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Nature Feb 2022Mutations in the protein kinase PINK1 lead to defects in mitophagy and cause autosomal recessive early onset Parkinson's disease. PINK1 has many unique features that...
Mutations in the protein kinase PINK1 lead to defects in mitophagy and cause autosomal recessive early onset Parkinson's disease. PINK1 has many unique features that enable it to phosphorylate ubiquitin and the ubiquitin-like domain of Parkin. Structural analysis of PINK1 from diverse insect species with and without ubiquitin provided snapshots of distinct structural states yet did not explain how PINK1 is activated. Here we elucidate the activation mechanism of PINK1 using crystallography and cryo-electron microscopy (cryo-EM). A crystal structure of unphosphorylated Pediculus humanus corporis (Ph; human body louse) PINK1 resolves an N-terminal helix, revealing the orientation of unphosphorylated yet active PINK1 on the mitochondria. We further provide a cryo-EM structure of a symmetric PhPINK1 dimer trapped during the process of trans-autophosphorylation, as well as a cryo-EM structure of phosphorylated PhPINK1 undergoing a conformational change to an active ubiquitin kinase state. Structures and phosphorylation studies further identify a role for regulatory PINK1 oxidation. Together, our research delineates the complete activation mechanism of PINK1, illuminates how PINK1 interacts with the mitochondrial outer membrane and reveals how PINK1 activity may be modulated by mitochondrial reactive oxygen species.
Topics: Animals; Cryoelectron Microscopy; Insect Proteins; Mitochondria; Mitophagy; Pediculus; Phosphorylation; Protein Conformation; Protein Kinases; Ubiquitin
PubMed: 34933320
DOI: 10.1038/s41586-021-04340-2 -
Nature Cell Biology May 2023Activation of receptor protein kinases is prevalent in various cancers with unknown impact on ferroptosis. Here we demonstrated that AKT activated by insulin-like growth...
Activation of receptor protein kinases is prevalent in various cancers with unknown impact on ferroptosis. Here we demonstrated that AKT activated by insulin-like growth factor 1 receptor signalling phosphorylates creatine kinase B (CKB) T133, reduces metabolic activity of CKB and increases CKB binding to glutathione peroxidase 4 (GPX4). Importantly, CKB acts as a protein kinase and phosphorylates GPX4 S104. This phosphorylation prevents HSC70 binding to GPX4, thereby abrogating the GPX4 degradation regulated by chaperone-mediated autophagy, alleviating ferroptosis and promoting tumour growth in mice. In addition, the levels of GPX4 are positively correlated with the phosphorylation levels of CKB T133 and GPX4 S104 in human hepatocellular carcinoma specimens and associated with poor prognosis of patients with hepatocellular carcinoma. These findings reveal a critical mechanism by which tumour cells counteract ferroptosis by non-metabolic function of CKB-enhanced GPX4 stability and underscore the potential to target the protein kinase activity of CKB for cancer treatment.
Topics: Animals; Humans; Mice; Carcinoma, Hepatocellular; Creatine Kinase; Ferroptosis; Liver Neoplasms; Phosphorylation
PubMed: 37156912
DOI: 10.1038/s41556-023-01133-9 -
Cell Reports Jul 2023Protein phosphorylation modification is crucial for signaling transduction in plant development and environmental adaptation. By precisely phosphorylating crucial... (Review)
Review
Protein phosphorylation modification is crucial for signaling transduction in plant development and environmental adaptation. By precisely phosphorylating crucial components in signaling cascades, plants can switch on and off the specific signaling pathways needed for growth or defense. Here, we have summarized recent findings of key phosphorylation events in typical hormone signaling and stress responses. More interestingly, distinct phosphorylation patterns on proteins result in diverse biological functions of these proteins. Thus, we have also highlighted latest findings that show how the different phosphosites of a protein, also named phosphocodes, determine the specificity of downstream signaling in both plant development and stress responses.
Topics: Phosphorylation; Signal Transduction; Plants; Plant Development; Plant Proteins
PubMed: 37405922
DOI: 10.1016/j.celrep.2023.112729 -
Nature Plants Apr 2023Phosphorylation modification is required for the modulation of phytochrome B (phyB) thermal reversion, but the kinase(s) that phosphorylate(s) phyB and the biological...
Phosphorylation modification is required for the modulation of phytochrome B (phyB) thermal reversion, but the kinase(s) that phosphorylate(s) phyB and the biological significance of the phosphorylation are still unknown. Here we report that FERONIA (FER) phosphorylates phyB to regulate plant growth and salt tolerance, and the phosphorylation not only regulates dark-triggered photobody dissociation but also modulates phyB protein abundance in the nucleus. Further analysis indicates that phosphorylation of phyB by FER is sufficient to accelerate the conversion of phyB from the active form (Pfr) to the inactive form (Pr). Under salt stress, FER kinase activity is inhibited, leading to delayed photobody dissociation and increased phyB protein abundance in the nucleus. Our data also show that phyB mutation or overexpression of PIF5 attenuates growth inhibition and promotes plant survival under salt stress. Together, our study not only reveals a kinase that controls phyB turnover via a signature of phosphorylation, but also provides mechanistic insights into the role of the FER-phyB module in coordinating plant growth and stress tolerance.
Topics: Phytochrome B; Arabidopsis; Arabidopsis Proteins; Phosphorylation; Salt Tolerance; Plants; Light; Gene Expression Regulation, Plant
PubMed: 37012430
DOI: 10.1038/s41477-023-01390-4 -
Chemical Society Reviews Jul 2022Protein phosphorylation is a crucial regulator of protein and cellular function, yet, despite identifying an enormous number of phosphorylation sites, the role of most... (Review)
Review
Protein phosphorylation is a crucial regulator of protein and cellular function, yet, despite identifying an enormous number of phosphorylation sites, the role of most is still unclear. Each phosphoform, the particular combination of phosphorylations, of a protein has distinct and diverse biological consequences. Aberrant phosphorylation is implicated in the development of many diseases. To investigate their function, access to defined protein phosphoforms is essential. Materials obtained from cells often are complex mixtures. Recombinant methods can provide access to defined phosphoforms if site-specifically acting kinases are known, but the methods fail to provide homogenous material when several amino acid side chains compete for phosphorylation. Chemical and chemoenzymatic synthesis has provided an invaluable toolbox to enable access to previously unreachable phosphoforms of proteins. In this review, we selected important tools that enable access to homogeneously phosphorylated protein and discuss examples that demonstrate how they can be applied. Firstly, we discuss the synthesis of phosphopeptides and proteins through chemical and enzymatic means and their advantages and limitations. Secondly, we showcase illustrative examples that applied these tools to answer biological questions pertaining to proteins involved in signal transduction, control of transcription, neurodegenerative diseases and aggregation, apoptosis and autophagy, and transmembrane proteins. We discuss the opportunities and challenges in the field.
Topics: Biology; Phosphopeptides; Phosphorylation; Proteins; Signal Transduction
PubMed: 35726784
DOI: 10.1039/d1cs00991e -
Nature Plants Jan 2022The root:shoot ratio has long been known to be enhanced in plants under drought stress. Here we discovered that osmotic stress enhances long-distance sucrose transport...
The root:shoot ratio has long been known to be enhanced in plants under drought stress. Here we discovered that osmotic stress enhances long-distance sucrose transport to increase the root:shoot ratio in an abscisic-acid-dependent manner. The Arabidopsis sucrose transporters SWEET11 and 12, key players in phloem loading, are rapidly phosphorylated upon drought and abscisic acid treatments. The drought- and abscisic-acid-activated SnRK2 protein kinases phosphorylate the carboxy-terminal cytosolic regions of SWEET11 and 12. This phosphorylation enhances the oligomerization and sucrose transport activity of SWEETs, which results in elevated sucrose contents in roots and improved root growth under drought stress, leading to the enhanced root:shoot ratio of biomass and drought resistance. Notably, the expression of phospho-mimic SWEETs led to improved root growth even under non-stressed conditions. The phosphorylation of sucrose transporters provides an explanation for the long-standing observation that drought stress enhances the root:shoot ratio in plants and suggests a strategy for engineering drought-resistant crops.
Topics: Abscisic Acid; Droughts; Gene Expression Regulation, Plant; Phosphorylation; Plant Roots; Stress, Physiological; Sucrose
PubMed: 34949800
DOI: 10.1038/s41477-021-01040-7 -
Developmental Cell Sep 2023Salt stress is one of the unfavorable environmental factors to affect plants. Salinity represses root growth, resulting in reduced biomass of agricultural plants. Little...
Salt stress is one of the unfavorable environmental factors to affect plants. Salinity represses root growth, resulting in reduced biomass of agricultural plants. Little is known about how plants maintain root growth to counteract salt stress. The AP2-domain transcription factors PLETHORA1/2 (PLT1/2) act as master regulators in root meristem maintenance in Arabidopsis. In this study, we report that the salt overly sensitive (SOS) pathway component SOS2 regulates PLT1/2 at the post-transcriptional level. Salt-activated SOS2 interacts and phosphorylates PLT1/2 through their conserved C-terminal motifs to stabilize PLT1/2, critical for root apical meristem maintenance under salt stress. The phospho-mimetic version of PLT1/2 restored meristem and primary root length reduction of sos2-2 and plt1-4 plt2-2 mutants on salt treatment. Moreover, SOS2-mediated PLT1/2 phosphorylation improves root growth recovery after salt stress alleviation. We identify a SOS2-PLT1/2 core protein module that is required for protecting primary root growth and meristem maintenance from salt stress.
Topics: Meristem; Signal Transduction; Arabidopsis; Phosphorylation; Salt Stress
PubMed: 37480843
DOI: 10.1016/j.devcel.2023.06.012 -
Signal Transduction and Targeted Therapy Mar 2023Epithelial to mesenchymal transition (EMT) plays a crucial role in cancer metastasis, accompanied with vast epigenetic changes. AMP-activated protein kinase (AMPK), a...
Epithelial to mesenchymal transition (EMT) plays a crucial role in cancer metastasis, accompanied with vast epigenetic changes. AMP-activated protein kinase (AMPK), a cellular energy sensor, plays regulatory roles in multiple biological processes. Although a few studies have shed light on AMPK regulating cancer metastasis, the inside epigenetic mechanisms remain unknown. Herein we show that AMPK activation by metformin relieves the repressive H3K9me2-mediated silencing of epithelial genes (e.g., CDH1) during EMT processes and inhibits lung cancer metastasis. PHF2, a H3K9me2 demethylase, was identified to interact with AMPKα2. Genetic deletion of PHF2 aggravates lung cancer metastasis and abolishes the H3K9me2 downregulation and anti-metastasis effect of metformin. Mechanistically, AMPK phosphorylates PHF2 at S655 site, enhancing PHF2 demethylation activity and triggering the transcription of CDH1. Furthermore, the PHF2-S655E mutant that mimics AMPK-mediated phosphorylation status further reduces H3K9me2 and suppresses lung cancer metastasis, while PHF2-S655A mutant presents opposite phenotype and reverses the anti-metastasis effect of metformin. PHF2-S655 phosphorylation strikingly reduces in lung cancer patients and the higher phosphorylation level predicts better survival. Altogether, we reveal the mechanism of AMPK inhibiting lung cancer metastasis via PHF2 mediated H3K9me2 demethylation, thereby promoting the clinical application of metformin and highlighting PHF2 as the potential epigenetic target in cancer metastasis.
Topics: Humans; Phosphorylation; AMP-Activated Protein Kinases; Epithelial-Mesenchymal Transition; Lung Neoplasms; Metformin; Homeodomain Proteins
PubMed: 36872368
DOI: 10.1038/s41392-022-01302-6 -
Nature Communications Aug 2023PPARα corepressor NCoR1 is a key regulator of fatty acid β-oxidation and ketogenesis. However, its regulatory mechanism is largely unknown. Here, we report that...
PPARα corepressor NCoR1 is a key regulator of fatty acid β-oxidation and ketogenesis. However, its regulatory mechanism is largely unknown. Here, we report that oncoprotein p21-activated kinase 4 (PAK4) is an NCoR1 kinase. Specifically, PAK4 phosphorylates NCoR1 at T1619/T2124, resulting in an increase in its nuclear localization and interaction with PPARα, thereby repressing the transcriptional activity of PPARα. We observe impaired ketogenesis and increases in PAK4 protein and NCoR1 phosphorylation levels in liver tissues of high fat diet-fed mice, NAFLD patients, and hepatocellular carcinoma patients. Forced overexpression of PAK4 in mice represses ketogenesis and thereby increases hepatic fat accumulation, whereas genetic ablation or pharmacological inhibition of PAK4 exhibites an opposite phenotype. Interestingly, PAK4 protein levels are significantly suppressed by fasting, largely through either cAMP/PKA- or Sirt1-mediated ubiquitination and proteasome degradation. In this way, our findings provide evidence for a PAK4-NCoR1/PPARα signaling pathway that regulates fatty acid β-oxidation and ketogenesis.
Topics: Animals; Mice; Co-Repressor Proteins; Fatty Acids; p21-Activated Kinases; PPAR alpha; Nuclear Receptor Co-Repressor 1; Humans; Phosphorylation; Signal Transduction
PubMed: 37591884
DOI: 10.1038/s41467-023-40597-z -
Molecular Cell Jun 2022Protein phosphorylation is a reversible post-translational modification. Nine of the 20 natural amino acids in proteins can be phosphorylated, but most of what we know... (Review)
Review
Protein phosphorylation is a reversible post-translational modification. Nine of the 20 natural amino acids in proteins can be phosphorylated, but most of what we know about the roles of protein phosphorylation has come from studies of serine, threonine, and tyrosine phosphorylation. Much less is understood about the phosphorylation of histidine, lysine, arginine, cysteine, aspartate, and glutamate, so-called non-canonical phosphorylations. Phosphohistidine (pHis) was discovered 60 years ago as a mitochondrial enzyme intermediate; since then, evidence for the existence of histidine kinases and phosphohistidine phosphatases has emerged, together with examples where protein function is regulated by reversible histidine phosphorylation. pHis is chemically unstable and has thus been challenging to study. However, the recent development of tools for studying pHis has accelerated our understanding of the multifaceted functions of histidine phosphorylation, revealing a large number of proteins that are phosphorylated on histidine and implicating pHis in a wide range of cellular processes.
Topics: Histidine; Phosphorylation; Phosphotyrosine; Proteins
PubMed: 35654043
DOI: 10.1016/j.molcel.2022.05.007