Plasma membrane associated membranes (PAM) from Jurkat cells contain STIM1 protein: Is PAM involved in the capacitative calcium entry?

A proper cooperation between the plasma membrane, the endoplasmic reticulum and the mitochondria seems to be essential for numerous cellular processes involved in Ca²⁺ signalling and maintenance of Ca²⁺ homeostasis. A presence of microsomal and mitochondrial proteins together with those characteristic for the plasma membrane in the fraction of the plasma membrane associated membranes (PAM) indicates a formation of stabile interactions between these three structures. We isolated the plasma membrane associated membranes from Jurkat cells and found its significant enrichment in the plasma membrane markers including plasma membrane Ca²⁺-ATPase, Na⁺, K⁺-ATPase and CD3 as well as sarco/endoplasmic reticulum Ca²⁺ ATPase as a marker of the endoplasmic reticulum membranes. In addition, two proteins involved in the store-operated Ca²⁺ entry, Orai1 located in the plasma membrane and an endoplasmic reticulum protein STIM1 were found in this fraction. Furthermore, we observed a rearrangement of STIM1-containing protein complexes isolated from Jurkat cells undergoing stimulation by thapsigargin. We suggest that the inter-membrane compartment composed of the plasma membrane and the endoplasmic reticulum, and isolated as a stabile plasma membrane associated membranes fraction, might be involved in the store-operated Ca²⁺ entry, and their formation and rebuilding have an important regulatory role in cellular Ca²⁺ homeostasis.     

Phosphoglycerate mutase in mammalian striated muscles: Subcellular localization and binding partners

Contrary to previously published data, we have found that in mammalian skeletal muscles, phosphoglycerate mutase (PGM) is organized in a regular, striated fashion within the sarcomere. In the absence of the enzyme effectors, PGM localizes mainly at the M-line, but under conditions typical for contracting muscle, the enzyme accumulates within the I-band of the sarcomere. Searching for muscle PGM binding partners, we have found that PGM interacts with several enzymes of triose phosphate metabolism. It might suggest that PGM is a central structural element of the muscle glycolytic complex located within the isotropic region of the sarcomere.

Structured summary

MINT-7034028: PGM (uniprotkb:P16290) physically interacts (MI:0218) with lactate dehydrogenase B (uniprotkb:P42123), lactate dehydrogenase A (uniprotkb:P04642), G3PDH (uniprotkb:P04797), aldolase (uniprotkb:P05065), Creatine kinase (uniprotkb:P07335), phPhosphoglycerate kinase (uniprotkb:P16617) and Enolase (uniprotkb:P04764) by pull down (MI:0096)     

Genome balance in six successive generations of the allotetraploid Festuca pratensis × Lolium perenne

In the allotetraploid, Festuca pratensis Huds. (2n = 4x = 28) × Lolium perenne L. (2n = 4x = 28) the balance of chromatin, as determined by GISH, changes over successive generations of open pollination in favour of L. perenne. There is extensive recombination between chromosomes of the two parental genomes, as well as substitution of whole Festuca chromosomes by whole Lolium chromosomes. The total number of Lolium chromosomes increased from a mean 14.36 in the F₂ to 16.26 in the F₆, and the total number of Festuca chromosomes decreased correspondingly from a mean of 13.57 to a value of 11.56. The number of recombinant chromosomes and recombination breakpoints per genotype also increased from generation to generation, although the respective values of both characters were higher for Festuca (0.86–8.41 and 1.14–15.22) than for Lolium (0.68–4.59 and 0.68–6.0). The proportion of total genome length contributed by the L. perenne chromatin increased from about 50% in F₂ to 59.5% in F₆. The results are based on the sample of 134 plants studied (26–28 plants per generation), and are discussed in terms of the dominance of Lolium chromosomes over those of Festuca, and possible mechanisms underlying this phenomenon of chromatin substitution.     

Stiffness of normal and pathological erythrocytes studied by means of atomic force microscopy

During recent years, atomic force microscopy has become a powerful technique for studying the mechanical properties (such as stiffness, viscoelasticity, hardness and adhesion) of various biological materials. The unique combination of high-resolution imaging and operation in physiological environment made it useful in investigations of cell properties. In this work, the microscope was applied to measure the stiffness of human red blood cells (erythrocytes). Erythrocytes were attached to the poly-L-lysine-coated glass surface by fixation using 0.5% glutaraldehyde for 1 min. Different erythrocyte samples were studied: erythrocytes from patients with hemolytic anemias such as hereditary spherocytosis and glucose-6-phosphate-dehydrogenase deficiency patients with thalassemia, and patients with anisocytosis of various causes. The determined Young's modulus was compared with that obtained from measurements of erythrocytes from healthy subjects. The results showed that the Young's modulus of pathological erythrocytes was higher than in normal cells. Observed differences indicate possible changes in the organization of cell cytoskeleton associated with various diseases.     

Light and scanning electron microscopic study of the tongue in the cormorant Phalacrocorax carbo (Phalacrocoracidae, Aves)

The tongue of the cormorant Phalacrocorax carbo is a small, immobile structure with a length of 1.4 cm, situated in the middle part of the elongated lower bill. The uniquely shaped tongue resembles a mushroom, with a short base and an elongated dorsal part with sharpened anterior and posterior tips. A median crest can be observed on the surface of the tongue. Examination by light and scanning electron microscopy revealed that the whole tongue is formed by a dense connective tissue with many bundles of elastic fibers. The lingual mucosa is covered by a multilayered keratinized epithelium. The thickest, horny layer of the lingual epithelium was observed on the surface of the median crest and on the posterior tip of the tongue. Lingual glands are absent in cormorants. The framework of the tongue is composed of a hyoid cartilage incorporated into the base. The localization and structure of the tongue in the cormorant show that it is a rudimentary organ and that the lingual body, usually well-developed in birds, is conserved.     

Endosomal hyperacidification in cystic fibrosis is due to defective nitric oxide-cylic GMP signalling cascade

Endosomal hyperacidification in cystic fibrosis (CF) respiratory epithelial cells is secondary to a loss of sodium transport control owing to a defective form of the CF transmembrane conductance regulator CFTR. Here, we show that endosomal hyperacidification can be corrected by activating the signalling cascade controlling sodium channels through cyclic GMP. Nitric oxide (NO) donors corrected the endosomal hyperacidification in CF cells. Stimulation of CF cells with guanylate cyclase agonists corrected the pH in endosomes. Exposure of CF cells to an inhibitor of cGMP-specific phosphodiesterase PDE5, Sildenafil, normalized the endosomal pH. Treatment with Sildenafil reduced secretion by CF cells of the proinflammatory chemokine interleukin 8 following stimulation with Pseudomonas aeruginosa products. Thus, the endosomal hyperacidification and excessive proinflammatory response in CF are in part due to deficiencies in NO- and cGMP-regulated processes and can be pharmacologically reversed using PDE5 inhibitors.     

The level of cotinine - marker of tobacco smoking, in patients with hyperthyroidism

INTRODUCTION: Orbitopathy associated with Graves' disease TAO (thyroid associated orbitopathy) is likewise connected with environmental factors including tobacco smoking. Tobacco smoking increases the risk of ophthalmologic pathology and leads to the orbital tissue damage due to the hypoxia. The aim of the study was to correlate the concentration of urinecotinine (marker of tobacco smoking) in Graves' disease patients with TAO with points received on the Fagerstr?m questionnaire. MATERIALS AND METHODS: Thirty women with different degree of exacerbation of TAO in Graves' disease (34.28 +/- 12.04 yr.) were examined. As control we used 29 women with Graves' disease without TAO (29.35 +/- 12.33 yr.). The diagnosis was established according to level of TSH and level of thyrotropin receptor antibodies (TRAb). Measure of tobacco smoking or exposure to second hand tobacco smoke ETS (environmental tobacco smoke) carried out according to the Fagerstr?m questionnaire, and measured cotinine level (the major metabolite of nicotine in urine). The level of clinical ophtalmopathy was measured according to the CAS (Clinical Activity Score) scale and degree of progression of TAO according to American Thyroid Association (NOSPECS scale). Additionally increase of exophthalmus was measured using the Hertl's exophthalmometer. RESULTS: In the group with TAO: 19 (63.3%) persons had mild exophthalmos (according to CAS), medium in 13 (43.3%) patients, and pronounced exophthalmous in 10 (33%) patients. There was statistically significant difference in the level of TRAb (18.4 +/- 8.2 vs. 5.1 +/- 3.4 IU/l; p < 0.0001) between TAO and controls. There was no correlation between TSH (0.6 +/- 0.02 vs. 0.4 +/- 0.04; p = 0.18) and fT4 (38.8 +/- 29.3 vs. 26.1 +/- 17.3; p = 0.026) in both analyzed groups. Smokers and non-smokers with TAO had no statistically significant in level of TRAb (Mann-Whitney test p = 0.16). No correlation was found between smoking tobacco (cotinine level) and the level of TRAb in patients with TAO (Pearson r = 0.28 p = 0.58). There was a statistically significant difference between the level of urine cotinine in smoking patients with TAO in the highest level of ophthalmopathy exacerbation (> 24 mm; > or = 4 points according to CAS) and those without TAO (Mann-Whitney's test p = 0.04). CONCLUSIONS: 1. In patients with Graves' disease with TAO the highest score of tobacco smoking has been found in persons with the highest ophthalmopathy exacerbation. 2. Estimation of cotinine concentration in urine is the most objective and useful method of tobacco smoking.     

Role of intraluteal prostaglandin F(2alpha), progesterone and oxytocin in basal and pulsatile progesterone release from developing bovine corpus luteum

The present study examined the role of intra-luteal prostaglandin (PG) F(2alpha), progesterone (P4) and oxytocin (OT) on the corpus luteum function by using specific hormone antagonists. Luteal cells from the developing CL (days 5-7 of the estrous cycle) were exposed to P4 antagonist (onapristone, OP, 10(-4)M), OT antagonist (atosiban, AT; 10(-6)M) or indomethacin (INDO; 10(-4)M), for 12h and then stimulated with PGF(2alpha) (10(-8)M) for 4h. Pre-treatment of the cells with OP, AT or INDO resulted in an increase in P4 secretion in response to PGF(2alpha). To examine the temporal effects of P4, OT and PGs on P4 secretion, dispersed luteal cells were pre-exposed to OP, AT or INDO for 1, 2, 4, 6 or 12h. Prostaglandin F(2alpha) stimulated P4 secretion (P<0.05) after 2h of pre-exposition. In the microdyalisis study, the spontaneous release of P4 from developing CL tissue was of pulsatile nature with irregular peaks at 1-2h intervals. Treatment with OP increased the number of P4 peaks (P<0.05), whereas AT and INDO significantly reduced the number of P4 peaks detected (P<0.05). Interestingly, INDO completely blocked the pulsatile nature in the release of P4, but it secretion remained stable throughout the experimental period. These results demonstrate that luteal PGF(2alpha), OT, and P4 are components of an autocrine/paracrine intra-ovarian regulatory system responsible for the episodic (pulsatile) release of P4 from the bovine CL during the early luteal phase.