SMARCA2 deficiency in NSCLC: a clinicopathologic and immunohistochemical analysis of a large series from a single institution

BACKGROUND@#SMARCA2 (SWI/SNF Related, Matrix Associated, Actin Dependent Regulator of Chromatin, Subfamily A, Member 2) is an important ATPase catalytic subunit in the switch-sucrose nonfermenting (SWI/SNF) complex. However, its relationship with the pathological features of NSCLC and its prognosis remain unclear.@*METHODS@#We retrospectively reviewed 2390 patients with surgically resected NSCLC, constructed tissue microarrays (TMAs) and performed immunohistochemical assays. We analyzed the correlation of SAMRCA2 with clinicopathological features and evaluated its prognostic value.@*RESULTS@#Among 2390 NSCLC cases, the negative expression ratios of SAMRCA2, SMARCA4, ARID1A, ARID1B and INI1 were 9.3%, 1.8%, 1.2%, 0.4% and 0%, respectively. In NSCLC, male sex, T3 and T4 stage, moderate and poor differentiation, tumor ≥ 2 cm, Ki67 ≥ 15%, SOX-2 negative expression, middle lobe lesion and adenocarcinoma were relative risk factors affecting SMARCA2-negative expression. In lung adenocarcinomas, high-grade nuclei, histological morphology of acinar and papillary, solid and micropapillary and TTF-1-negative expression were relative risk factors affecting SMARCA2-negative expression. Kaplan-Meier survival analysis showed that the OS was shorter in the SMARCA2-negative group. Multivariate survival analysis revealed that SMARCA2-negative expression was an independent factor correlated with a poor prognosis in NSCLC.@*CONCLUSION@#In conclusion, SMARCA2-negative expression is an independent predictor of a poor outcome of NSCLC and is a potential target for NSCLC treatment.

Advances in plant heavy metal transporter P1B-ATPases / 生物工程学报

P1B-ATPases are a group of proteins that can transport heavy metal ions across membranes by hydrolyzing ATP and they are a subclass of the P-type ATPase family. It was found that P1B-ATPases are mainly responsible for the active transport of heavy metal ions in plants and play an important role in the regulation of heavy metal homeostasis in plants. In this paper, we dissusses the mechanism of P1B-ATPases from the structure and classification of P1B-ATPases, and review the current research progress in the function of P1B-ATPases, in order to provide reference for future research and application of P1B-ATPases in improving crop quality and ecological environment management.

Prevalência da baixa densidade mineral óssea em mulheres na pós-menopausa tratadas de câncer de mama / Prevalence of low bone mineral density in postmenopausal breast cancer survivors

OBJETIVO: Avaliar a prevalência da baixa densidade mineral óssea (DMO) em mulheres na pós-menopausa tratadas de câncer de mama. MÉTODOS: Estudo de corte transversal que incluiu 115 mulheres tratadas de câncer de mama atendidas em Hospital Universitário do Sudeste do Brasil. Foram incluídas mulheres com amenorreia há 12 meses ou mais e 45 anos ou mais de idade, tratadas de câncer de mama e livres de doença há pelo menos 5 anos. A DMO foi mensurada pelos raios-X de dupla energia em coluna lombar (L1 a L4) e colo de fêmur. Considerou-se baixa DMO quando valores de T-score de coluna total e/ou colo de fêmur <-1,0 Score de Delphi (DP) (osteopenia e osteoporose). Por meio de entrevista, foram avaliados fatores de risco para baixa DMO. Na análise estatística, empregaram-se os testes do χ2 ou Exato de Fisher. RESULTADOS: A média de idade das pacientes foi 61,6±10,1 anos e o tempo de menopausa, 14,2±5,6 anos, com tempo médio de seguimento de 10,1±3,9 anos. Considerando coluna e colo de fêmur, 60% das mulheres tratadas de câncer de mama apresentavam baixa DMO. Avaliando os fatores de risco para baixa DMO, foi encontrada diferença significativa na distribuição percentual quanto à idade (maior porcentagem de mulheres com mais de 50 anos e baixa DMO), história pessoal de fratura prévia (11,6% com baixa DMO e nenhuma com DMO normal) e índice de massa corpórea. Maior frequência de obesidade foi observada entre mulheres com DMO normal (63%) quando comparadas àquelas com baixa DMO (26,1%; p<0,05). CONCLUSÃO: Mulheres na pós-menopausa tratadas de câncer de mama apresentaram elevada prevalência de baixa DMO (osteopenia e/ou osteoporose). .

PURPOSE: To evaluate the prevalence of low bone mineral density (BMD) in postmenopausal breast cancer survivors. METHODS: In this cross-sectional study, 115 breast cancer survivors, seeking healthcare at a University Hospital in Brazil, were evaluated. Eligibility criteria included women with amenorrhea ≥12 months and age ≥45 years, treated for breast cancer and metastasis-free for at least five years. BMD was measured by DEXA at the lumbar spine (L1-L4) and femoral neck. Low BMD was considered when total-spine and/or femoral-neck T-score values were <-1.0 Delphi Score (DP) (osteopenia and osteoporosis). The risk factors for low BMD were assessed by interview. Data were analyzed statistically by the χ2 test and Fisher's exact test. RESULTS: The mean age of breast cancer survivors was 61.6±10.1 years and time since menopause was 14.2±5.6 years, with a mean follow-up of 10.1±3.9 years. Considering spine and femoral neck, 60% of breast cancer survivors had low BMD. By evaluating the risk factors for low BMD, a significant difference was found in the percent distribution for age (higher % of women >50 years with low BMD), personal history of previous fracture (11.6% with low BMD versus 0% with normal BMD) and BMI. A higher frequency of obesity was observed among women with normal BMD (63%) compared to those with low BMD (26.1%) (p<0.05). CONCLUSION: Postmenopausal breast cancer survivors had a high prevalence of osteopenia and osteoporosis. .

Mitochondrial ATPase activity and membrane fluidity changes in rat liver in response to intoxication with Buckthorn (Karwinskia humboldtiana)

BACKGROUND: Karwinskia humboldtiana (Kh) is a poisonous plant of the rhamnacea family. To elucidate some of the subcellular effects of Kh toxicity, membrane fluidity and ATPase activities as hydrolytic and as proton-pumping activity were assessed in rat liver submitochondrial particles. Rats were randomly assigned into control non-treated group and groups that received 1,1.5 and 2 g/Kg body weight of dry powder of Kh fruit, respectively. Rats were euthanized at day 1 and 7 after treatment. RESULTS: Rats under Kh treatment at all dose levels tested, does not developed any neurologic symptoms. However, we detected alterations in membrane fluidity and ATPase activity. Lower dose of Kh on day 1 after treatment induced higher mitochondrial membrane fluidity than control group. This change was strongly correlated with increased ATPase activity and pH gradient driven by ATP hydrolysis. On the other hand, membrane fluidity was hardly affected on day 7 after treatment with Kh. Surprisingly, the pH gradient driven by ATPase activity was significantly higher than controls despite an diminution of the hydrolytic activity of ATPase. CONCLUSIONS: The changes in ATPase activity and pH gradient driven by ATPase activity suggest an adaptive condition whereby the fluidity of the membrane is altered.

Protective effect of some amino acids synthesized derivatives and their chelates on Escherichia coli under X-ray irradiation.

The protective effects of novel synthesized derivatives of some amino acids — nicotinyl-L-tyrosinate and nicotinyl-L-tryptophanate schiff bases and their Cu(II) and Mn(II) chelates on growth, survival and membrane-associated ATPase activity of E. coli under X-ray irradiation were investigated. The specific growth rate and survival of E. coli were decreased at 10, 20 and 30 Gy doses. However, as 30 Gy was found to be the most effective irradiation dose, it was chosen for studying the radio-protective properties of different compounds. These compounds could increase the bacterial cell protection against X-ray irradiation in concentration-dependent manner. They had a role in stimulation of synthesis or regulation of activity of metal-dependent enzymes, required for reversing the X-ray irradiation damage. The study may prove useful for further estimation of the effectiveness of different compounds as radio-protectors on bacteria and other cells, especially mammalian cells under X-ray irradiation.

Effects of low concentrations of a polychlorinated biphenyl, Aroclor 1254 on membrane bound ion dependent ATPases in mice liver.

Aroclor 1254, a polychlorinated biphenyl, is present in the environment in low concentration but references on its toxic effects on liver cell membrane proteins and the mechanism of actions are not abundantly available. Therefore, the present study was undertaken to investigate the low level, sub-acute dose and exposure duration dependent effects of Aroclor 1254 on total, Na+, K+, Ca2+ and Mg2+-ATPases of the mouse liver. The hypotheses tested in the present study were, (a) whether the low, environmentally available dose and the exposure durations of Aroclor 1254 affects the membrane-bound ion dependent ATPases, and (b) if a response was observed, whether it is a direct or indirect effects of the toxicant. Groups of mice were exposed to different doses (0.1 and 1mg kg-1 body weight d-1) and exposure durations (4 d, 8 d and 12 d) of Aroclor 1254. The results indicated significant exposure duration dependent changes in the specific activity of the selected membrane bound ATPases. As the observed changes were mostly enzyme stimulation after toxication through oral administration, the effects of the Aroclor were possibly indirect, through complex chain of reactions.

The role of micro RNAs let7c, 100 and 218 expression and their target RAS, C-MYC, BUB1, RB, SMARCA5, LAMB3 and Ki-67 in prostate cancer

OBJECTIVE: The aim of this study is to verify the expression of proteins that are controlled by miR-let7c, 100 and 218 using immunohistochemistry in tissue microarray representative of localized and metastasized the lymph nodes and bone prostate cancer. METHODS: To verify the expression of proteins that are controlled by miR-let7c (C-MYC, BUB1, RAS) 100 (SMARCA5, RB) and 218 (LAMB3) and cell proliferation (Ki-67) we used immunohistochemistry and computerized image system ImageJ MacBiophotonics in three tissue microarrays representative of localized prostate cancer and lymph node and bone metastases. miRNA expression was evaluated by qRT-PCR using 60 paraffin blocks to construct the tissue microarray representative of localized disease. RESULTS: RAS expression was increased in localized prostate cancer and bone metastases compared to the lymph nodes (p=0.017). RB showed an increase in expression from localized prostate cancer to lymph node and bone metastasis (p=0.036). LAMB3 was highly expressed in localized and lymph node metastases (p<0.001). Cell proliferation evaluated by Ki-67 showed an increase from localized prostate cancer to metastases (p<0.001). We did not found any relationship between C-MYC (p=0.253), BUB1 (p=0.649) and SMARCA5 (p=0.315) protein expression with prognosis or tumor behavior. CONCLUSION: We found that the expression of RAS, RB, LAMB3 and Ki-67 changed in the different stages of prostate cancer. Furthermore, we confirmed the overexpression of the miRNAs let7c, 100 and 218 in localized prostate cancer but failed to show the control of protein expression by the putative controller miRNAs using immunohistochemistry. .

Mismatch repair genes expression defects & association with clinicopathological characteristics in colorectal carcinoma.

Background & objectives: DNA mismatch repair gene (MMR) abnormalities are seen in 95 per cent of hereditary nonpolyposis colorectal cancer (HNPCC) and 10-15 per cent of sporadic colorectal cancers. There are no data on MMR abnormalities in Malaysian colorectal cancer patients. This study was aimed to determine the frequency of abnormal MMR gene protein expression in colorectal carcinoma in Northern Peninsular Malaysia using immunohistochemistry. Methods: Clinicopathological information was obtained from 148 patients’ records who underwent bowel resection for colorectal cancer (CRC) at the three hospitals in Malaysia. Immunohistochemistry for MLH1, MSH2, MSH6 and PMS2 proteins were performed on paraffin embedded tissue containing carcinoma. Results: A total of 148 subjects and 150 colorectal carcinomas of sporadic and hereditary types were assessed. Three patients had synchronous tumours. Twenty eight cancers (18.6%) from 26 subjects (17.6%) had absent immunohistochemical expression of any one of the MMR gene proteins. This comprised absent MLH1 only – 3 cancers, absent MSH2 only – 3, absent MSH6 only – 2, absent PMS2 only – 3, absent MLH1 and PMS2 – 14, absent MSH2 and MSH6 – 2 and absent MLH1, MSH6 and PMS2 – 1. There was significant association between abnormal MMR gene protein expression and proximal colon cancers, mucinous, signet ring and poorly differentiated morphology. Interpretation & conclusions: Cancers with abnormal MMR gene expression were associated with microsatellite instability-high (MSI-H) phenotype. About 15 per cent demonstrated absent MSH2, MSH6 and PMS2 protein expression in isolation or in combination with other MMR genes, which often predicts a germline mutation, synonymous with a diagnosis of HNPCC. This appears to be high frequency compared to reported data.

Lack of evidence for regulation of cardiac P-type ATPases and MAP kinases in transgenic mice with cardiac-specific overexpression of constitutively active á1B-adrenoceptors

The regulatory function of á1B-adrenoceptors in mammalian heart homeostasis is controversial. The objective of the present study was to characterize the expression/activity of key proteins implicated in cardiac calcium handling (Na+/K+-ATPase and Ca2+-ATPases) and growth (ERK1/2, JNK1/2 and p38) in mice with cardiac-selective overexpression of constitutively active mutant á1B-adrenoceptor (CAMá1B-AR), which present a mild cardiac hypertrophy phenotype. Immunoblot assays showed that myocardial plasma membrane Ca2+-ATPase (PMCA) expression was increased by 30 percent in CAMá1B-AR mice (N = 6, P < 0.05), although there was no change in sarco/endoplasmic reticulum Ca2+-ATPase (SERCA2) expression. Moreover, total Ca2+-ATPase activity was not modified, but a significant increase in the activity of the thapsigargin-resistant (PMCA) to thapsigargin-sensitive (SERCA) ratio was detected. Neither Na+/K+-ATPase activity nor the expression of á1 and á2 subunit isoforms was changed in CAMá1B-AR mouse hearts. Moreover, immunoblot assays did not provide evidence for an enhanced activation of the three mitogen-activated protein kinases studied in this stage of hypertrophy. Therefore, these findings indicate that chronic cardiac á1B-AR activation in vivo led to mild hypertrophy devoid of significant signs of adaptive modifications concerning primary intracellular calcium control and growth-related proteins, suggesting a minor pathophysiological role of this adrenergic receptor in mouse heart at this stage of development.

Effects of cocaine on activities of ATPase, LDH and SDH in mouse splenocytes / 法医学杂志

OBJECTIVE@#To examine the effects of cocaine on the activities of ATPase, LDH and SDH in cultured mouse splenocytes in vitro.@*METHODS@#The ATPase, LDH and SDH activities in mouse splenocytes were detected at day 7 after continuous culturing the mouse cells exposed to cocaine hydrochloride in final concentration of 10, 20 and 100 microg/mL in vitro.@*RESULTS@#The activities of ATPase, LDH and SDH in mouse splenocytes exposed to cocaine hydrochloride in final concentration of 10, 20 and 100 microg/mL were significantly decreased after continuous culturing for 7 days.@*CONCLUSION@#The present study demonstrated that cocaine could inhibit the activities of ATPase, LDH and SDH in cultured splenocytes in vitro.

Apoptose, proliferação e histomorfometria do baço de ratas adultas com hipofunção tireoidiana e ovariana / Apoptosis, proliferation and spleen histomorphometry of adult female rats with thyroid and ovarian hypofunction

Apoptose, proliferação e histomorfometria do baço foram investigados em ratas Wistar adultas ovariectomizadas e não-ovariectomizadas, mantidas em hipotireoidismo induzido pela administração diária de propiltiouracil (PTU) por 120 dias. Dois grupos eutireóideos ovariectomizados e não-ovariectomizados serviram como controle. Foi colhido o plasma para dosagem de T4 livre e o baço para análise da histomorfometria, do índice apoptótico e da expressão imunohistoquímica de caspase 3 e CDC47. Valores de T4 livre foram menores nas ratas tratadas com PTU (p < 0,05). Nos grupos hipotireóideos houve redução do peso do baço, do número e do tamanho dos folículos linfóides e aumento do índice apoptótico e da expressão de caspase 3 (p < 0,05). Porém, o baço de ratas hipotireóideas ovariectomizadas apresentou aumento menos acentuado do índice apoptótico e da expressão de caspase 3 do que o baço de ratas hipotireóideas não-ovariectomizadas (p < 0,05). O grupo eutireóideo ovariectomizado apresentou hiperplasia da polpa branca em relação ao grupo eutireóideo não-ovariectomizado. Não houve diferença na expressão de CDC47 entre os grupos. Conclui-se que a hipofunção tireoidiana e gonadal apresentam efeitos distintos no baço e que na associação hipotireoidismo-hipogonadismo há aumento menos acentuado do índice apoptótico e da expressão de caspase-3 esplênica do que no hipotireoidismo isolado.

Apoptosis, proliferation and histomorphometry of spleen were investigated in ovariectomized and non-ovariectomized adult Wistar rats maintained in hypothyroidism induced by daily administration of propylthiouracil (PTU) during 120 days. Two groups ovariectomized euthyroid and non-ovariectomized euthyroid were used as controls. Plasma was collected for free T4 dosage and the spleen for histomorphometry analysis, apoptosis index and the immunohistochemistry expression of caspase 3 and CDC47. Values of free T4 were lower in rats treated with PTU (p<0.05). In the hypothyroid groups there was some decrease in the spleen weight as well as the number and size of lymphoid follicles and there was some increase in the apoptotic index and the caspase 3 expression (p<0.05). However, the increase in the apoptosis index and the expression of caspase 3 in ovariectomized hypothyroid rats spleen was less accentuated than non-ovariectomized hypothyroid ones (p<0.05). The ovariectomized euthyroid group presented white pulp hyperplasia in comparison to the non-ovariectomized euthyroid group. There was no difference in the CDC47 expression between groups. It was concluded that the thyroid and ovarian hypofunction have distinct effects on the spleen and that in the hypothyroidism-hypogonadism association, the increase in the apoptosis index and in the expression of splenic caspase 3 is not as much as in isolated hypothyroidism.

Histoenzymological study on the toxicity of copper sulphate in the digestive glands of Lymnaea luteola.

During 24 and 48 hr of exposure, the digestive glands of Lymnaea treated with a lethal concentration of 0.038 mgl(-1) CuSO4 revealed intense activity of acid phosphatase in perilobular margin. On the other hand, same area of the gland showed moderate activity of ATPase during 24 and 48 hr of exposure. However, alkaline phosphatase showed average activity in perialveolar region and perilobular margin during 24 and 48, and 72 hr of exposure respectively The changes in the activity of these enzymes were nonsignificant in alveolar margin and perialveolar region of the gland. It is interesting to note moderate activity of acid phosphatase in perialveolar region during 24 hr of exposure.

Chelation therapy and vanadium: effect on reproductive organs in rats.

Present investigation was planned to evaluate the therapeutic effectiveness of chelating agents against vanadium intoxication on blood and reproductive organs of rats. Male and female albino rats were injected vanadyl sulphate (7.5 mg/kg, po, for 21 days, 5 days in a week). Chelating agents tiron (T) alone and in combination with lipoic acid (LA), vitamin E (vit E) and selenium (Se) were given for 2 days/week. With the administration of vanadyl sulphate to rats fructose level in seminal vesicles was significantly (P< or =0.05) declined. The activities of alkaline phosphatase and adenosine triphosphatase were also decreased, whereas glycogen content and acid phosphatase activity increased in testis, seminal vesicles, ovaries and uterus after toxicant exposure. Significant changes in serum transaminases, serum alkaline phosphatase and lactate dehydrogenase were recouped by chelation therapy. Lipid peroxidation, reduced glutathione level and triglycerides levels altered significantly after exposure to vanadium in rats. The ultrastructural damage in spermatogenic stages in treated animals showed recovery pattern after therapy. Co-treatment with antioxidants restored these activities. The most effective combination was tiron + selenium followed by tiron + vitamin E, and tiron + lipoic acid.

Total ATPases activity in different tissues of albino mice exposed to aluminium acetate.

This study has revealed significant variation in total ATPase activity after administration of aluminium acetate in different tissues of albino mice. With sublethal dose (3.5 mg/kg body weight) of aluminium acetate, total ATPase activity was decreased in brain (-50.61), liver (-50.69), kidney (-30.74), heart (-64.07), muscle (-51.50) and testis (-65.53) of albino mice. The decrement was enhanced with the increase of aluminium acetate. ATPases play an important role in the maintenance of cell permeability and energy transformation in the biological system. The results suggest that ATPase has a particular sensitivity to aluminium acetate.

'DEAD-box' helicase from Plasmodium falciparum is active at wide pH and is schizont stage-specific.

BACKGROUND & OBJECTIVES: DNA helicases catalyse unwinding of duplex DNA in an ATP-dependent manner and are involved in all the basic genetic processes. In order to study these important enzymes in the human malaria parasite we have recently cloned the first full-length 'DEAD-box' helicase gene from Plasmodium falciparum (3D7). In the present study, we report some of the important activities of the encoded protein. METHODS: We have expressed the P. falciparum helicase in Escherichia coli and characterised the encoded biochemically active helicase protein. The characterisation of the protein was carried out using radioactively labeled substrate and the standard strand displacement assay. The localisation of the enzyme was studied using immunofluorescence assay. RESULTS & CONCLUSION: P. falciparum helicase gene is 1551 bp in length and encodes for a protein consisting of 516 amino acid residues with a predicted molecular mass of 59.8 kDa. The protein is designated as Plasmodium falciparum DEAD-box helicase 60 kDa in size (PfDH60). Purified PfDH60 showed ATP and Mg2+ dependent DNA unwinding, ssDNA-dependent ATPase and ATP-binding activities. Interestingly, this is a unique helicase because it works at a wide pH range (from 5.0-10.0). The peak expression of PfDH60 is mainly in schizont stages of the development of P. falciparum, where DNA replication is active. The cell-cycle dependent expression suggests that PfDH60 may be involved in the process of DNA replication and distinct cellular processes in the parasite and this study should make an important contribution in our better understanding of DNA metabolic pathways in the parasite.

Clinicopathological Characteristics of Colorectal Cancer with Family History: an Evaluation of Family History as a Predictive Factor for Microsatellite Instability

To determine whether family history of cancer may be a risk factor for the mutator phenotype in colorectal cancer, we recruited 143 consecutive colorectal cancer patients with a family history of accompanying cancers not meeting the Amsterdam criteria. Microsatellite instability (MSI) at 5 markers, hMLH1-promoter methylation, and expression of mismatch repair (MMR) proteins (hMLH1, hMSH2, hMSH6, hMPS1, and hPMS2) were determined. Among the relatives of familial colorectal cancer patients, colorectal cancer was the most common tumor type. Of the proband colorectal cancers, 26 (18.2%) showed high-level MSI (MSI-H); 47 additional tumors with mutator phenotype (32.9%) were identified by hMLH1-promoter methylation and/or loss of MMR protein expression. Mutator phenotype was associated with right-sided colon cancer and the type of accompanying cancer. Family history, which was differentially quantified according to the degree of relatives and the type of accompanying cancers, effectively discriminated MSI-H from microsatellite stable (MSS) and low-level microsatellite instability (MSI-L) and mutator phenotypes. Our findings indicate that familial colorectal cancer may be associated with multiple occurrences of colorectal or accompanying cancers and that family history could be correlated with microsatellite instability.

Cross-immunoreactivity between anti-potato apyrase antibodies and mammalian ATP diphosphohydrolases: potential use of the vegetal protein in experimental schistosomiasis

We have previously showed that Schistosoma mansoni ATP-diphosphohydrolase and Solanum tuberosum potato apyrase share epitopes and the vegetable protein has immunostimulatory properties. Here, it was verified the in situ cross-immunoreactivity between mice NTPDases and anti-potato apyrase antibodies produced in rabbits, using confocal microscopy. Liver samples were taken from Swiss Webster mouse 8 weeks after infection with S. mansoni cercariae, and anti-potato apyrase and TRITC-conjugated anti-rabbit IgG antibody were tested on cryostat sections. The results showed that S. mansoni egg ATP diphosphohydrolase isoforms, developed by anti-potato apyrase, are expressed in miracidial and egg structures, and not in granulomatous cells and hepatic structures (hepatocytes, bile ducts, and blood vessels). Therefore, purified potato apyrase when inoculated in rabbit generates polyclonal sera containing anti-apyrase antibodies that are capable of recognizing specifically S. mansoni ATP diphosphohydrolase epitopes, but not proteins from mammalian tissues, suggesting that autoantibodies are not induced during potato apyrase immunization. A phylogenetic tree obtained for the NTPDase family showed that potato apyrase had lower homology with mammalian NTPDases 1-4, 7, and 8. Further analysis of potato apyrase epitopes could implement their potential use in schistosomiasis experimental models.

Does melatonin have a time-dependent effect on brain and gill ionic metabolism in a teleost, Anabas testudineus (Bloch)?

Exogenous administration of 0.20, 0.40 and 0.60 microg/g body weight melatonin over a 24 hr cycle caused an inhibition of Na+, K+ ATPase activity in both brain and gills of A. testudineus. However, Ca2+ ATPase activity in the brain was significantly inhibited by the highest dose, and that in the gill at all the doses of melatonin. Evening injection of melatonin had an inhibitory effect on both brain and gill Na+ K+ and Ca2+ ATPase activity. Melatonin treatment in the morning for 12 hrs did not have an effect on brain Na+, K+ ATPase, while Ca2+ ATPase was inhibited. Similar treatment stimulated Na+, K+ and Ca2+ ATPase activity in the gills. Sodium, potassium and calcium ions in the gill were significantly reduced in the evening treated group while no change was observed in the morning melatonin injected group. The results suggest that melatonin elicits a time-dependent effect on the enzymes and ionic content in the brain and gills of A. testudineus.

Cop-like operon: Structure and organization in species of the Lactobacillale order

Copper is an essential and toxic trace metal for bacteria and, therefore, must be tightly regulated in the cell. Enterococcus hirae is a broadly studied model for copper homeostasis. The intracellular copper levels in E. hirae are regulated by the cop operon, which is formed by four genes: copA and copB that encode ATPases for influx and efflux of copper, respectively; copZ that encodes a copper chaperone; and copY, a copper responsive repressor. Since the complete genome sequence for E. hirae is not available, it is possible that other genes may encode proteins involved in copper homeostasis. Here, we identified a cop-like operon in nine species of Lactobacillale order with a known genome sequence. All of them always encoded a CopY-like repressor and a copper ATPase. The alignment of the cop-like operon promoter region revealed two CopY binding sites, one of which was conserved in all strains, and the second was only present in species of Streptococcus genus and L. johnsonii. Additional proteins associated to copper metabolism, CutC and Cupredoxin, also were detected. This study allowed for the description of the structure and organization of the cop operon and discussion of a phylogenetic hypothesis based on the differences observed in this operon's organization and its regulation in Lactobacillale order.

Effect of sodium nitroprusside on H+-ATPase activity and ATP concentration in Candida albicans.

ATP hydrolysis by plasma membrane H+-ATPase from Candida albicans has been investigated in presence of nitric oxide and various nutrients (sugars and amino acids). Sodium nitroprusside (SNP) was used as nitric oxide donor. It was found that ATP concentration decreased in SNP treated cells which was more in presence of sugars like glucose, xylose and 2-deoxy-D-glucose and amino acids as compared to their respective controls. The activity of H+-ATPase from plasma membrane decreased by 70 % in SNP treated cells. Both in vivo and in vitro treatments of SNP showed almost similar effects of decrease in ATPase activity. Effect of SNP was more pronounced in presence of nutrients. Interestingly, it was observed that vanadate did not show any independent effect in presence of nitric oxide. Several workers have reported similar type of results with other P-type ATPases. For the first time, it was observed in the present study that in presence of nitric oxide, H+-ATPase activity decreased like other P-type ATPases. Our study indicated that NO had a significant effect on ATP synthesis and activity of H+- ATPase. In the presence of NO, the ATP concentration was decreased indicating it affected mitochondrial electron transport chain. It may be concluded that NO, not only affects (inhibit) mitochondrial electron transport chain but also interferes with H+- ATPase of plasma membrane by changing its conformation resulting in decreased activity.