Osmoregulation is important for plant growth, development and response to environmental changes. SNF1-related protein kinase 2s (SnRK2s) are quickly activated by osmotic stress and are central ...components in osmotic stress and abscisic acid (ABA) signaling pathways; however, the upstream components required for SnRK2 activation and early osmotic stress signaling are still unknown. Here, we report a critical role for B2, B3 and B4 subfamilies of Raf-like kinases (RAFs) in early osmotic stress as well as ABA signaling in Arabidopsis thaliana. B2, B3 and B4 RAFs are quickly activated by osmotic stress and are required for phosphorylation and activation of SnRK2s. Analyses of high-order mutants of RAFs reveal critical roles of the RAFs in osmotic stress tolerance and ABA responses as well as in growth and development. Our findings uncover a kinase cascade mediating osmoregulation in higher plants.
The perception and relay of cell-wall signals are critical for plants to regulate growth and stress responses, but the underlying mechanisms are poorly understood. We found that the cell-wall ...leucine-rich repeat extensins (LRX) 3/4/5 are critical for plant salt tolerance in Arabidopsis. The LRXs physically associate with the RAPID ALKALINIZATION FACTOR (RALF) peptides RALF22/23, which in turn interact with the plasma membrane-localized receptor-like protein kinase FERONIA (FER). The lrx345 triplemutant as well as fer mutant plants display retarded growth and salt hypersensitivity, which are mimicked by overexpression of RALF22/23. Salt stress promotes S1P protease-dependent release of mature RALF22 peptides. Treatment of roots with mature RALF22/23 peptides or salt stress causes the internalization of FER. Our results suggest that the LRXs, RALFs, and FER function as a module to transduce cell-wall signals to regulate plant growth and salt stress tolerance.
Protein kinases are major regulatory components in almost all cellular processes in eukaryotic cells. By adding phosphate groups, protein kinases regulate the activity, localization, protein–protein ...interactions, and other features of their target proteins. It is known that protein kinases are central components in plant responses to environmental stresses such as drought, high salinity, cold, and pathogen attack. However, only a few targets of these protein kinases have been identified. Moreover, how these protein kinases regulate downstream biological processes and mediate stress responses is still largely unknown. In this study, we introduce a strategy based on isotope-labeled in vitro phosphorylation reactions using in vivo phosphorylated peptides as substrate pools and apply this strategy to identify putative substrates of nine protein kinases that function in plant abiotic and biotic stress responses. As a result, we identified more than 5,000 putative target sites of osmotic stress-activated SnRK2.4 and SnRK2.6, abscisic acid-activated protein kinases SnRK2.6 and casein kinase 1-like 2 (CKL2), elicitor-activated protein kinase CDPK11 and MPK6, cold-activated protein kinase MPK6, H₂O₂-activated protein kinase OXI1 and MPK6, and saltinduced protein kinase SOS1 and MPK6, as well as the lowpotassium-activated protein kinase CIPK23. These results provide comprehensive information on the role of these protein kinases in the control of cellular activities and could be a valuable resource for further studies on the mechanisms underlying plant responses to environmental stresses.
Abstract
During the last decade, there has been great interest in elucidating the biological role of extracellular vesicles (EVs), particularly, their hormone-like role in cell-to-cell communication. ...The field of endocrinology is uniquely placed to provide insight into the functions of EVs, which are secreted from all cells into biological fluids and carry endocrine signals to engage in paracellular and distal interactions. EVs are a heterogeneous population of membrane-bound vesicles of varying size, content, and bioactivity. EVs are specifically packaged with signaling molecules, including lipids, proteins, and nucleic acids, and are released via exocytosis into biofluid compartments. EVs regulate the activity of both proximal and distal target cells, including translational activity, metabolism, growth, and development. As such, EVs signaling represents an integral pathway mediating intercellular communication. Moreover, as the content of EVs is cell-type specific, it is a “fingerprint” of the releasing cell and its metabolic status. Recently, changes in the profile of EV and bioactivity have been described in several endocrine-related conditions including diabetes, obesity, cardiovascular diseases, and cancer. The goal of this statement is to highlight relevant aspects of EV research and their potential role in the field of endocrinology.
The outbreak of Zika virus (ZIKV) in 2016 created worldwide health emergency which demand urgent research efforts on understanding the virus biology and developing therapeutic strategies. Here, we ...present a time-resolved chemical proteomic strategy to track the early-stage entry of ZIKV into host cells. ZIKV was labeled on its surface with a chemical probe, which carries a photocrosslinker to covalently link virus-interacting proteins in living cells on UV exposure at different time points, and a biotin tag for subsequent enrichment and mass spectrometric identification of the receptor or other host proteins critical for virus internalization. We identified Neural Cell Adhesion Molecule (NCAM1) as a potential ZIKV receptor and further validated it through overexpression, knockout, and inhibition of NCAM1 in Vero cells and human glioblastoma cells U-251 MG. Collectively, the strategy can serve as a universal tool to map virus entry pathways and uncover key interacting proteins.
Auxin determines the developmental fate of plant tissues, and local auxin concentration is precisely controlled. The role of auxin transport in modulating local auxin concentration has been widely ...studied but the regulation of local auxin biosynthesis is less well understood. Here, we show that TRYPTOPHAN AMINOTRANSFERASE OF ARABIDOPSIS (TAA1), a key enzyme in the auxin biosynthesis pathway in Arabidopsis thaliana is phosphorylated at Threonine 101 (T101). T101 phosphorylation status can act as an on/off switch to control TAA1-dependent auxin biosynthesis and is required for proper regulation of root meristem size and root hair development. This phosphosite is evolutionarily conserved suggesting post-translational regulation of auxin biosynthesis may be a general phenomenon. In addition, we show that auxin itself, in part via TRANS-MEMBRANE KINASE 4 (TMK4), can induce T101 phosphorylation of TAA1 suggesting a self-regulatory loop whereby local auxin signalling can suppress biosynthesis. We conclude that phosphorylation-dependent control of TAA1 enzymatic activity may contribute to regulation of auxin concentration in response to endogenous and/or external cues.
Aptamer in Bioanalytical Applications Iliuk, Anton B; Hu, Lianghai; Tao, W. Andy
Analytical chemistry (Washington),
06/2011, Letnik:
83, Številka:
12
Journal Article
The state of protein phosphorylation can be a key determinant of cellular physiology such as early-stage cancer, but the development of phosphoproteins in biofluids for disease diagnosis remains ...elusive. Here we demonstrate a strategy to isolate and identify phosphoproteins in extracellular vesicles (EVs) from human plasma as potential markers to differentiate disease from healthy states. We identified close to 10,000 unique phosphopeptides in EVs isolated from small volumes of plasma samples. Using label-free quantitative phosphoproteomics, we identified 144 phosphoproteins in plasma EVs that are significantly higher in patients diagnosed with breast cancer compared with healthy controls. Several biomarkers were validated in individual patients using paralleled reaction monitoring for targeted quantitation. This study demonstrates that the development of phosphoproteins in plasma EV as disease biomarkers is highly feasible and may transform cancer screening and monitoring.
As sessile organisms, plants must adapt to variations in the environment. Environmental stress triggers various responses, including growth inhibition, mediated by the plant hormone abscisic acid ...(ABA). The mechanisms that integrate stress responses with growth are poorly understood. Here, we discovered that the Target of Rapamycin (TOR) kinase phosphorylates PYL ABA receptors at a conserved serine residue to prevent activation of the stress response in unstressed plants. This phosphorylation disrupts PYL association with ABA and with PP2C phosphatase effectors, leading to inactivation of SnRK2 kinases. Under stress, ABA-activated SnRK2s phosphorylate Raptor, a component of the TOR complex, triggering TOR complex dissociation and inhibition. Thus, TOR signaling represses ABA signaling and stress responses in unstressed conditions, whereas ABA signaling represses TOR signaling and growth during times of stress. Plants utilize this conserved phospho-regulatory feedback mechanism to optimize the balance of growth and stress responses.
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•The TOR kinase phosphorylates ABA receptor PYLs at a conserved serine residue•PYLs phosphorylation inhibits stress responses by abolishing PYLs activities•Stress- and ABA-activated SnRK2s phosphorylate Raptor and inhibit TOR activity•TOR and ABA signaling balance plant growth and stress responses
Wang et al. reveal that the TOR kinase phosphorylates ABA receptors to repress stress responses under unstressed conditions and to promote growth recovery once environmental stresses subside. Under stress conditions, SnRK2s phosphorylate Raptor, a regulatory component in the TOR complex, to prevent growth by inhibiting TOR activity.
Significance Drought stress induces the accumulation of the plant stress hormone abscisic acid (ABA). ABA then quickly activates the protein kinase OST1/SnRK2.6 to phosphorylate a number of proteins ...in guard cells, resulting in stomatal closure to reduce transpirational water loss. How SnRK2.6 is deactivated and how ABA signaling may be desensitized are unclear. This study found that nitric oxide (NO) resulting from ABA signaling causes S-nitrosylation of SnRK2.6 at a cysteine residue close to the kinase catalytic site, which blocks the kinase activity. Dysfunction of S-nitrosoglutathione (GSNO) reductase causes GSNO overaccumulation in guard cells and ABA insensitivity in stomatal regulation. This work thus reveals how ABA-induced NO functions in guard cells to inactivate SnRK2.6 to negatively feedback regulate ABA signaling.
The phytohormone abscisic acid (ABA) plays important roles in plant development and adaptation to environmental stress. ABA induces the production of nitric oxide (NO) in guard cells, but how NO regulates ABA signaling is not understood. Here, we show that NO negatively regulates ABA signaling in guard cells by inhibiting open stomata 1 (OST1)/sucrose nonfermenting 1 (SNF1)-related protein kinase 2.6 (SnRK2.6) through S-nitrosylation. We found that SnRK2.6 is S-nitrosylated at cysteine 137, a residue adjacent to the kinase catalytic site. Dysfunction in the S-nitrosoglutathione (GSNO) reductase (GSNOR) gene in the gsnor1-3 mutant causes NO overaccumulation in guard cells, constitutive S-nitrosylation of SnRK2.6, and impairment of ABA-induced stomatal closure. Introduction of the Cys137 to Ser mutated SnRK2.6 into the gsnor1-3/ost1-3 double-mutant partially suppressed the effect of gsnor1-3 on ABA-induced stomatal closure. A cysteine residue corresponding to Cys137 of SnRK2.6 is present in several yeast and human protein kinases and can be S-nitrosylated, suggesting that the S-nitrosylation may be an evolutionarily conserved mechanism for protein kinase regulation.
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