Fusicoccin, 14-3-3 proteins, and defense responses in tomato plants

Fusicoccin (FC) is a fungal toxin that activates the plant plasma membrane H+-ATPase by binding with 14-3-3 proteins, causing membrane hyperpolarization. Here we report on the effect of FC on a gene-for-gene pathogen-resistance response and show that FC application induces the expression of several genes involved in plant responses to pathogens. Ten members of the FC-binding 14-3-3 protein gene family were isolated from tomato (Lycopersicon esculentum) to characterize their role in defense responses. Sequence analysis is suggestive of common biochemical functions for these tomato 14-3-3 proteins, but their genes showed different expression patterns in leaves after challenges. Different specific subsets of 14-3-3 genes were induced after treatment with FC and during a gene-for-gene resistance response. Possible roles for the H+-ATPase and 14-3-3 proteins in responses to pathogens are discussed.     

Divergent expression of claudin -1, -3, -4, -5 and -7 in developing human lung

Background

Claudins are the main components of tight junctions, structures which are associated with cell polarity and permeability. The aim of this study was to analyze the expression of claudins 1, 3, 4, 5, and 7 in developing human lung tissues from 12 to 40 weeks of gestation.

Methods

47 cases were analyzed by immunohistochemisty for claudins 1, 3, 4, 5 and 7. 23 cases were also investigated by quantitative RT-PCR for claudin-1, -3 and -4.

Results

Claudin-1 was expressed in epithelium of bronchi and large bronchioles from week 12 onwards but it was not detected in epithelium of developing alveoli. Claudin-3, -4 and -7 were strongly expressed in bronchial epithelium from week 12 to week 40, and they were also expressed in alveoli from week 16 to week 40. Claudin-5 was expressed strongly during all periods in endothelial cells. It was expressed also in epithelium of bronchi from week 12 to week 40, and in alveoli during the canalicular period. RT-PCR analyses revealed detectable amounts of RNAs for claudins 1, 3 and 4 in all cases studied.

Conclusion

Claudin-1, -3, -4, -5, and -7 are expressed in developing human lung from week 12 to week 40 with distinct locations and in divergent quantities. The expression of claudin-1 was restricted to the bronchial epithelium, whereas claudin-3, -4 and -7 were positive also in alveolar epithelium as well as in the bronchial epithelium. All claudins studied are linked to the development of airways, whereas claudin-3, -4, -5 and -7, but not claudin-1, are involved in the development of acinus and the differentiation of alveolar epithelial cells.     

The 14-3-3 Proteins: Gene,Gene Expression,and Function

14-3-3 Proteins were discovered by Moore and Perez in the soluble extract of bovine brain. These proteins are highly abundant in the brain. In this review 14-3-3 cDNA cloning, nucleotide sequence of 14-3-3 cDNA, the structure of 14-3-3 gene and 14-3-3 gene expression, in situ hybridization of 14-3-3 mRNA in the brain, the function and regulation of 14-3-3 protein, the binding of 14-3-3 protein to other proteins, the effects of 14-3-3 protein on the binding of a protein to other proteins, and the effect on protein kinase, etc., are concisely described. From the recent rapid development of proteom technology, markedly more target proteins of 14-3-3 protein should be discovered.     

Dkk1, -2, and -3 expression in mouse craniofacial development

Summary The Dickkopf family is important for embryogenesis and postnatal development and growth. Dkk1 is a strong head inducer and knockout of this gene leads to absence of anterior head structures, which are predominantly formed through neural crest migration. During early craniofacial development, Dkk1 to Dkk3 show developmentally regulated expression in a number of elements. However, their expression and roles in late times of craniofacial development are largely unknown. This study focuses on the expression profile of Dkk1-3 on late embryonic and early postnatal stages. It was found that Dkks were involved in a variety of craniofacial developmental processes, including facial outgrowth, myogenesis, osteogenesis, palatogenesis, olfactory epithelium and tooth development; and the expression persisted to postnatal stage in the muscles and bones. Their expression patterns suggest important roles in these processes; further study is warranted to elucidate these roles.     

Investigation of the Axonal Transport of Three Acidic, Soluble Proteins (14-3-2, 14-3-3, and S-100) in the Rabbit Visual System

Abstract: The question of whether three acidic, water-soluble proteins (14-3-2, 14-3-3, and S-100, the first and last known to be brain-specific) are axonally transported was investigated in the rabbit visual system. The water-soluble proteins were obtained from individual optic nerves, combined optic tracts and lateral geniculate bodies, superior colliculi, and, in some instances, retinas at various times (1–56 days) after monocular injections of [³H]leucine. These proteins were separated by a two-step polyacrylamide gel electrophore-sis procedure that isolated 14-3-2, 14-3-3, and S-100 almost uncontaminated by other radioactivity. The isolated 14-3-2 and S-100 were demonstrated to be approx. 90% pure by a new method based on retarding the migration of these proteins by immunoadsorption during the first step of electrophoresis. An analysis of the radioactive labeling of the total soluble proteins (TSP) and the isolated acidic proteins revealed that: (1) S-100 was not axonally transported; (2) both 14-3-2 and 14-3-3 were part of one of the slow components of axonal transport (2-4 mm/day); (3) the radioactivity of 14-3-2 and 14-3-3 represented about 2.7% and 3.2%, respectively, of the radioactivity incorporated into the axonally transported TSP; (4) the ultimate distributions of the radioactively labeled 14-3-2 and 14-3-3 were the same (about 70% of each destined for the superior colliculus) and differed from that of the TSP; and (5) the rates of catabolism of the axonally transported 14-3-2 and 14-3-3 were slightly greater than that of the TSP, with half-lives for 14-3-2 and 14-3-3 estimated to be 11 and 10 days, respectively.     

14-3-3 proteins in neurodegeneration

Among the first reported functions of 14-3-3 proteins was the regulation of tyrosine hydroxylase (TH) activity suggesting a possible involvement of 14-3-3 proteins in Parkinson's disease. Since then the relevance of 14-3-3 proteins in the pathogenesis of chronic as well as acute neurodegenerative diseases, including Alzheimer's disease, polyglutamine diseases, amyotrophic lateral sclerosis and stroke has been recognized. The reported function of 14-3-3 proteins in this context are as diverse as the mechanism involved in neurodegeneration, reaching from basal cellular processes like apoptosis, over involvement in features common to many neurodegenerative diseases, like protein stabilization and aggregation, to very specific processes responsible for the selective vulnerability of cellular populations in single neurodegenerative diseases.Here, we review what is currently known of the function of 14-3-3 proteins in nervous tissue focussing on the properties of 14-3-3 proteins important in neurodegenerative disease pathogenesis.     

BtrI, a novel restriction endonuclease, recognises the non-palindromic sequence 5'-CACGTC(-3/-3)-3'

The recognition sequence and cleavage positions of a new restriction endonuclease BTR:I isolated from Bacillus stearothermophilus SE-U62 have been determined. BTR:I belongs to a rare type IIQ of restriction endonucleases, which recognise non-palindromic nucleotide sequences and cleave DNA symmetrically within them.     

The 14-3-3 proteins of Trypanosoma brucei function in motility, cytokinesis, and cell cycle

The cDNAs for two isoforms (I and II) of the 14-3-3 proteins have been cloned and functionally characterized in Trypanosoma brucei. The amino acid sequences of isoforms I and II have 47 and 50% identity to the human tau isoform, respectively, with important conserved features including a potential amphipathic groove for the binding of phosphoserine/phosphothreonine-containing motifs and a nuclear export signal-like domain. Both isoforms are abundantly expressed at approximately equal levels (1-2 x 10(6) molecules/cell) and localized mainly in the cytoplasm. Knockdown by induction of double-stranded RNA of isoform I and/or II in both bloodstream and procyclic forms resulted first in a reduction of cell motility and then significant reduction in cell growth rates and morphological changes; the changes include aberrant numbers of organelles and abnormal shapes and sizes that mimic phenotypes produced by various cytokinesis inhibitors. Morphological and fluorescence-activated cell sorting analysis of the cell cycle suggested that isoforms I and II might play important roles in nuclear (G2-M transition) and cell (M-G1 transition) division. These findings indicate that the 14-3-3 proteins play important roles in cell motility, cytokinesis, and the cell cycle.     

Purification, Properties, and Immunohistochemical Localisation of Human Brain 14-3-3 Protein

Abstract: A protein has been purified from human brain that appears to be the human equivalent of bovine 14-3-3 protein. On polyacrylamide gel electrophoresis the protein migrates as a faster major component, termed 14-3-3-2 protein, and a slower minor component, termed 14-3-3-1 protein, which consists of approximately 12% of the total protein. Both 14-3-3-1 and 14-3-3-2 have a native molecular weight of approximately 67,000. 14-3-3-2 appears to have the subunit composition (αβ; 14-3-3-1 has the composition ββ. Peptide mapping with Stuphvlococcus aureus V8 proteinase shows that α and β subunits are unrelated but the β and β' subunits show some common peptides. Immunoperoxidase labelling shows that 14-3-3 is localised in neurones in the human cerebral cortex. 14-3-3 shows no enolase, creatine kinase, triose phosphate isomerase, ATPase, cyclic nucleotide-dependent protein kinase, or purine nucleoside phosphorylase activity. 14-3-3 does not bind calcium and does not appear to be related to calmodulin, calcineurin, tubulin, neurofilament proteins, clathrin-associated proteins, or tropomyosin. The functional significance of this neuronal protein remains obscure.     

14-3-3蛋白研究进展

14-3-3蛋白是高度保守的、所有真核生物细胞中都普遍存在的、在大多数生物物种中由一个基因家族编码的一类蛋白调控家族。它几乎参与生命体所有的生理反应过程,人们在各种组织细胞中发现了各种不同的14-3-3蛋白。作为与磷酸丝氨酸／苏氨酸结合的第一信号分子,14-3-3蛋白在细胞的信号转导中起着至关重要的作用,尤其是它直接参与调节蛋白激酶和蛋白磷酸化酶的活性,被称为蛋白质与蛋白质相互作用的”桥梁蛋白”;它可以与转录因子结合形成复合体,调节相关基因的表达。一些研究表明,14-3-3蛋白调控机制的紊乱可以直接导致疾病的发生,在临床上14-3-3蛋白常常可以作为诊断的标志物。     

Bioactive recombinant neuregulin-1, -2, and -3 expressed in Escherichia coli

The neuregulins (NRGs) are a family of signaling proteins that are ligands for receptor tyrosine kinase of the ErbB family (namely ErbB3 and ErbB4). To date, four different neuregulin genes have been identified (neuregulin1-4). While NRG1 isoforms have been extensively studied, little is yet known about the other genes of the family. We report the expression of recombinant NRG1beta1, NRG2alpha, NRG2beta, and NRG3 as recombinant fusion proteins in Escherichia coli. The cDNA encoding for the EGF-like domain of each protein was cloned from the mouse olfactory bulb and inserted into the pET-19b vector allowing for bacterial expression of the protein fused to an N-terminal His tag. The recombinant NRGs expressed in the inclusion bodies were solubilized under denaturing conditions, purified by affinity chromatography, and refolded via dialysis in the presence of reducing agents. Purified recombinant NRGs were active as they bound to their receptors and induced their phosphorylation. In particular, and in agreement with data on the native proteins, all the molecules were able to bind and activate ErbB4 while only the rNRG1 and the two rNRG2 (but not rNRG3) bound ErbB3.     

Functional polymorphisms in matrix metalloproteinases -1, -3, -9 and -12 in relation to cervical artery dissection

Background  

Cervical artery dissection is a leading cause of cerebral ischemia in young adults. Morphological investigations have shown alterations in the extracellular matrix (ECM) of affected vessel walls. As matrix metalloproteinases (MMP) play a central role in the regulation of the ECM, an increased expression of these enzymes might lead to the endothelial damage in spontaneous cervical artery dissection (sCAD). Five different DNA polymorphisms in MMP-1, -3, -9 and -12 were tested for their frequency in patients with sCAD and compared with those of a control population.     

Genome-Wide Identification,Classification, and Expression Analysis of 14-3-3 Gene Family in Populus

Background

In plants, 14-3-3 proteins are encoded by a large multigene family and are involved in signaling pathways to regulate plant development and protection from stress. Although twelve Populus 14-3-3s were identified based on the Populus trichocarpa genome V1.1 in a previous study, no systematic analysis including genome organization, gene structure, duplication relationship, evolutionary analysis and expression compendium has been conducted in Populus based on the latest P. trichocarpa genome V3.0.

Principal Findings

Here, a comprehensive analysis of Populus 14-3-3 family is presented. Two new 14-3-3 genes were identified based on the latest P. trichocarpa genome. In P. trichocarpa, fourteen 14-3-3 genes were grouped into ε and non-ε group. Exon-intron organizations of Populus 14-3-3s are highly conserved within the same group. Genomic organization analysis indicated that purifying selection plays a pivotal role in the retention and maintenance of Populus 14-3-3 family. Protein conformational analysis indicated that Populus 14-3-3 consists of a bundle of nine α-helices (α1-α9); the first four are essential for formation of the dimer, while α3, α5, α7, and α9 form a conserved peptide-binding groove. In addition, α1, α3, α5, α7, and α9 were evolving at a lower rate, while α2, α4, and α6 were evolving at a relatively faster rate. Microarray analyses showed that most Populus 14-3-3s are differentially expressed across tissues and upon exposure to various stresses.

Conclusions

The gene structures and their coding protein structures of Populus 14-3-3s are highly conserved among group members, suggesting that members of the same group might also have conserved functions. Microarray and qRT-PCR analyses showed that most Populus 14-3-3s were differentially expressed in various tissues and were induced by various stresses. Our investigation provided a better understanding of the complexity of the 14-3-3 gene family in poplars.     

14-3-3 proteins in neurological disorders

14-3-3 proteins were originally discovered as a family of proteins that are highly expressed in the brain. Through interactions with a multitude of binding partners, 14-3-3 proteins impact many aspects of brain function including neural signaling, neuronal development and neuroprotection. Although much remains to be learned and understood, 14-3-3 proteins have been implicated in a variety of neurological disorders based on evidence from both clinical and laboratory studies. Here we will review previous and more recent research that has helped us understand the roles of 14-3-3 proteins in both neurodegenerative and neuropsychiatric diseases.     

14-3-3 proteins, FHA domains and BRCT domains in the DNA damage response

The DNA damage response depends on the concerted activity of protein serine/threonine kinases and modular phosphoserine/threonine-binding domains to relay the damage signal and recruit repair proteins. The PIKK family of protein kinases, which includes ATM/ATR/DNA-PK, preferentially phosphorylate Ser-Gln sites, while their basophilic downstream effecter kinases, Chk1/Chk2/MK2 preferentially phosphorylate hydrophobic-X-Arg-X-X-Ser/Thr-hydrophobic sites. A subset of tandem BRCT domains act as phosphopeptide binding modules that bind to ATM/ATR/DNA-PK substrates after DNA damage. Conversely, 14-3-3 proteins interact with substrates of Chk1/Chk2/MK2. FHA domains have been shown to interact with substrates of ATM/ATR/DNA-PK and CK2. In this review we consider how substrate phsophorylation together with BRCT domains, FHA domains and 14-3-3 proteins function to regulate ionizing radiation-induced nuclear foci and help to establish the G₂/M checkpoint. We discuss the role of MDC1 a molecular scaffold that recruits early proteins to foci, such as NBS1 and RNF8, through distinct phosphodependent interactions. In addition, we consider the role of 14-3-3 proteins and the Chk2 FHA domain in initiating and maintaining cell cycle arrest.