Physiological and Molecular Modulations to Drought Stress in the Brassica Species.

Climate change, particularly drought stress, significantly impacts plant growth and development, necessitating the development of resilient crops. This study investigated physiological and molecular modulations to drought stress between diploid parent species and their polyploid progeny in the Brassica species. While no significant phenotypic differences were observed among the six species, drought stress reduced growth parameters by 2.4% and increased oxidative stress markers by 1.4-fold. Drought also triggered the expression of genes related to stress responses and led to the accumulation of specific metabolites. We also conducted the first study of perfluorooctane sulfonic acid (PFOS) levels in leaves as a drought indicator. Lower levels of PFOS accumulation were linked to plants taking in less water under drought conditions. Both diploid and polyploid species responded to drought stress similarly, but there was a wide range of variation in their responses. In particular, responses were less variable in polyploid species than in diploid species. This suggests that their additional genomic components acquired through polyploidy may improve their flexibility to modulate stress responses. Despite the hybrid vigor common in polyploid species, Brassica polyploids demonstrated intermediate responses to drought stress. Overall, this study lays the framework for future omics-level research, including transcriptome and proteomic studies, to deepen our understanding of drought tolerance mechanisms in Brassica species.

Proteomic and bioinformatic analyses of proteins in the outer membrane and extracellular compartments and outer membrane vesicles of Candidatus Liberibacter species.

Citrus Huanglongbing (HLB) is the most devastating citrus disease in the world. Candidatus Liberibacter asiaticus (Las) is the prevalent HLB pathogen, which is yet to be cultivated. A recent study demonstrates that Las does not contain pathogenicity factors that are directly responsible for HLB symptoms. Instead, Las triggers systemic and chronic immune responses, representing a pathogen-triggered immune disease. Importantly, overproduction of reactive oxygen species (ROS) causes systemic cell death of phloem tissues, thus causing HLB symptoms. Because Las resides in the phloem tissues, it is expected that phloem cell might recognize outer membrane proteins, outer membrane vesicle (OMV) proteins and extracellular proteins of Las to contribute to the immune responses. Because Las has not been cultivated, we used Liberibacter crescens (Lcr) as a surrogate to identify proteins in the OM fraction, OMV proteins and extracellular proteins by liquid chromatography with tandem mass spectrometry (LC-MS/MS). We observed OMVs of Lcr under scanning electron microscope, representing the first experimental evidence that Liberibacter can deliver proteins to the extracellular compartment. In addition, we also further analyzed LC-MS/MS data using bioinformatic tools. Our study provides valuable information regarding the biology of Ca. Liberibacter species and identifies many putative proteins that may interact with host proteins in the phloem tissues.

Carry-over effects of dry period heat stress on the mammary gland proteome and phosphoproteome in the subsequent lactation of dairy cows.

Exposure to heat stress during a cow's dry period disrupts mammary gland remodeling, impairing mammary function and milk production during the subsequent lactation. Yet, proteomic changes in the mammary gland underlying these effects are not yet known. We investigated alterations in the mammary proteome and phosphoproteome during lactation as a result of dry period heat stress using an isobaric tag for relative and absolute quantitation (iTRAQ)-based approach. Cows were cooled (CL; n = 12) with fans and water soakers in a free stall setting or were heat stressed through lack of access to cooling devices (HT; n = 12) during the entire dry period (approximately 46 days). All cows were cooled postpartum. Mammary biopsies were harvested from a subset of cows (n = 4 per treatment) at 14, 42, and 84 days in milk. Overall, 251 proteins and 224 phosphorylated proteins were differentially abundant in the lactating mammary gland of HT compared to CL cows. Top functions of differentially abundant proteins and phosphoproteins affected were related to immune function and inflammation, amino acid metabolism, reactive oxygen species production and metabolism, tissue remodeling, and cell stress response. Patterns of protein expression and phosphorylation are indicative of increased oxidative stress, mammary gland restructuring, and immune dysregulation due to prior exposure to dry period heat stress. This study provides insights into the molecular underpinnings of disrupted mammary function and health during lactation arising from prior exposure to dry period heat stress, which might have led to lower milk yields.

Identification of proteins associated with two diverse Caulobacter phicbkvirus particles.

Genomic evolution among bacteriophages infecting Caulobacter crescentus is inevitable. However, the conservation of the proteins associated with intact phage particles has not been investigated. In this study, we compared the structural proteins associated with two genomically diverse but morphologically similar C. crescentus-infecting bacteriophages, phiCbK and CcrSC. We were able to detect more than 20 proteins that are part of the bacteriophage particle in both phages, and we were able to identify a small number of proteins that were found in only one of the two phage particles. All but one of the genes coding for these structural proteins were located in a region of the genome that had been designated a structural region, confirming the idea that the genes in these phage genomes are clustered according to their function. During the purification process, we also discovered that phiCbk has a replication complex that can be recovered from the cell lysate, and this complex allowed us to identify many of the phage proteins involved in phage genome replication.

Diversity in Aß deposit morphology and secondary proteome insolubility across models of Alzheimer-type amyloidosis.

A hallmark pathology of Alzheimer's disease (AD) is the formation of amyloid ß (Aß) deposits that exhibit diverse localization and morphologies, ranging from diffuse to cored-neuritic deposits in brain parenchyma, with cerebral vascular deposition in leptomeningeal and parenchymal compartments. Most AD brains exhibit the full spectrum of pathologic Aß morphologies. In the course of studies to model AD amyloidosis, we have generated multiple transgenic mouse models that vary in the nature of the transgene constructs that are expressed; including the species origin of Aß peptides, the levels and length of Aß that is deposited, and whether mutant presenilin 1 (PS1) is co-expressed. These models recapitulate features of human AD amyloidosis, but interestingly some models can produce pathology in which one type of Aß morphology dominates. In prior studies of mice that primarily develop cored-neuritic deposits, we determined that Aß deposition is associated with changes in cytosolic protein solubility in which a subset of proteins become detergent-insoluble, indicative of secondary proteome instability. Here, we survey changes in cytosolic protein solubility across seven different transgenic mouse models that exhibit a range of Aß deposit morphologies. We find a surprisingly diverse range of changes in proteome solubility across these models. Mice that deposit human Aß40 and Aß42 in cored-neuritic plaques had the most robust changes in proteome solubility. Insoluble cytosolic proteins were also detected in the brains of mice that develop diffuse Aß42 deposits but to a lesser extent. Notably, mice with cored deposits containing only Aß42 had relatively few proteins that became detergent-insoluble. Our data provide new insight into the diversity of biological effects that can be attributed to different types of Aß pathology and support the view that fibrillar cored-neuritic plaque pathology is the more disruptive Aß pathology in the Alzheimer's cascade.

Seaweed natural products modify the host inflammatory response via Nrf2 signaling and alter colon microbiota composition and gene expression.

Seaweeds are an important component of human diets, especially in Asia and the Pacific islands, and have shown chemopreventive as well as anti-inflammatory properties. However, structural characterization and mechanistic insight of seaweed components responsible for their biological activities are lacking. We isolated cymopol and related natural products from the marine green alga Cymopolia barbata and demonstrated their function as activators of transcription factor Nrf2-mediated antioxidant response to increase the cellular antioxidant status. We probed the reactivity of the bioactivation product of cymopol, cymopol quinone, which was able to modify various cysteine residues of Nrf2's cytoplasmic repressor protein Keap1. The observed adducts are reflective of the polypharmacology at the level of natural product, due to multiple electrophilic centers, and at the amino acid level of the cysteine-rich target protein Keap1. The non-polar C. barbata extract and its major active component cymopol, reduced inflammatory gene transcription in vitro in macrophages and mouse embryonic fibroblasts in an Nrf2-dependent manner. Cymopol-containing extracts attenuated neutrophil migration in a zebrafish tail wound model. RNA-seq analysis of colonic tissues of mice exposed to non-polar extract or cymopol showed an antioxidant and anti-inflammatory response, with more pronounced effects exhibited by the extract. Cymopolia extract reduced DSS-induced colitis as measured by fecal lipocalin concentration. RNA-seq showed that mucosal-associated bacterial composition and transcriptional profile in large intestines were beneficially altered to varying degrees in mice treated with either the extract or cymopol. We conclude that seaweed-derived compounds, especially cymopol, alter Nrf2-mediated host and microbial gene expression, thereby providing polypharmacological effects.

The Proteome of Pancreatic Cancer-Derived Exosomes Reveals Signatures Rich in Key Signaling Pathways.

Exosomes are membrane-bound vesicles that traffic small molecular cargos. These cargos participate in cell-cell communication and contribute to the pathogenesis of many disease including cancer. How these mechanisms contribute to communication within the pancreatic adenocarcinoma (PDAC) microenvironment and how they contribute to PDAC biology are poorly understood. Performed in this study are comprehensive, quantitative comparisons of the proteomes of three PDAC cell lines to those of the exosomes they produce. Approximately 35% of whole cell proteins sort into exosomes. Analysis of composition of microbiomes (ANCOM) determined a cluster of 98 enriched pancreatic cancer exosome core proteins (ePC-ECPs). Further, these proteins are predicted by ingenuity pathway analysis (IPA) as actively involved in signaling pathways regulating cell death and survival, cellular movement, and cell-to-cell signaling and interaction in particular (top three p-value significant pathways). Significant enrichment of canonical pathways of acute phase response signaling (inflammatory response signaling pathways) and FXR and RXR activation in biosynthetic pathways are also predicted; 97 ePC-ECPs are associated with cancer and among them, 34 are specifically associated with PDAC. In conclusion, exosomes from PDAC are enriched with cancer-associated signaling proteins. Further assessment of these proteins as PDAC biomarkers or therapeutic targets is warranted.

cysTMTRAQ-An integrative method for unbiased thiol-based redox proteomics.

Protein redox regulation plays important roles in many biological processes. Protein cysteine thiols are sensitive to redox changes and may function as redox switches, which turn signaling and metabolic pathways on or off to ensure speedy responses to environmental stimuli or stresses. Here we report a novel integrative proteomics method called cysTMTRAQ that combines two types of isobaric tags, cysteine tandem mass tags and isobaric tag for relative and absolute quantification, in one experiment. The method not only enables simultaneous analysis of cysteine redox changes and total protein level changes, but also allows the determination of bona fide redox modified cysteines in proteins through the correction of protein turnover. Overlooking the factor of protein-level changes in the course of protein posttranslational modification experiments could lead to misleading results. The capability to analyze protein posttranslational modification dynamics and protein-level changes in one experiment will advance proteomic studies in many areas of biology and medicine.