Eukaryotic-like protein serine/threonine kinases in Myxococcus xanthus,a developmental bacterium exhibiting social behavior

Myxococcus xanthus, a gram-negative bacterium exhibits a spectacular life cycle and social behavior. Its developmental cycle and multicellular morphogenesis resemble those of eukaryotic slime molds such as Dictyostelium discoideum. On the basis of this resemblance, we explored the existence of eukaryotic-like protein serine/threonine kinases which are known to play important roles in signal transduction during development of D. discoideum. It was indeed found that M. xanthus contains a large family of protein serine/threonine kinases related to the eukaryotic enzymes. This is the first unambiguous demonstration of eukaryotic-like protein serine/threonine kinases in the prokaryotes. © 1993 Wiley-Liss, Inc.     

Effects of 6-Dimethylaminopurine, 2-Aminopurine, Olomoucine and Sodium Vanadate on DNA Endoreduplication in Primary Roots of Pisum Sativum

The effects of 2-aminopurine, olomoucine, 6-dimethylaminopurine (inhibitors of cyclin-dependent kinases; CDK), and sodium vanadate (a potent inhibitor of protein phosphatases) on DNA endoreduplication were investigated during elongation and differentiation of the primary roots in Pisum sativum. When compared with the untreated control plants, at least one additional round of DNA replication was evidenced to occur within most cells, the majority of which have attained 4C DNA level, and a considerably greater portion of cells represented the endopolyploid state with nuclear DNA content approximating the 8C value. It is concluded that cellular commitment to DNA endoreduplication may appear not only as a consequence of suppression imposed directly upon CDK activity, but also as an indirect output connected with the decreased activity of cdc25 protein phosphatase, an enzyme necessary to turn the switch on for appropriate conformation of the CDK/cyclin B complex. By calculating the absorption profiles of Feulgen-stained nuclei, specific phosphorylation-dependent changes in chromatin condensation of endopolyploid cells have been revealed. It is proposed that acquisition of a certain critical level of chromatin condensation constitutes a prerequisite for additional rounds of DNA synthesis in plants.     

Developmental regulation of tau phosphorylation, tau kinases, and tau phosphatases

Tau is a neuronal microtubule-associated protein. Its hyperphosphorylation plays a critical role in Alzheimer disease (AD). Expression and phosphorylation of tau are regulated developmentally, but its dynamic regulation and the responsible kinases or phosphatases remain elusive. Here, we studied the developmental regulation of tau in rats during development from embryonic day 15 through the age of 24 months. We found that tau expression increased sharply during the embryonic stage and then became relatively stable, whereas tau phosphorylation was much higher in developing brain than in mature brain. However, the extent of tau phosphorylation at seven of the 14 sites studied was much less in developing brain than in AD brain. Tau phosphorylation during development matched the period of active neurite outgrowth in general. Tau phosphorylation at various sites had different topographic distributions. Several tau kinases appeared to regulate tau phosphorylation collectively at overlapping sites, and the decrease of overall tau phosphorylation in adult brain might be due to the higher levels of tau phosphatases in mature brain. These studies provide new insight into the developmental regulation of site-specific tau phosphorylation and identify the likely sites required for the abnormal hyperphosphorylation of tau in AD.     

蛋白激酶研究进展：Ⅰ.结构和分类

蛋白激酶含有２５０～３００个氨基酸残基构成的催化功能域,包括１２个功能亚域．这些氨基酸序列折叠成为一个核心催化结构．根据蛋白激酶功能域氨基酸序列比较分析可以找出各蛋白激酶在进化上的亲缘关系,这是蛋白激酶的分类基础．蛋白激酶可分为二大类,一类是丝氨酸／苏氨酸蛋白激酶,另一类是酪氨酸蛋白激酶．丝氨酸／苏氨酸蛋白激酶在进化上出现较早,可分为几个组,每个组又分为许多家族．酪氨酸蛋白激酶在进化上出现较晚,亲缘关系较紧密,同属于一个组,该组也分为许多家族．最近发现的许多双底物特异蛋白激酶,分散在丝氨酸／苏氨酸蛋白激酶的不同家族中．     

Signaling pathway of MAPK/ERK in cell proliferation,differentiation, migration,senescence and apoptosis

Abstract

The generic mitogen-activated protein kinases (MAPK) signaling pathway is shared by four distinct cascades, including the extracellular signal-related kinases (ERK1/2), Jun amino-terminal kinases (JNK1/2/3), p38-MAPK and ERK5. Mitogen-activated protein kinases/extracellular signal-regulated kinase (MAPK/ERK) pathway is reported to be associated with the cell proliferation, differentiation, migration, senescence and apoptosis. The literatures were searched extensively and this review was performed to review the role of MAPK/ERK signaling pathway in cell proliferation, differentiation, migration, senescence and apoptosis.     

Nature of the intrinsic protein kinases involved in phosphorylation of non-histone proteins in intact prostatic nuclei: further identification of androgen-sensitive protein kinase reactions

Nuclei isolated from rat ventral prostate contain a number of messenger-dependent and -independent protein kinases. Studies were undertaken to determine the relative contribution of these protein kinases in phosphorylation of non-histone proteins (NHPs) in isolated nuclei. The data suggest that messenger-dependent protein kinases such as those dependent on cAMP or Ca²⁺/calmodulin or Ca^2–/phospholipid may be present in very small amounts in intact isolated nuclei, and thus appear not to be significantly involved in phosphorylation of endogenous NHPs. Messenger-independent nuclear associated protein kinases PK-N1 and PK-N2 are known to catalyze the phosphorylation of NHPs in vitro (Goueli SA, et al., Eur J Biochem 113: 45–51, 1980). Of these, the intrinsic heparin-sensitive PK-N2 as compared with heparin-insensitive PK-N1 appeared to be the predominant protein kinase engaged in phosphorylation of NHPs in intact nuclei. About 78–88% of NHP phosphorylation in intact nuclei was inhibited by heparin suggesting that the remaining 12–22% phosphorylation of NHPs was catalyzed via the heparin-insensitive protein kinase(s). Further, the data provide additional evidence that heparin-sensitive PK-N2 is the one that is most responsive to androgenic status in the animal.Abbreviations NHP Non-Histone Protein - PMSF Phenylmethylsulfonyl Fluoride - DTT Dithiothreitol - SDS Sodium Dodecyl Sulfate     

Transmembrane signaling,then and now: The decade of the eighties

The chronology of the major discoveries important in elucidating certain aspects of the molecular basis of transmembrane signaling is briefly reviewed. Recent developments linking cell stimulation and transformation are intimated.The author is the recipient of a Research Career Development Award (AI-00672) and a research grant (AI-28342) from the National Institute of Allergy and Infectious Diseases.     

Pramanicin induces apoptosis in Jurkat leukemia cells: A role for JNK,p38 and caspase activation

Pramanicin is a novel anti-fungal drug with a wide range of potential application against human diseases. It has been previously shown that pramanicin induces cell death and increases calcium levels in vascular endothelial cells. In the present study, we showed that pramanicin induced apoptosis in Jurkat T leukemia cells in a dose- and time-dependent manner. Our data reveal that pramanicin induced the release of cytochrome c and caspase-9 and caspase-3 activation, as evidenced by detection of active caspase fragments and fluorometric caspase assays. Pramanicin also activated c-jun N-terminal kinase (JNK), p38 and extracellular signal-regulated kinases (ERK 1/2) with different time and dose kinetics. Treatment of cells with specific MAP kinase and caspase inhibitors further confirmed the mechanistic involvement of these signalling cascades in pramanicin-induced apoptosis. JNK and p38 pathways acted as pro-apoptotic signalling pathways in pramanicin-induced apoptosis, in which they regulated release of cytochrome c and caspase activation. In contrast the ERK 1/2 pathway exerted a protective effect through inhibition of cytochrome c leakage from mitochondria and caspase activation, which were only observed when lower concentrations of pramanicin were used as apoptosis-inducing agent and which were masked by the intense apoptosis induction by higher concentrations of pramanicin. These results suggest pramanicin as a potential apoptosis-inducing small molecule, which acts through a well-defined JNK- and p38-dependent apoptosis signalling pathway in Jurkat T leukemia cells.     

Insulin-like Effects of Vanadium: Mechanisms of Action,Clinical and Basic Implications

Summary Most or all mammalian cells contain vanadium at a concentration of 20 nM. The bulk of the vanadium in cells is probably in the reduced vanadyl (IV) form. Although this element is essential and should be present in the diet in minute quatities, no known physiological role for vanadium has been found thus far. In the years 1975–1980 the vanadate ion was shown to act as an efficient inhibitor of Na⁺, K⁺-ATPase and of other related phosphohydrolases as well. In 1980 it was observed that vanadate and vanadyl, when added to intact rat adipocytes, mimic the biological actions of insulin in stimulating hexose uptake and glucose oxidation. This initiated a long, currently active, field of research among basic scientists and diabetologists. Several of the aspects studied are reviewed here.     

Scaffolding by ERK3 regulates MK5 in development

Extracellular-regulated kinase 3 (ERK3, MAPK6) is an atypical member of the ERKs, lacking the threonine and tyrosine residues in the activation loop, carrying a unique C-terminal extension and being mainly regulated by its own protein stability and/or by autophosphorylation. Here we show that ERK3 specifically interacts with the MAPK-activated protein kinase 5 (MK5 or PRAK) in vitro and in vivo. Expression of ERK3 in mammalian cells leads to nuclear-cytoplasmic translocation and activation of MK5 and to phosphorylation of both ERK3 and MK5. Remarkably, activation of MK5 is independent of ERK3 enzymatic activity, but depends on its own catalytic activity as well as on a region in the C-terminal extension of ERK3. In mouse embryonic development, mRNA expression patterns of ERK3 and MK5 suggest spatiotemporal coexpression of both kinases. Deletion of MK5 leads to strong reduction of ERK3 protein levels and embryonic lethality at about stage E11, where ERK3 expression in wild-type mice is maximum, indicating a role of this signalling module in development.     

Insulin-like Effects of Vanadium: Mechanisms of Action, Clinical and Basic Implications

Most or all mammalian cells contain vanadium at a concentration of 20 nM. The bulk of the vanadium in cells is probably in the reduced vanadyl (IV) form. Although this element is essential and should be present in the diet in minute quantities, no known physiological role for vanadium has been found thus far. In the years 1975–1980 the vanadate ion was shown to act as an efficient inhibitor of Na⁺,K⁺-ATPase and of other related phosphohydrolases as well. In 1980 it was observed that vanadate and vanadyl, when added to intact rat adipocytes, mimic the biological actions of insulin in stimulating hexose uptake and glucose oxidation. This initiated a long, currently active, field of research among basic scientists and diabetologists. Several of the aspects studied are reviewed here.     

酪氨酸蛋白激酶和蛋白激酶C对N-乙酰氨基葡萄糖转移酶V的双重调控

在人肝癌细胞７７２１中研究了酪氨酸蛋白激酶（ＴＰＫ）和蛋白激酶Ｃ（ＰＫＣ）的激活剂［分别为表皮生长因子（ＥＧＦ）和佛波酯（ＰＭＡ）］和各种蛋白激酶抑制剂对Ｎ－乙酰氨基葡萄糖转移酶Ｖ（ＧｎＴ－Ｖ）活力的影响,以探讨ＴＰＫ和ＰＫＣ对ＧｎＴ－Ｖ的调节。结果发现,ＥＧＦ或ＰＭＡ处理细胞４８ｈ后,ＧｎＴ－Ｖ的活力明显增高;蛋白激酶的非特异性抑制剂槲皮素和染料木黄酮（ｇｅｎｉｓｔｅｉｎ）在抑制ＴＰＫ和ＰＫＣ的同时,抑制ＧｎＴ－Ｖ的基础活力,并完全阻断ＥＧＦ或ＰＭＡ对ＧｎＴ－Ｖ的增高作用;ＴＰＫ的特异性抑制剂Ｔｙｒｐｈｏｓｔｉｎ－２５和ＰＫＣ的特异性抑制剂Ｄ－鞘氨醇分别应用时,各自只能部分地取消ＥＧＦ或ＰＭＡ对ＧｎＴ－Ｖ的诱导。但当Ｔｙｒｐｈｏｓｔｉｎ－２５和Ｄ－鞘氨醇同时加入培养基中则可完全阻断ＥＧＦ或ＰＭＡ对ＧｎＴ－Ｖ的诱导激活。蛋白质合成抑制剂环己亚胺和蛋白激酶抑制剂作用相仿,不但可抑制ＧｎＴ－Ｖ的基础活力,也可完全消除ＥＧＦ或ＰＭＡ对ＧｎＴ－Ｖ的激活。以上结果提示ＥＧＦ或ＰＭＡ通过蛋白激酶调节ＧｎＴ－Ｖ的酶蛋白合成,并且ＧｎＴ－Ｖ受到膜性ＴＰＫ和ＰＫＣ的双重调节,其中ｍ－ＴＰＫ较ｍ－ＰＫＣ更为重要。     

The eukaryotic protein kinase superfamily of the necrotrophic fungal plant pathogen,Sclerotinia sclerotiorum

Protein kinases have been implicated in the regulation of many processes that guide pathogen development throughout the course of infection. A survey of the Sclerotinia sclerotiorum genome for genes encoding proteins containing the highly conserved eukaryotic protein kinase (ePK) domain, the largest protein kinase superfamily, revealed 92 S. sclerotiorum ePKs. This review examines the composition of the S. sclerotiorum ePKs based on conserved motifs within the ePK domain family, and relates this to orthologues found in other filamentous fungi and yeasts. The ePKs are also discussed in terms of their proposed role(s) in aspects of host pathogenesis, including the coordination of mycelial growth/development and deployment of pathogenicity determinants in response to environmental stimuli, nutrients and stress.     

The proline-rich,extensin-like receptor kinase-1 (PERK1) gene is rapidly induced by wounding

We report the isolation and characterization of PERK1 (Proline Extensin-like Receptor Kinase 1), a novel plant RLK from Brassica napusthat is predicted to consist of a proline-rich extracellular domain with sequence similarity to extensins, a transmembrane region, and a catalytic domain possessing serine/threonine kinase activity. Database searches with the predicted PERK1 amino acid sequence also led to the identification of a predicted family of related genes in the Arabidopsis genome. Using biolistic bombardment of onion epidermal cells, we have shown that a PERK1-GFP fusion is localized to the plasma membrane as predicted for a receptor kinase. Given the similarity of PERK1's extracellular domain to extensins, a possible role in plant defense responses was investigated by treating B. napus tissue with mechanical stresses and infection with the fungal pathogen, Sclerotinia sclerotiorum. Various wounding stimuli resulted in a dramatic and rapid accumulation of PERK1 mRNA. Levels of PERK1 mRNA also increased moderately in response to infection by the fungal pathogen S. sclerotiorum. Given the kinetics of PERK1 mRNA accumulation in response to these treatments, PERK1 may be involved early on in the general perception and response to a wound and/or pathogen stimulus.     

Phosphorylation of vertebrate nonmuscle and smooth muscle myosin heavy chains and light chains

In this article we review the various amino acids present in vertebrate nonmuscle and smooth muscle myosin that can undergo phosphorylation. The sites for phosphorylation in the 20 kD myosin light chain include serine-19 and threonine-18 which are substrates for myosin light chain kinase and serine-1 and/or-2 and threonine-9 which are substrates for protein kinase C. The sites in vertebrate smooth muscle and nonmuscle myosin heavy chains that can be phosphorylated by protein kinase C and casein kinase II are also summarized.Original data indicating that treatment of human T-lymphocytes (Jurkat cell line) with phorbol 12-myristate 13-acetate results in phosphorylation of both the 20 kD myosin light chain as well as the 200 kD myosin heavy chain is presented. We identified the amino acids phosphorylated in the human T-lymphocytes myosin light chains as serine-1 or serine-2 and in the myosin heavy chains as serine-1917 by 1-dimensional isoelectric focusing of tryptic phosphopeptides. Untreated T-lymphocytes contain phosphate in the serine-19 residue of teh myosin light chain and in a residue tentatively identified as serine-1944 in the myosin heavy chain.Abbreviations MLC myosin light chain - MHC myosin heavy chain - Tris tris(hydroxymethyl)aminomethane - EGTA [ethylenebis(oxyethylenenitrilo)]tetraacetic acid - EDTA ethylenediaminetetraacetate - TPCK N-tosyl-L-phenylalanine chloromethyl ketone - PMA phorbol 12-myristate 13-acetate     

植物中的CDPK/SnRK蛋白激酶家族

在植物中至少存在五类蛋白激酶,这五类都属于CDPK/SnRK家族,它们的结构中含有EF手型结构或者与其相互作用的蛋白质中包含着EF手型结构。CDPK和CCaMKs在C-端都包含EF手型结构,在与钙离子结合后而被激活。SnRK3s结合蛋白包含着三个EF手型,其中一些被钙离子激活。植物两类其它的蛋白激酶家族成员-CaMK和CRK,与钙调素相结合。本文将阐述这些蛋白激酶结构、活性调节以及参与钙信号传递的潜能。     

Phosphorylation of human microtubule-associated protein tau by protein kinases of the AGC subfamily

Intraneuronal inclusions made of hyperphosphorylated microtubule-associated protein tau are a defining neuropathological characteristic of Alzheimer's disease, and of several other neurodegenerative disorders. Many phosphorylation sites in tau are S/TP sites that flank the microtubule-binding repeats. Others are KXGS motifs in the repeats. One site upstream of the repeats lies in a consensus sequence for AGC kinases. This site (S214) is believed to play an important role in the events leading from normal, soluble to filamentous, insoluble tau. Here, we show that all AGC kinases tested phosphorylated S214. RSK1 and p70 S6 kinase also phosphorylated the neighbouring T212, a TP site that conforms weakly to the AGC kinase consensus sequence. MSK1 phosphorylated S214, as well as S262, a KXGS site in the first repeat, and S305 in the second repeat.     

Protein phosphorylation in rat liver mitochondria

Summary Incubation of rat liver mitochondria in the presence of either [³²P] Pi or ₃₂ ^y -P] ATP resulted in a phosphorylation of four proteins with Mr 50, 47, 44 and 36 kDa, respectively. The endogenous phosphorylation of these proteins in the presence of [³²P] Pi was markedly influenced by the osmolarity of the incubation medium and differentially affected by various effectors of mitochondrial functions, such as Ca²⁺, oligomycin, FCCP, arsenite and dichloroacetate. In particular, the 36 kDa protein, unlike the other proteins, appears to be phosphorylated also by direct incorporation of [³²P], independently of respiratory chain-linked ATP synthesis. The four proteins, located in the mitoplasts, seem to be phosphorylated by diiferent protein kinases, as suggested by the observation that the endogenous phosphorylation of 36 kDa protein resulted selectively increased by addition of exogenous protein kinases, such as casein kinases S and TS. A tentative identification of these phosphorylatable protein is discussed.     

Protein Kinases in Neutrophils: A Review

In chemotactic factor-stimulated neutrophils, rapid increases of intracellular levels of cyclic AMP, calcium, and diacylglycerol have been observed and may be linked to protein kinase activation. The study of the physiological role and regulation of protein kinases in the neutrophil and the identification of their substrates has provided valuable information on the molecular mechanism of neutrophil activation. The focus of this review is on those aspects of protein kinases that are relevant to neutrophil activation and on the substrate proteins for these protein kinases. The possible role of protein phosphorylation in neutrophil function is also discussed.