Characterization of the caleosin gene family in the Triticeae

Background

The caleosin genes encode proteins with a single conserved EF hand calcium-binding domain and comprise small gene families found in a wide range of plant species. Some members of the gene family have been shown to be upregulated by environmental stresses including low water availability and high salinity. Caleosin 3 from wheat has been shown to interact with the α-subunit of the heterotrimeric G proteins, and to act as a GTPase activating protein (GAP). This study characterizes the size and diversity of the gene family in wheat and related species and characterizes the differential tissue-specific expression of members of the gene family.

Results

A total of 34 gene family members that belong to eleven paralogous groups of caleosins were identified in the hexaploid bread wheat, T. aestivum. Each group was represented by three homeologous copies of the gene located on corresponding homeologous chromosomes, except the caleosin 10, which has four gene copies. Ten gene family members were identified in diploid barley, Hordeum vulgare, and in rye, Secale cereale, seven in Brachypodium distachyon, and six in rice, Oryza sativa. The analysis of gene expression was assayed in triticale and rye by RNA-Seq analysis of 454 sequence sets and members of the gene family were found to have diverse patterns of gene expression in the different tissues that were sampled in rye and in triticale, the hybrid hexaploid species derived from wheat and rye. Expression of the gene family in wheat and barley was also previously determined by microarray analysis, and changes in expression during development and in response to environmental stresses are presented.

Conclusions

The caleosin gene family had a greater degree of expansion in the Triticeae than in the other monocot species, Brachypodium and rice. The prior implication of one member of the gene family in the stress response and heterotrimeric G protein signaling, points to the potential importance of the caleosin gene family. The complexity of the family and differential expression in various tissues and under conditions of abiotic stress suggests the possibility that caleosin family members may play diverse roles in signaling and development that warrants further investigation.

Electronic supplementary material

The online version of this article (doi:10.1186/1471-2164-15-239) contains supplementary material, which is available to authorized users.     

Acidic extracellular pH induces p120-catenin-mediated disruption of adherens junctions via the Src kinase-PKCδ pathway

An acidic microenvironment induces disruption of adherens junctions (AJs) of hepatoma cells. This study investigated the impact of an acidic extracellular pH (pHe) on p120-catenin (p120-ctn) serine phosphorylation. pH 6.6 treatment increased intracellular calcium levels, activated protein kinase C (PKC)α and PKCδ, and decreased serine phosphorylation of p120-ctn. Further knockdown of PKCα and δ by small interference RNA (siRNA) prevented the pH 6.6-induced downregulation of p120-ctn at AJ and the serine dephosphorylation of p120-ctn. Moreover, PP2 pretreatment and siRNA of c-Src abrogated the pH 6.6-induced PKCδ activation. Together, the c-Src-PKCδ cascade and PKCα regulate the acidic pHe-induced AJ disruption.     

Maternal obesity is associated with younger age at obesity onset in U.S. adolescent offspring followed into adulthood

Objective: The objective was to test the hypothesis that maternal obesity is associated with younger age of offspring's obesity onset. Research Methods and Procedures: We used prospective, nationally representative, longitudinal data collected across Waves I (1995; 12 to 20 years), II (1996; 13 to 20 years), and III (2001; 18 to 28 years) of the National Longitudinal Study of Adolescent Health (N = 14,654; 49% female). Interval regression analysis was used to assess the association between maternal obesity and age at offspring's obesity onset (International Obesity Task Force BMI ≥30 equivalent age‐ and sex‐specific cut‐off points for adolescents and BMI ≥30 for young adults) using self‐reported heights and weights, adjusting for race/ethnicity, sex, parental education, and family income, accounting for complex sampling design. Results: The net effect of having an obese mother varied by race/ethnicity and was associated with a significantly earlier age at obesity onset (p = 0.0001) for whites [β= ?8.1 year, 95% confidence interval (CI), ?9.3; ?6.9)], blacks (β = ?10.8 years, 95% CI, ?12.4; ?9.2), Hispanics (β = ?7.0 years, 95% CI, ?9.2; ?4.8), and Asians (β = ?8.6 years, 95% CI, ?13.3; ?3.9). Earlier obesity onset (<18 years) was associated with increased severity at young adulthood (mean BMI, 36.0 ± 0.3 kg/m²) vs. onset after age 18 (mean BMI, 34.4 ± 0.2 kg/m²; p = 0.0001). There were no sex differences in the association of maternal obesity to age at obesity onset. Conclusions: Having an obese mother was associated with earlier age at obesity onset across all race/ethnic groups, particularly non‐Hispanic blacks. Early obesity onset has important health consequences because of its association with more severe adult obesity.     

Mechanistic studies on the intramolecular one-electron transfer between the two flavins in the human endothelial NOS reductase domain

The object of this study was to clarify the mechanism of electron transfer in the human endothelial nitric oxide synthase (eNOS) reductase domain using recombinant eNOS reductase domains; the FAD/NADPH domain containing FAD- and NADPH-binding sites and the FAD/FMN domain containing FAD/NADPH-, FMN-, and a calmodulin-binding sites. In the presence of molecular oxygen or menadione, the reduced FAD/NADPH domain is oxidized via the neutral (blue) semiquinone (FADH(*)), which has a characteristic absorption peak at 520 nm. The FAD/NADPH and FAD/FMN domains have high activity for ferricyanide, but the FAD/FMN domain has low activity for cytochrome c. In the presence or absence of calcium/calmodulin (Ca(2+)/CaM), reduction of the oxidized flavins (FAD-FMN) and air-stable semiquinone (FAD-FMNH(*)) with NADPH occurred in at least two phases in the absorbance change at 457nm. In the presence of Ca(2+)/CaM, the reduction rate of both phases was significantly increased. In contrast, an absorbance change at 596nm gradually increased in two phases, but the rate of the fast phase was decreased by approximately 50% of that in the presence of Ca(2+)/CaM. The air-stable semiquinone form was rapidly reduced by NADPH, but a significant absorbance change at 520 nm was not observed. These findings indicate that the conversion of FADH(2)-FMNH(*) to FADH(*)-FMNH(2) is unfavorable. Reduction of the FAD moiety is activated by CaM, but the formation rate of the active intermediate, FADH(*)-FMNH(2) is extremely low. These events could cause a lowering of enzyme activity in the catalytic cycle.     

Analysis of ALS5 and ALS6 allelic variability in a geographically diverse collection of Candida albicans isolates

The Candida albicans ALS (agglutinin-like sequence) gene family encodes eight cell-surface glycoproteins, some of which function in adhesion to host surfaces. ALS genes have a central tandem repeat-encoding domain comprised entirely of head-to-tail copies of a conserved 108-bp sequence. The number of copies of the tandemly repeated sequence varies between C. albicans strains and often between alleles within the same strain. Because ALS alleles can encode different-sized proteins that may have different functional characteristics, defining the range of allelic variability is important. Genomic DNA from C. albicans strains representing the major genetic clades was PCR amplified to determine the number of tandemly repeated sequence copies within the ALS5 and ALS6 central domain. ALS5 alleles had 2-10 tandem repeat sequence copies (mean=4.82 copies) while ALS6 alleles had 2-8 copies (mean=4.00 copies). Despite this variability, tandem repeat copy number was stable in C. albicans strains passaged for 3000 generations. Prevalent alleles and allelic distributions varied among the clades for ALS5 and ALS6. Overall, ALS6 exhibited less variability than ALS5. ALS5 deletions can occur naturally in C. albicans via direct repeats flanking the ALS5 locus. Deletion of both ALS5 alleles was associated particularly with clades III and SA. ALS5 exhibited allelic polymorphisms in the coding region 5' of the tandem repeats; some alleles resembled ALS1, suggesting recombination between these contiguous loci. Natural deletion of ALS5 and the sequence variation within its coding region suggest relaxed selective pressure on this locus, and that Als5p function may be dispensable in C. albicans or redundant within the Als family.     

EGFR and beta1 integrins utilize different signaling pathways to activate Akt

Akt, also called PKB, is a serine/threonine kinase that plays a major role in cell survival. It can be activated by several cellular receptors, including integrins and growth factor receptors, in PI3K-dependent manners. In this study, we analyzed the two current models for Akt activation upon beta1 integrin-mediated adhesion: via focal adhesion kinase and via transactivation of the EGF receptor. Distinct differences in the pathways leading to phosphorylation and activation of Akt from stimulated beta1 integrins and EGF receptor were observed, including opposing sensitivity to the tyrosine kinase inhibitors PP2 and Gefitinib. Using knockout cells and integrin mutant cells, we show that beta1 integrins can induce phosphorylation of Akt at Ser473 and Thr308 and Akt kinase activity independently of the EGF receptor activity, focal adhesion kinase, and the Src family members. In contrast to stimulation with EGF, beta1 integrin-mediated adhesion did not induce Akt tyrosine phosphorylation. Moreover, tyrosine phosphorylation of Akt was found not to be required for its catalytic activity. The results identify a previously unrecognized mechanism by which beta1 integrins activate the PI3K/Akt pathway.     

The chalcone synthase multigene family of Petunia hybrida (V30): sequence homology,chromosomal localization and evolutionary aspects

Chalcone synthase (CHS) genes in Petunia hybrida comprise a multigene family containing at least 7 complete members in the strain Violet 30 (V30). Based on a high sequence homology in both coding and non-coding sequence, a number of CHS genes can be placed into two subfamilies. By restriction fragment length polymorphism (RFLP) analysis it was shown that both chromosomes II and V carry one of these subfamilies, in addition to the other CHS genes identified so far. Members of a subfamily were found to be closely linked genetically. Analysis of the Petunia species that contributed to the hybrid nature of P. hybrida (P. axillaris, P. parodii, P. inflata and P. violacea) shows that none of the CHS gene clusters is specific for either one of the parents and therefore did not arise as a consequence of the hybridization. The number of CHS genes within a subfamily varies considerably among these Petunia species. From this we infer that the CHS subfamilies arose from very recent gene duplications.     

Kindlins: essential regulators of integrin signalling and cell-matrix adhesion

Integrin-mediated cell-ECM (extracellular matrix) adhesion is a fundamental process that controls cell behaviour. For correct cell-ECM adhesion, both the ligand-binding affinity and the spatial organization of integrins must be precisely controlled; how integrins are regulated, however, is not completely understood. Kindlins constitute a family of evolutionarily conserved cytoplasmic components of cell-ECM adhesions that bind to beta-integrin cytoplasmic tails directly and cooperate with talin in integrin activation. In addition, kindlins interact with many components of cell-ECM adhesions--such as migfilin and integrin-linked kinase--to promote cytoskeletal reorganization. Loss of kindlins causes severe defects in integrin signalling, cell-ECM adhesion and cytoskeletal organization, resulting in early embryonic lethality (kindlin-2), postnatal lethality (kindlin-3) and Kindler syndrome (kindlin-1). It is therefore clear that kindlins, together with several other integrin-proximal proteins, are essential for integrin signalling and cell-ECM adhesion regulation.     

受体相互作用蛋白质激酶3通过上调整合素β3促进骨关节炎相关病变

骨关节炎（osteoarthritis,OA）是最常见的慢性致残性关节疾患,目前尚无针对病因的有效治疗手段。程序性坏死在多种疾病中扮演关键角色,受体相互作用蛋白质激酶3（receptor-interacting protein kinase 3, RIP3）是程序性坏死进程的关键调控因子。有研究显示,RIP3在人与鼠骨关节炎退变软骨组织中表达水平显著上调,提示程序性坏死的发生,但RIP3在软骨中的具体病生理角色仍不明确。本研究拟对过表达RIP3前后的软骨细胞转录物组进行测序分析,探索RIP3在骨关节炎进程中发挥作用的具体机制。RNA测序结果显示,RIP3的过表达诱发软骨细胞中244个基因表达上调,277个表达下调。通过进一步构建基因间共表达作用网络,筛选出16个候选靶基因在mRNA水平进行验证,证实RIP3对磷脂酰肌醇3激酶调节亚单位5（phosphoinositide-3-kinase, regulatory subunit 5,Pik3r5）、整合素β3（integrin subunit beta 3,Itgb3）及成髓细胞瘤转录因子第2亚型（MYB proto-oncogene like 2,Mybl2）的表达上调作用最为显著。CCK-8以及乳酸脱氢酶活性检测结果表明,利用siRNA沉默Itgb3的表达可显著抑制RIP3诱发的软骨细胞活力下降及程序性坏死,同时也抑制了RIP3对软骨细胞中分解代谢相关基因Mmp1、Mmp13与Il6的表达上调作用,以及其对合成代谢相关基因Acan、Col2a1与Sox9的下调作用。本研究证实,RIP3通过上调软骨细胞中Itgb3的表达诱发软骨细胞坏死与软骨基质代谢紊乱,并最终导致软骨退变,为骨关节炎的临床治疗提供了新靶点,同时进一步明确了程序性坏死的病理生理学意义。