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1. LEA motifs promote desiccation tolerance in vivo

   https://doi.org/10.1186/s12915-021-01176-0  

   Hibshman, Jonathan D. ; Goldstein, Bob ( December 2021 , BMC Biology)

   Cells and organisms typically cannot survive in the absence of water. However, some animals including nematodes, tardigrades, rotifers, and some arthropods are able to survive near-complete desiccation. One class of proteins known to play a role in desiccation tolerance is the late embryogenesis abundant (LEA) proteins. These largely disordered proteins protect plants and animals from desiccation. A multitude of studies have characterized stress-protective capabilities of LEA proteins in vitro and in heterologous systems. However, the extent to which LEA proteins exhibit such functions in vivo, in their native contexts in animals, is unclear. Furthermore, little is known about the distribution of LEA proteins in multicellular organisms or tissue-specific requirements in conferring stress protection. Here, we used the nematodeC. elegansas a model to study the endogenous function of an LEA protein in an animal.

   We created a null mutant ofC. elegansLEA-1, as well as endogenous fluorescent reporters of the protein. LEA-1 mutant animals formed defective dauer larvae at high temperature. We confirmed thatC. eleganslacking LEA-1 are sensitive to desiccation. LEA-1 mutants were also sensitive to heat and osmotic stress and were prone to protein aggregation. During desiccation, LEA-1 expression increased and became more widespread throughout the body. LEA-1 was required at high levels in body wall muscle for animals to survive desiccation and osmotic stress, but expression in body wall muscle alone was not sufficient for stress resistance, indicating a likely requirement in multiple tissues. We identified minimal motifs withinC. elegansLEA-1 that were sufficient to increase desiccation survival ofE. coli. To test whether such motifs are central to LEA-1’s in vivo functions, we then replaced the sequence oflea-1with these minimal motifs and found thatC. elegansdauer larvae formed normally and survived osmotic stress and mild desiccation at the same levels as worms with the full-length protein.

   Our results provide insights into the endogenous functions and expression dynamics of an LEA protein in a multicellular animal. The results show that LEA-1 buffers animals from a broad range of stresses. Our identification of LEA motifs that can function in both bacteria and in a multicellular organism in vivo suggests the possibility of engineering LEA-1-derived peptides for optimized desiccation protection.

    

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2. Nonredundant functions of Mycobacterium tuberculosis chaperones promote survival under stress

   https://doi.org/10.1111/mmi.14615  

   Harnagel, Alexa ; Lopez Quezada, Landys ; Park, Sae Woong ; Baranowski, Catherine ; Kieser, Karen ; Jiang, Xiuju ; Roberts, Julia ; Vaubourgeix, Julien ; Yang, Amy ; Nelson, Brock ; et al ( November 2020 , Molecular Microbiology)

   Bacterial chaperones ClpB and DnaK, homologs of the respective eukaryotic heat shock proteins Hsp104 and Hsp70, are essential in the reactivation of toxic protein aggregates that occur during translation or periods of stress. In the pathogenMycobacterium tuberculosis(Mtb), the protective effect of chaperones extends to survival in the presence of host stresses, such as protein‐damaging oxidants. However, we lack a full understanding of the interplay of Hsps and other stress response genes in mycobacteria. Here, we employ genome‐wide transposon mutagenesis to identify the genes that supportclpBfunction in Mtb. In addition to validating the role of ClpB in Mtb's response to oxidants, we show that HtpG, a homolog of Hsp90, plays a distinct role from ClpB in the proteotoxic stress response. While loss of neitherclpBnorhtpGis lethal to the cell, loss of both through genetic depletion or small molecule inhibition impairs recovery after exposure to host‐like stresses, especially reactive nitrogen species. Moreover, defects in cells lackingclpBcan be complemented by overexpression of other chaperones, demonstrating that Mtb's stress response network depends upon finely tuned chaperone expression levels. These results suggest that inhibition of multiple chaperones could work in concert with host immunity to disable Mtb.

    

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3. Chaperone Activity and Protective Effect against Aβ-Induced Cytotoxicity of Artocarpus camansi Blanco and Amaranthus dubius Mart. ex Thell Seed Protein Extracts

   https://doi.org/10.3390/ph16060820  

   Sanchez-Rodriguez, David ; Gonzalez-Figueroa, Idsa ; Alvarez-Berríos, Merlis P. ( June 2023 , Pharmaceuticals)

   Alzheimer’s disease (AD) is the most common type of dementia and is listed as the sixth-leading cause of death in the United States. Recent findings have linked AD to the aggregation of amyloid beta peptides (Aβ), a proteolytic fragment of 39–43 amino acid residues derived from the amyloid precursor protein. AD has no cure; thus, new therapies to stop the progression of this deadly disease are constantly being searched for. In recent years, chaperone-based medications from medicinal plants have gained significant interest as an anti-AD therapy. Chaperones are responsible for maintaining the three-dimensional shape of proteins and play an important role against neurotoxicity induced by the aggregation of misfolded proteins. Therefore, we hypothesized that proteins extracted from the seeds of Artocarpus camansi Blanco (A. camansi) and Amaranthus dubius Mart. ex Thell (A. dubius) could possess chaperone activity and consequently may exhibit a protective effect against Aβ1–40-induced cytotoxicity. To test this hypothesis, the chaperone activity of these protein extracts was measured using the enzymatic reaction of citrate synthase (CS) under stress conditions. Then, their ability to inhibit the aggregation of Aβ1–40 using a thioflavin T (ThT) fluorescence assay and DLS measurements was determined. Finally, the neuroprotective effect against Aβ1–40 in SH-SY5Y neuroblastoma cells was evaluated. Our results demonstrated that A. camansi and A. dubius protein extracts exhibited chaperone activity and inhibited Aβ1–40 fibril formation, with A. dubius showing the highest chaperone activity and inhibition at the concentration assessed. Additionally, both protein extracts showed neuroprotective effects against Aβ1–40-induced toxicity. Overall, our data demonstrated that the plant-based proteins studied in this research work can effectively overcome one of the most important characteristics of AD. 

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4. Holistic insights from pollen omics : co-opting stress-responsive genes and ER-mediated proteostasis for male fertility

   https://doi.org/10.1093/plphys/kiab463  

   Sze, Heven ; Palanivelu, Ravishankar ; Harper, Jeffrey F ; Johnson, Mark A ( October 2021 , Plant Physiology)

   Abstract Sexual reproduction in flowering plants takes place without an aqueous environment. Sperm are carried by pollen through air to reach the female gametophyte, though the molecular basis underlying the protective strategy of the male gametophyte is poorly understood. Here we compared the published transcriptomes of Arabidopsis thaliana pollen, and of heat-responsive genes, and uncovered insights into how mature pollen (MP) tolerates desiccation, while developing and germinating pollen are vulnerable to heat stress. Germinating pollen expresses molecular chaperones or “heat shock proteins” in the absence of heat stress. Furthermore, pollen tubes that grew through pistils at basal temperature showed induction of the endoplasmic reticulum (ER) stress response, which is a characteristic of stressed vegetative tissues. Recent studies show MP contains mRNA–protein (mRNP) aggregates that resemble “stress” granules triggered by heat or other stresses to protect cells. Based on these observations, we postulate that mRNP particles are formed in maturing pollen in response to developmentally programmed dehydration. Dry pollen can withstand harsh conditions as it is dispersed in air. We propose that, when pollen lands on a compatible pistil and hydrates, mRNAs stored in particles are released, aided by molecular chaperones, to become translationally active. Pollen responds to osmotic, mechanical, oxidative, and peptide cues that promote ER-mediated proteostasis and membrane trafficking for tube growth and sperm discharge. Unlike vegetative tissues, pollen depends on stress-protection strategies for its normal development and function. Thus, heat stress during reproduction likely triggers changes that interfere with the normal pollen responses, thereby compromising male fertility. This holistic perspective provides a framework to understand the basis of heat-tolerant strains in the reproduction of crops. 

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5. Liquid-liquid phase separation promotes animal desiccation tolerance

   https://doi.org/10.1073/pnas.2014463117  

   Belott, Clinton ; Janis, Brett ; Menze, Michael A. ( October 2020 , Proceedings of the National Academy of Sciences)

   Proteinaceous liquid-liquid phase separation (LLPS) occurs when a polypeptide coalesces into a dense phase to form a liquid droplet (i.e., condensate) in aqueous solution. In vivo, functional protein-based condensates are often referred to as membraneless organelles (MLOs), which have roles in cellular processes ranging from stress responses to regulation of gene expression. Late embryogenesis abundant (LEA) proteins containing seed maturation protein domains (SMP; PF04927) have been linked to storage tolerance of orthodox seeds. The mechanism by which anhydrobiotic longevity is improved is unknown. Interestingly, the brine shrimpArtemia franciscanais the only animal known to express such a protein (AfrLEA6) in its anhydrobiotic embryos. Ectopic expression ofAfrLEA6 (AWM11684) in insect cells improves their desiccation tolerance and a fraction of the protein is sequestered into MLOs, while aqueousAfrLEA6 raises the viscosity of the cytoplasm. LLPS ofAfrLEA6 is driven by the SMP domain, while the size of formed MLOs is regulated by a domain predicted to engage in protein binding.AfrLEA6 condensates formed in vitro selectively incorporate target proteins based on their surface charge, while cytoplasmic MLOs formed inAfrLEA6-transfected insect cells behave like stress granules. We suggest thatAfrLEA6 promotes desiccation tolerance by engaging in two distinct molecular mechanisms: by raising cytoplasmic viscosity at even modest levels of water loss to promote cell integrity during drying and by forming condensates that may act as protective compartments for desiccation-sensitive proteins. Identifying and understanding the molecular mechanisms that govern anhydrobiosis will lead to significant advancements in preserving biological samples.

    

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