Annexins--proteins involved in organization and function of biological membranes--from Arabidopsis thaliana to Homo sapiens

The mini-review series presented in this issue of Postepy Biochemii is focussed on some aspects of biology of calcium- and membrane-binding proteins, annexins, ubiquitous in all eucaryotic organisms (excluding yeasts), from Arabidopsis thaliana to Homo sapiens. Annexins are encoded by twelve genes in verterbrates and by eight in higher plants. Their physiological significance is underlined by two facts: the numer of the annexin genes seems to grow during evolution and in some cell types they comprise up to 2% of total protein. In the present review the hypothesis is discussed suggesting that multiplication of annexin genes in evolution represents mechanism of organism adaptation to changes in environment. In addition, the experimental data are presented suggestive of annexins playing a crucial role in functioning of plasma membrane, such as signal transduction, ion and vesicular transport and membrane repair. The review is then followed by articlesdealing in details with participation of annexins in plant response to abiotic stress (Arabidopsis thaliana), in tissue mineralization (Gallus gallus), in exocytosis of catecholamines by PC12 cells (mammals) and in Niemann-Pick type C disease related to abnormal transport and intracellular storage of cholesterol (Homo sapiens).     

Annexins in Niemann-Pick type C disease

Niemann-Pick disease is a genetic disorder, affecting approximately 1 to 150,000 living births per year; in Poland 1-5 cases. Usually diagnosed in the childhood, Niemann-Pick disease results in death in the teenage years. Niemann-Pick disease is defined as a lysosomal storage disorder and is related to impaired transport and/or accumulation of specific lipids inside the cell. In this report, we provide evidence about potential role of annexins, calcium- and membrane-binding proteins, in the formation and stabilization of cholesterol-rich microdomains and their possible function in organizing the membranes of early and late endosomes, organelles affected in the type C Niemann-Pick disease characterized by abnormal accumulation of cholesterol and glycosphingolipids in lysosomal like organelles.     

Equol producer status, salivary estradiol profile and urinary excretion of isoflavones in Irish Caucasian women, following ingestion of soymilk

Equol production, isoflavone excretion, and the salivary estradiol profile among 36 females, native Irish Caucasian volunteers following ingestion of 200mL soymilk is reported. The soymilk contained daidzein (73+/-6.7mg) and genistein (86+/-10.2mg). Volunteers provided personal and family medical history. Dietary analysis revealed that all volunteers regularly consumed soy-based or soy-supplemented food products. The mean age, mean age at menarche, and body mass index of volunteers were 46.6+/-12.3 years, 13.1 years and 26.1, respectively. The average number of children per volunteer was 2.13. Twelve (34%) of the volunteers were found to be first-degree relatives of breast cancer patients. Following consumption of the soymilk, equol was detected in the urine of 18 (51%) of the volunteers. Mean urinary daidzein and genistein concentrations during the hours following soymilk ingestion were 13.5 and 16.7microg/mg creatinine, respectively, however, some volunteers excreted little (less than 4.0microg/mg) or no isoflavone. Salivary estradiol in most (24) volunteers had decreased from 51.5+/-28.67pmol/L pre-ingestion to 29.75+/-16.13pmol/L 5h after drinking the soymilk. However, the salivary estradiol in 12 subjects (34%) increased from 33.76+/-13.4pmol/L to 137.4+/-65.64pmol/L over the same period. Individuals whose salivary estradiol increased had significantly less children (1.58 (P<0.05)), were more likely to (a) return urine samples with low isoflavone content (50.3% compared to 25%), (b) to be equol producers (67% compared to 41.7%), and (c) to be first-degree relatives of breast cancer patients (41.7% compared to 25%). Volunteers who reported a first-degree link to breast cancer were more likely to have a higher body mass index (29.0 compared to 26.1 (P<0.05)), to be equol producers (75% compared to 51%), and to excrete isoflavones in low quantities only (60% compared to 50%). First-degree relatives also had fewer children (1.75 (P<0.05)). The results indicate a significant, distinctive variation in equol production, isoflavone excretion and salivary estradiol profile among individual volunteers following ingestion of soymilk.     

Trypanosoma cruzi prevalence and epidemiologic trends in lemurs on St. Catherines Island, Georgia

Lemurs on St. Catherines Island, Georgia were tested for Trypanosoma cruzi infection to develop a better understanding of the epizootiology of the parasite in nonhuman primates in the southeastern United States. Fifty-six ring-tailed (Lemur catta), blue-eyed black (Eulemur macaco flavifrons), and black-and-white ruffed (Varecia variegata variegata) lemurs were tested by hemoculture and serology to determine the prevalence of T. cruzi in the population. Of those tested 3 (5%) were identified as culture positive and 25 (44.6%) as seropositive. When hemoculture results were compared with those from a similar study performed in 1997, prevalence remained unchanged. Genetic characterization of the 3 culture isolates indicated they belong to the T. cruzi IIa group, which is identical to strains previously isolated from raccoons on the island. Despite the occurrence of T. cruzi in the population, there was no evidence that the health of the lemurs was compromised as a result of infection. Based upon prevalence and available breeding records we speculate that both vertical and vector-mediated transmission play significant roles in the epidemiology of T. cruzi on the island. This also represents the first report of autochthonous infection in blue-eyed black and black-and-white ruffed lemurs.     

Application of a changing field of growth rates to a description of root apex formation

Based on the growth tensor method an unsteady field of growth rates for developing root apex is presented. Maps of growth rates distribution as well as simulations in which the field was applied to initially uniform grid of points are presented. In the simulations, the grid undergoes deformation that resembles new root formation in its axial plane. Four variants of field operation on the grid are shown.     

Protoporphyrin IX interacts with wild-type p53 protein in vitro and induces cell death of human colon cancer cells in a p53-dependent and -independent manner

Photodynamic therapy (PDT) of cancer is an alternative treatment for tumors resistant to chemo- and radiotherapy. It induces cancer cell death mainly through generation of reactive oxygen species by a laser light-activated photosensitizer. It has been suggested that the p53 tumor suppressor protein sensitizes some human cancer cells to PDT. However, there is still no direct evidence for this. We have demonstrated here for the first time that the photosensitizer protoporphyrin IX (PpIX) binds to p53 and disrupts the interaction between p53 tumor suppressor protein and its negative regulator HDM2 in vitro and in cells. Moreover, HCT116 colon cancer cells exhibited a p53-dependent sensitivity to PpIX in a dose-dependent manner, as was demonstrated using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay and fluorescence-activated cell sorter (FACS) analysis of cell cycle profiles. We have also observed induction of p53 target pro-apoptotic genes, e.g. puma (p53-up-regulated modulator of apoptosis), and bak in PpIX-treated cells. In addition, p53-independent growth suppression by PpIX was detected in p53-negative cells. PDT treatment (2 J/cm2) of HCT116 cells induced p53-dependent activation of pro-apoptotic gene expression followed by growth suppression and induction of apoptosis.     

Effects of a particle film on biology and behavior of Diaphorina citri (Hemiptera: Psyllidae) and its infestations in citrus

Studies were conducted to investigate the effects of a kaolin-based hydrophilic particle film, Surround WP, on the biology and behavior of the psyllid Diaphorina citri Kuwayama (Hemiptera: Psyllidae) and to assess population densities of D. citri in citrus subjected to monthly applications of Surround WP. Laboratory investigations indicated a 3% (wt:vol) suspension of Surround WP in water applied directly was not acutely toxic to eggs, older nymphs or adults. Presence of the dried particle film on leaves interfered with the ability of adults to grasp and walk on citrus leaves. During a 30-s period, adults spent an average of 5 s moving on leaves with particle film compared with 16 s on leaves without particle film. When leaves were inverted, a significantly higher percentage of adults fell or flew from treated leaves (53%) than untreated leaves (16%). In a 12-mo study investigating infestations of D. citri on citrus treated monthly with Surround WP, cumulative reductions of 78% in adult numbers on mature leaves and of 60% in adult numbers on flush shoots (immature leaves) were observed in treated trees compared with untreated trees. Numbers of eggs and nymphs per flush shoot were reduced by 85 and 78%, respectively, in trees treated with particle film. Reductions in infestation levels of D. citri in treated trees were attributed to the negative effects of the particle film on the ability of adults to grasp, move, and oviposit. The suppressive effects of a Surround treatment against adult psyllids were degraded by rain.     

The in vitro effect of pH on osteoclasts and bone resorption in the cat: implications for the pathogenesis of FORL

Dental disease due to osteoclast over-activity reaches epidemic proportions in older domestic cats and has also been reported in wild cats. Feline osteoclastic resorptive lesions (FORL) involve extensive resorption of the tooth leaving it liable to root fracture and subsequent tooth loss. The aetio-pathogenesis of FORL is not known. Recent work has shown that systemic acidosis causes increased osteoclast activation and that loci of infection or inflammation in cat mouth are likely to be acidotic. To investigate this, we generated osteoclasts from cat blood and found that they formed in large numbers (approximately 400) in cultures on bovine cortical bone slices. Acidosis caused an increase in the size of cells-in cultures maintained up to 14 days at basal pH 7.25, mean osteoclast area was 0.01 +/- 0.003 mm(2), whereas an 8.6-fold increase was observed in cells cultured between 11 and 14 days at pH 7.15 (0.086 +/- 0.004 mm(2)). Acidosis caused a modest increase in the number of osteoclasts. Exposure to pH 6.92 exhibited a 5-fold increase in the area of bone slices covered by resorption lacunae ( approximately 70% bone slice resorbed). In line with this finding, significant increases were observed in the expression of cathepsin K and proton pump enzymes (both approximately 3-fold) that are key enzymes reflective of resorptive activity in osteoclasts. These results demonstrate that acidosis is a major regulator of osteoclast formation and functional activation in the cat, and suggest that local pH changes may play a significant role in the pathogenesis of FORL.     

Zinc modulation of water permeability reveals that aquaporin 0 functions as a cooperative tetramer

We previously showed that the water permeability of AQP0, the water channel of the lens, increases with acid pH and that His40 is required (Németh-Cahalan, K.L., and J.E. Hall. 2000. J. Biol. Chem. 275:6777-6782; Németh-Cahalan, K.L., K. Kalman, and J.E. Hall. 2004. J. Gen. Physiol. 123:573-580). We have now investigated the effect of zinc (and other transition metals) on the water permeability of AQP0 expressed in Xenopus oocytes and determined the amino acid residues that facilitate zinc modulation. Zinc (1 mM) increased AQP0 water permeability by a factor of two and prevented any additional increase induced by acid pH. Zinc had no effect on water permeability of AQP1, AQP4 or MIPfun (AQP0 from killifish), or on mutants of AQP1 and MIPfun with added external histidines. Nickel, but not copper, had the same effect on AQP0 water permeability as zinc. A fit of the concentration dependence of the zinc effect to the Hill equation gives a coefficient greater than three, suggesting that binding of more than one zinc ion is necessary to enhance water permeability. His40 and His122 are necessary for zinc modulation of AQP0 water permeability, implying structural constraints for zinc binding and functional modulation. The change in water permeability was highly sensitive to a coinjected zinc-insensitive mutant and a single insensitive monomer completely abolished zinc modulation. Our results suggest a model in which positive cooperativity among subunits of the AQP0 tetramer is required for zinc modulation, implying that the tetramer is the functional unit. The results also offer the possibility of a pharmacological approach to manipulate the water permeability and transparency of the lens.     

Staphylococcus aureus-derived staphopain B, a potent cysteine protease activator of plasma chemerin

Chemerin is an attractant for cells that express the serpentine receptor CMKLR1, which include immature plasmacytoid dendritic cells (pDC) and macrophages. Chemerin circulates in the blood where it exhibits low biological activity, but upon proteolytic cleavage of its C terminus, it is converted to a potent chemoattractant. Enzymes that contribute to this conversion include host serine proteases of the coagulation, fibrinolytic, and inflammatory cascades, and it has been postulated that recruitment of pDC and macrophages by chemerin may serve to balance local tissue immune and inflammatory responses. In this work, we describe a potent, pathogen-derived proteolytic activity capable of chemerin activation. This activity is mediated by staphopain B (SspB), a cysteine protease secreted by Staphylococcus aureus. Chemerin activation is triggered by growth medium of clinical isolates of SspB-positive S. aureus, but not by that of a SspB(null) mutant. C-terminal processing by SspB generates a chemerin isoform identical with the active endogenous attractant isolated from human ascites fluid. Interestingly, SspB is a potent trigger of chemerin even in the presence of plasma inhibitors. SspB may help direct the recruitment of specialized host cells, including immunoregulatory pDC and/or macrophages, contributing to the ability of S. aureus to elicit and maintain a chronic inflammatory state.