MSA with two different approaches; the first method measures consistency while the second measures the underlying phylogenetic signal. Proteins that contained both regions of disorder and order were analyzed along with proteins that were fully disordered and fully ordered, using nucleotide, codon and peptide sequence alignments. We observed that IDPs had less similar MSAs than ordered proteins, which is most likely linked to the lower sequence conservation in IDPs. However, comparisons of tree distances found that trees from the ordered sequence MSAs were not significantly closer to the species tree than those inferred from disordered sequence MSAs. Our results show that it is correct to say that IDPs are difficult to align on the basis of MSA consistency, but that this does not equate with alignments being of poor quality when assessed by their ability to correctly infer a species tree.

Student Evaluations of Teaching (SETs) are a ubiquitous tool in higher education. Though they are not effective means of evaluating teaching ability, they are useful in formative teaching development of a teaching career. We characterise the current formative use of and attitudes towards SETs by instructors across all disciplines (STEM and non-STEM). We found that tenured instructors used SETs for formative development more than untenured, and that non-STEM instructors had more negative associations with SETs than STEM instructors. Based upon these data, we make recommendations to redesign the SET instrument and change the way in which the data are used to support formative teaching development.

Dehydrins are intrinsically disordered proteins expressed ubiquitously throughout the plant kingdom in response to desiccation. Dehydrins have been found to have a cryoprotective effect on lactate dehydrogenase (LDH) in vitro, which is in large part influenced by their hydrodynamic radius rather than the order of the amino acids within the sequence (alternatively, this may be a sequence specific effect). However, it seems that a different mechanism may underpin the cryoprotection that they confer to the cold-labile yeast frataxin homolog-1 (Yfh1). Circular dichroism spectroscopy (CD) was used to assess the degree of helicity of Yfh1 at 1 °C, both alone and in the presence of several dehydrin constructs. Three constructs were compared to the wild type: YSK₂-K→R (lysine residues substituted with arginine), YSK₂-Neutral (locally neutralized charge), and YSK₂-SpaceK (evenly distributed positive charge). The results show that sequence rearrangements and minor substitutions have little impact on the ability of the dehydrin to preserve LDH activity. However, when the positive charge of the dehydrin is locally neutralized or evenly distributed, the dehydrin becomes less efficient at promoting structure in Yfh1 at low temperatures. This suggests that a stabilizing, charge-based interaction occurs between dehydrins and Yfh1. Dehydrins are intrinsically disordered proteins, expressed by certain organisms to improve desiccation tolerance. These proteins are thought to serve many cellular roles, such as the stabilization of membranes, DNA, and proteins. However, the molecular mechanisms underlying the function of dehydrins are not well understood. Here, we examine the importance of positive charges in dehydrin sequences by making substitutions and comparing their effects in the cryoprotection of two different proteins.

Plants have evolved a number of different ways to deal with different types of abiotic stresses; at the molecular level, dehydration can cause multiple forms of damage to different biomolecules

Editorial: Structural bioinformatics and biophysical approaches for understanding the plant responses to biotic and abiotic stresses

To deal with increasingly severe periods of dehydration related to global climate change, it becomes increasingly important to understand the complex strategies many organisms have developed to cope with dehydration and desiccation. While it is undisputed that late embryogenesis abundant (LEA) proteins play a key role in the tolerance of plants and many anhydrobiotic organisms to water limitation, the molecular mechanisms are not well understood. In this review, we summarize current knowledge of the physiological roles of LEA proteins and discuss their potential molecular functions. As these are ultimately linked to conformational changes in the presence of binding partners, post-translational modifications, or water deprivation, we provide a detailed summary of current knowledge on the structure–function relationship of LEA proteins, including their disordered state in solution, coil to helix transitions, self-assembly, and their recently discovered ability to undergo liquid–liquid phase separation. We point out the promising potential of LEA proteins in biotechnological and agronomic applications, and summarize recent advances. We identify the most relevant open questions and discuss major challenges in establishing a solid understanding of how these intriguing molecules accomplish their tasks as cellular sentinels at the limits of surviving water scarcity.

Plants can be exposed to cold temperatures and have therefore evolved several mechanisms to prevent damage caused by freezing. One of the most important targets are membranes, which are particularly susceptible to cold damage. To protect against such abiotic stresses, plants express a family of proteins known as late embryogenesis abundant (LEA) proteins. Many LEA proteins are intrinsically disordered, that is, they do not contain stable secondary or tertiary structures alone in solution. These proteins have been shown in a number of studies to protect plants from damage caused by cold, drought, salinity, and osmotic stress. In this family, the most studied proteins are the type II LEA proteins, better known as dehydrins (dehydration-induced proteins). Many physiological studies have shown that dehydrins are often located near the membrane during abiotic stress and that the expression of dehydrins helps to prevent the formation of oxidation-modified lipids and reduce the amount of electrolyte leakage, two hallmarks of damaged membranes. One of the earliest biophysical clues that dehydrins are involved in membrane cryoprotection came from in vitro studies that demonstrated a binding interaction between the protein and membranes. Subsequent work has shown that one conserved motif, known as K-segments, is involved in binding, while recent studies have used NMR to explore the residue specific structure of dehydrins when bound to membranes. The biophysical techniques also provide insight into the mechanism by which dehydrins protect the membrane from cold stress, which appears to mainly involve the lowering of the transition temperature.

Bimodal grade distributions indicate a gap in learning, where the highest quartile of students is skilful in the subject matter, but the lowest quartile is not retaining course material that demonstrates a good level of understanding. Students in the lower quartile do not necessarily have the same challenges as remedial students (i.e., those that do not meet the minimum course requirement) and should therefore be directed differently. We suggest that it is important to consider which elements of the course can be modified to reduce or eliminate bimodality. We provide here an approach to detect bimodality, explore the causes, and provide potential solutions that could be applied to any course. Our case study is on a third-year biochemistry course where several semesters showed a bimodal grade distribution. While student composition and timing of the course may have contributed to this result, underlying causes that can be controlled by the instructor include the lack of student engagement and academic motivation. Increasing the opportunities to earn marks and receive feedback, adding online components using social media, implementing seminars, and training teaching assistants to lead seminars can help reduce this problem.