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PubMed2000

Hydrophobic pairwise interactions stabilize alpha-conotoxin MI in the muscle acetylcholine receptor binding site.

The present work delineates pairwise interactions underlying the nanomolar affinity of alpha-conotoxin MI (CTx MI) for the alpha-delta site of the muscle acetylcholine receptor (AChR). We mutated all non-cysteine residues in CTx MI, expressed the alpha(2)betadelta(2) pentameric form of the AChR in 293 human embryonic kidney cells, and measured binding of the mutant toxins by competition against the initial rate of (125)I-alpha-bungarotoxin binding. The CTx MI mutations P6G, A7V, G9S, and Y12T all decrease affinity for alpha(2)betadelta(2) pentamers by 10,000-fold. Side chains at these four positions localize to a restricted region of the known three-dimensional structure of CTx MI. Mutations of the AChR reveal major contributions to CTx MI affinity by Tyr-198 in the alpha subunit and by the selectivity determinants Ser-36, Tyr-113, and Ile-178 in the delta subunit. By using double mutant cycles analysis, we find that Tyr-12 of CTx MI interacts strongly with all three selectivity determinants in the delta subunit and that deltaSer-36 and deltaIle-178 are interdependent in stabilizing Tyr-12. We find additional strong interactions between Gly-9 and Pro-6 in CTx MI and selectivity determinants in the delta subunit, and between Ala-7 and Pro-6 and Tyr-198 in the alpha subunit. The overall results reveal the orientation of CTx MI when bound to the alpha-delta interface and show that primarily hydrophobic interactions stabilize the complex.

Amino Acid Sequence↗

PubMed1998

The root knot nematode resistance gene Mi from tomato is a member of the leucine zipper, nucleotide binding, leucine-rich repeat family of plant genes.

The Mi locus of tomato confers resistance to root knot nematodes. Tomato DNA spanning the locus was isolated as bacterial artificial chromosome clones, and 52 kb of contiguous DNA was sequenced. Three open reading frames were identified with similarity to cloned plant disease resistance genes. Two of them, Mi-1.1 and Mi-1.2, appear to be intact genes; the third is a pseudogene. A 4-kb mRNA hybridizing with these genes is present in tomato roots. Complementation studies using cloned copies of Mi-1.1 and Mi-1.2 indicated that Mi-1.2, but not Mi-1.1, is sufficient to confer resistance to a susceptible tomato line with the progeny of transformants segregating for resistance. The cloned gene most similar to Mi-1.2 is Prf, a tomato gene required for resistance to Pseudomonas syringae. Prf and Mi-1.2 share several structural motifs, including a nucleotide binding site and a leucine-rich repeat region, that are characteristic of a family of plant proteins, including several that are required for resistance against viruses, bacteria, fungi, and now, nematodes.

Amino Acid Sequence↗

PubMed2003

Myocardial infarction (MI) at the work-site among Polish seafarers. The risk and the impact of occupational factors.

OBJECTIVES: To study the incidence of myocardial infarction (MI) on board ship, the impact of conditions of work on the mortality and survival rate, to analyze the recommendations and possible preventive interventions to reduce the loss of life at the work-site among Polish seafarers and deep-sea fishermen. METHODS: To use various sources of information for the retrospective study of the MI incidence among 11,325 Polish seamen and deep-sea fishermen. To analyse the impact of the marine environment and work-related factors on the course, treatment and prognosis of MI. The results were compared with the incidence and mortality rate of MI among general male workers population in Poland. RESULTS: In this retrospective study, 106 cases of the MI at sea were included. Their diagnosis was based on the medical documentation and legal investigation of cases. The most significant findings seem to indicate that the incidence of MI at the work-site among Polish seafarers was not higher than among general population of Polish males in the same decade, probably due to the "healthy worker" effect. However, the pre-hospital and one-month mortality due to MI were higher in the group of seafarers as compared to the group of general male population. The analysis of conditions of seafarers' work and life and the possibility of treatment at sea in a case of the MI on board strongly suggest that the work-related factors had significant impact on the loss of life among them. The preventive measures, very complex and difficult from the organizational and economic point of view, are still far from being recommended and implemented in the shipping industry. CONCLUSIONS: The study has confirmed that seafarers are at high risk of death in case of a MI on board. The impact of their work environment and work-related factors reduce significantly the chance of survival at sea in such cases. The preventive measures are difficult to implement.

Adult↗

PubMed1993

A helix-loop-helix transcription factor-like gene is located at the mi locus.

The mouse microphthalmia phenotype is complex and consists of one or more of the following phenotypic alterations: a lack of pigmentation, small eyes, a mast cell defect, and bone abnormalities. The locus for this allele has been assigned to chromosome 6. A single gene defect that produces such a pleiotropic effect has suggested some involvement at a control point in development. Recently a mutant line of mice carrying a transgene insertion, which represents a new allelic form of mi, was described. The integration site of the transgene from these mi(tg) mice was cloned and analyzed. An exon sequence was discovered adjacent to the insertion. Computer analysis of this nucleotide sequence revealed the presence of a motif indicative of the helix-loop-helix class of transcription factors. The gene was expressed in a number of tissues from wild type animals but was absent in the tissue RNA from mi(tg) mice. Southern blot analysis demonstrated a deletion of some of the genetic material for this gene in the mi(tg) mice. This is consistent with the lack of expression in the mi(tg) mice. Interestingly, when DNA from other mi allelic variants was subjected to a similar analysis, a deletion was also observed in this gene in two other mi lines. Taken together, these data suggest that the gene encoding this new helix-loop-helix DNA-binding protein, and residing in the mi locus, is a strong candidate for the mi gene.

Amino Acid Sequence↗

PubMed1995

The major dermatomyositis-specific Mi-2 autoantigen is a presumed helicase involved in transcriptional activation.

OBJECTIVE: To characterize the complementary DNA (cDNA) and protein sequences of autoantigens recognized by anti-Mi-2 antibodies, using recombinant Mi-2 proteins for improved autoantibody detection. METHODS: A cDNA expression library was immunoscreened, and cDNA isolation, alignment, and sequence analysis were performed. Northern blotting and in situ hybridization techniques were used. A recombinant protein (rMi-2) was synthesized. Immunoprecipitation of 35S-methionine-labeled HEp-2 cell proteins and immunoblotting of rMi-2 and natural nuclear proteins were performed. Immunofluorescence studies were done with anti-Mi-2 positive sera of dermatomyositis (DM) patients, and with human or rabbit antibodies specific for rMi-2. Antibody screening of systemic lupus erythematosus, rheumatoid arthritis, DM, and antinuclear antibody-positive human sera was performed using an rMi-2 protein enzyme-linked immunosorbent assay (ELISA). RESULTS: A major antigen recognized by anti-Mi-2 positive sera of DM patients was found to constitute a 218-kd nuclear protein (218-kd Mi-2) encoded on chromosome 12 and to belong to the SNF2/RAD 54 helicase family. Human and rabbit antibodies that were affinity purified using the recombinant protein reacted with and precipitated a nuclear protein of similar size, which was also recognized by anti-Mi-2 sera. Anti-218-kd Mi-2 antibodies detected by rMi-2 protein ELISA seemed to be mainly restricted to sera from patients with DM. CONCLUSION: The molecular characterization of the 218-kd Mi-2 antigen may contribute to our understanding of autoimmune phenomena in DM. The use of immunoreactive recombinant proteins allows structural and functional studies of the helicase and the development of sensitive and accurate antibody screening tests.

Adenosine Triphosphatases↗

PubMed2003

Mi-2 beta associates with BRG1 and RET finger protein at the distinct regions with transcriptional activating and repressing abilities.

Mi-2 beta is the main component of the nucleosome remodeling and deacetylase complex and plays an important role in epigenetic transcriptional repression. Here we show that the amino-terminal and carboxyl-terminal regions of Mi-2 beta have distinct transcriptional activities and bind to BRG1, a component of the SWI/SNF complex, and the RET finger protein (RFP), respectively. Analysis by luciferase reporter assay revealed that the amino-terminal region of Mi-2 beta has a strong transactivating ability, whereas its carboxyl-terminal region has transcriptional repressive activity. Co-localization and association of Mi-2, RFP, and histone deacetylase 1 suggested that these proteins cooperate in transcriptional repression. Furthermore, the functional importance of the association of Mi-2 beta and RFP was confirmed by using Rfp-/- fibroblasts. On the other hand, we demonstrated that Mi-2 and BRG1 were associated with each other and that the bromodomain region of BRG1 strongly suppressed transactivation by the amino-terminal region of Mi-2 beta. The findings that Mi-2 beta interacts with both transactivating and repressing proteins and directly associates with another chromatin remodeling protein, BRG1, provide new insight into the formation of multiprotein supercomplex involved in transcriptional regulation.

Adenosine Triphosphatases↗

PubMed2006

Posttraumatic stress disorder and general health problems following myocardial infarction (Post-MI PTSD) among older patients: the role of personality.

OBJECTIVES: This study aimed to: (1) investigate the degree to which older MI patients experienced post-MI PTSD and general health problems, and to (2) identify the association between five personality traits, post-MI PTSD and general health problems among these older people. METHODS: The sample comprised 96 older MI patients and were recruited from two general practices. They were interviewed using the Posttraumatic Stress Diagnostic Scale (PDS), the General Health Questionnaire-28 (GHQ-28) and the NEO-Five Factor Inventory (NEO-FFI). The control group comprised 92 older people who experienced no MI. They were assessed using the GHQ-28. RESULTS: The older MI patients experienced hyperarousal the most, followed by avoidance and re-experiencing symptoms. PDS revealed that 30% had full PTSD. The older patients were conscientious and agreeable individuals. They were also somewhat extraverted and open to experience. They also showed traits of neuroticism. Controlling for bypass surgery, heart failure, previous mental health difficulties, angina and angioplasty, regression analyses showed that neuroticism predicted re-experiencing and avoidance symptoms and general health total. Neuroticism and less agreeableness predicted hyperarousal symptoms. CONCLUSIONS: MI older patients could manifest PTSD symptoms and general health problems following their MI. Personality traits, particularly neuroticism and antagonism, played a role in the manifestation of these symptoms.

Aged↗

PubMed2006

The role of sexual satisfaction, age, and cardiac risk factors in the reduction of post-MI anxiety.

BACKGROUND: Anxiety is common after myocardial infarction (MI); however, little is known about the role of sexual satisfaction and return to sexual activity on anxiety post-MI. AIM: To examine the role of sexual satisfaction in reducing anxiety post-MI. METHODS: Patients with acute myocardial infarction (MI) recruited from one U.S. medical center completed questionnaires at baseline while hospitalized and at 1, 3, and 5 months post-MI. This analysis includes 64 patients compared on low or high anxiety at 5 months post-MI using sexual satisfaction and selected demographic and clinical variables in the analysis. RESULTS: Patients with high anxiety scores reported lower sexual satisfaction (p < .001) and a higher percentage of coronary risk factors (p < .01). The OLS regression model provided similar results with an adjusted R-square of .422, accounting for approximately 42% of anxiety (p < .001). Sexual satisfaction accounted for the most variance in the model, showing an inverse relationship between sexual satisfaction and anxiety. An inverse relationship also existed between age and anxiety. CONCLUSIONS: Anxiety is common after MI, and decreased sexual satisfaction appears to contribute to heightened anxiety. Attention to sexual concerns of MI patients before and after hospital discharge may improve psychosocial outcomes.

Age Factors↗

PubMed2005

Who is at risk of post-MI depressive symptoms?

OBJECTIVE: The aim of this study was to identify cardiologic, psychologic, and demographic risk factors in two groups of patients with post-myocardial infarction (MI) depressive symptoms (in-hospital and during the postdischarge year). METHODS: Patients admitted for MI were assessed for depressive symptoms with the Beck Depression Inventory (BDI) during hospitalization and 3, 6, and 12 months post-MI. We contrasted both groups with nondepressed patients. RESULTS: Pre-MI vital exhaustion, living alone, history of depressive disorder, history of MI, poor performance on exercise tolerance testing, and female gender were significantly and independently associated with in-hospital depressive symptoms. Pre-MI vital exhaustion, history of depressive disorder, female gender, poor ejection fraction, and longer hospital stay were independent predictors of the development of postdischarge depressive symptoms. CONCLUSIONS: Post-MI depressive symptoms seem largely driven by the psychological and social consequences of the MI in patients vulnerable to depression, as indexed by a history of depression and vital exhaustion.

Aged↗

PubMed2002

Dual abnormal effects of mutant MITF encoded by Mi(wh) allele on mouse mast cells: decreased but recognizable transactivation and inhibition of transactivation.

MITF is a basic helix-loop-helix leucine zipper-type transcription factor and is important for development of mast cells. MITF encoded by Mi(wh) allele (Mi(wh)-MITF) was mutated at a single amino acid of basic domain, and possessed a deficient but apparent DNA-binding ability. Here, we characterized the unique effects of Mi(wh)-MITF on the expression of mast cell-related genes. The expression level of mouse mast cell protease (mMCP)-4, -5, and -6 genes in Mi(wh)/Mi(wh) cultured mast cells (CMCs) was intermediate between levels of normal (+/+) CMCs and tg/tg CMCs, which did not express any MITFs. Mi(wh)-MITF appeared to show the positive transactivation effect through the remaining DNA-binding ability. On the other hand, the expression level of tryptophan hydroxylase gene was lower in Mi(wh)/Mi(wh) CMCs than in tg/tg CMCs, suggesting the inhibitory effect of Mi(wh)-MITF on the transactivation. Mi(wh)-MITF possessed dual abnormal effects on transactivation of mast cell-related genes.

Alleles↗

PubMedDate not supplied

[Can specific verbal descriptors be useful in differentiating those with and without MI? Findings from a two-year study].

This paper discusses the findings of a two-year study, which was based in an eleven-bedded coronary care unit in the South-West of England. The study aimed to explore the difference between the verbal descriptors used by those with and without MI in order to determine their contribution in assessing patients with a suspected MI. The study also examined whether any differences existed between the words men and women with MI used to describe their chest pain symptoms. All patients admitted with an episode of chest pain were eligible to participate providing that they were pain-free at 24 hours after admission, were over 18 years of age and could speak English. The sample comprised of 266 patients with MI and 275 without MI. All patients were offered a menu of 12 sensory and 10 effective words, which had been validated in previous research. The results suggest that there is little difference in the words patients with and without MI use to describe their chest pain and this may reinforce the complexity in obtaining an accurate differential diagnosis. However, there are some clear differences in the vocabulary of men and women with MI. Women with MI appeared to report more emotive language than men and expressed their chest pain in terms of being "frightened" (p < 0.05) and "terrified" (21.1 % vs 12.1 %). The discussion will examine the methodological issues and possible practice implications for the future.

Aged↗

PubMed2004

Iodo-alpha-conotoxin MI selectively binds the alpha/delta subunit interface of muscle nicotinic acetylcholine receptors.

The embryonic mouse muscle nicotinic acetylcholine receptor (nAChR) is a ligand-gated ion channel formed by alpha1, beta1, delta, and gamma subunits. The receptor contains two ligand binding sites at alpha/delta and alpha/gamma subunit interfaces. [(3)H]Curare preferentially binds the alpha/gamma interface. We describe the synthesis and properties of a high-affinity iodinated ligand that selectively binds the alpha/delta interface. An analogue of alpha-conotoxin MI was synthesized with an iodine attached to Tyr-12 (iodo-alpha-MI). The analogue potently blocks the fetal mouse muscle subtype of nAChR expressed in Xenopus oocytes. It failed, however, to block alpha3beta4, alpha4beta2, or alpha7 nAChRs. Iodo-alpha-MI potently blocks the alpha1beta1delta but not the alpha1beta1gamma subunit combination expressed in Xenopus oocytes indicating selectivity for the alpha/delta subunit interface. Alpha-conotoxin MI was subsequently radioiodinated, and its properties were further evaluated. Saturation experiments indicate that radioiodinated alpha-conotoxin MI binds to TE671 cell homogenates with a Hill slope of 0.95 +/- 0.0094. Kinetic studies indicate that the binding of [(125)I]alpha-conotoxin MI is reversible (k(off) = 0.084 +/- 0.0045 min(-1)); k(on) is 8.5 x 10(7) min(-1) M(-1). The calculated k(d) is 0.98 nM. This potency is approximately 20-fold higher than the unmodified alpha-MI peptide. Unlike [(125)I]alpha-bungarotoxin, [(125)I]alpha-conotoxin MI binding to TE671 cell homogenates is fully displaceable by the small molecule antagonist d-tubocurarine.

Animals↗

PubMed2003

Exercise intolerance during post-MI heart failure in rats: prevention with supplemental dietary propionyl-L-carnitine.

Exercise capacity in patients with several types of cardiovascular disease can be improved with dietary carnitine, or carnitine derivatives. Mechanisms underlying this improvement remain largely unknown in part due to a lack of animal models of cardiac pathology in which carnitine derivatives improve exercise tolerance. Our goal was to evaluate the ability of propionyl-L-carnitine (PLC) to improve exercise tolerance in a rat model of exercise intolerance. Fischer 344 rats were followed after either a moderate size MI (n = 22) or sham MI surgery (n = 14). Starting 10 days post-surgery 10 of the MI and 7 of the sham rats received 100 mg/kg/day PLC in drinking water, which increased plasma and LV total l-carnitine concentrations 15-23% (p < 0.05). Rats were followed longitudinally until a statistically significant decrease in exercise capacity occurred in one of the groups, at which time all rats were sacrificed for study of the isolated perfused hearts. At 12-weeks post-MI exercise capacity had decreased 16 +/- 7% (p < 0.05) in the MI group, but remained within 3% of baseline in the MI group that received PLC and the sham groups. Both MI groups exhibited the same degree of LV dilation, decrease in fractional shortening, and blunting of the response to isoproterenol. We conclude that supplemental dietary PLC attenuates the exercise intolerance that occurs secondary to post-MI heart failure in rats, but that this beneficial effect is not attributable to altered LV remodeling, an improved response to beta-adrenergic stimulation, or increased skeletal muscle citrate synthase activity.

Animals↗

PubMed1998

The nematode resistance gene Mi of tomato confers resistance against the potato aphid.

Resistance against the aphid Macrosiphum euphorbiae previously was observed in tomato and attributed to a novel gene, designated Meu-1, tightly linked to the nematode resistance gene, Mi. Recent cloning of Mi allowed us to determine whether Meu-1 and Mi are the same gene. We show that Mi is expressed in leaves, that aphid resistance is isolate-specific, and that susceptible tomato transformed with Mi is resistant to the same aphid isolates as the original resistant lines. We conclude that Mi and Meu-1 are the same gene and that Mi mediates resistance against both aphids and nematodes, organisms belonging to different phyla. Mi is the first example of a plant resistance gene active against two such distantly related organisms. Furthermore, it is the first isolate-specific insect resistance gene to be cloned and belongs to the nucleotide-binding, leucine-rich repeat family of resistance genes.

Animals↗

PubMed2005

Magnetoacoustic tomography with magnetic induction (MAT-MI).

We report our theoretical and experimental investigations on a new imaging modality, magnetoacoustic tomography with magnetic induction (MAT-MI). In MAT-MI, the sample is located in a static magnetic field and a time-varying (micros) magnetic field. The time-varying magnetic field induces an eddy current in the sample. Consequently, the sample will emit ultrasonic waves by the Lorentz force. The ultrasonic signals are collected around the object to reconstruct images related to the electrical impedance distribution in the sample. MAT-MI combines the good contrast of electrical impedance tomography with the good spatial resolution of sonography. MAT-MI has two unique features due to the solenoid nature of the induced electrical field. Firstly, MAT-MI could provide an explicit or simple quantitative reconstruction algorithm for the electrical impedance distribution. Secondly, it promises to eliminate the shielding effects of other imaging modalities in which the current is applied directly with electrodes. In the theoretical part, we provide formulae for both the forward and inverse problems of MAT-MI and estimate the signal amplitude in biological tissues. In the experimental part, the experimental setup and methods are introduced and the signals and the image of a metal object by means of MAT-MI are presented. The promising pilot experimental results suggest the feasibility of the proposed MAT-MI approach.

Acoustics↗

PubMed2004

The outcome of ICSI of immature MI oocytes and rescued in vitro matured MII oocytes.

BACKGROUND: The use of immature oocytes is limited to cases where these are the only available oocytes, and they are usually only microinjected with sperm after having undergone maturation in vitro. This study compares the outcome of injection of sperm into metaphase I oocytes immediately after their denudation (MI) performed 2 h after their retrieval, with the outcome of injection of sperm into rescued in vitro matured metaphase II (IVM MII) oocytes after their short incubation in routine laboratory conditions. METHODS: ICSI was performed on MI oocytes, rescued IVM MII oocytes and on MI oocytes that were incubated but failed to extrude their first polar body (arrested IVM MI). Fertilization and cleavage rates were compared with those achieved in mature metaphase II oocytes (MII). RESULTS: ICSI of MI oocytes showed impaired performance compared with ICSI of rescued IVM MII oocytes and MII oocytes, in terms of oocyte degeneration rate (11 versus 6 versus 4%; P < 0.0001), fertilization rate (28 versus 44 versus 68%; P < 0.0001) and multipronucleated fertilization (10 versus 4 versus 4%; P < 0.01). The cleavage rate was lower in rescued IVM MII oocytes compared with MII oocytes (86 versus 95%; P < 0.01). Arrested IVM MI oocytes showed similar results to those of MI oocytes but had a lower cleavage rate (72 versus 96%; P < 0.01). CONCLUSIONS: The injection of rescued IVM MII oocytes is preferred to the injection of MI oocytes.

Adult↗

PubMed2004

Rme1 is necessary for Mi-1-mediated resistance and acts early in the resistance pathway.

The tomato gene Mi-1 confers resistance to root-knot nematodes (Meloidogyne spp.), potato aphid, and whitefly. Using genetic screens, we have isolated a mutant, rme1 (resistance to Meloidogyne spp.), compromised in resistance to M. javanica and potato aphid. Here, we show that the rme1 mutant is also compromised in resistance to M. incognita, M. arenaria, and whitefly. In addition, using an Agrobacterium-mediated transient assay in leaves to express constitutive gain-of-function mutant Pto(L205D), we demonstrated that the rme1 mutation is not compromised in Pto-mediated hypersensitive response. Moreover, the mutation in rme1 does not result in increased virulence of pathogenic Pseudomonas syringae or Mi-1-virulent M. incognita. Using a chimeric Mi-1 construct, Mi-DS4, which confers constitutive cell death phenotype and A. rhizogenes root transformation, we showed that the Mi-1-mediated cell death pathway is intact in this mutant. Our results indicate that Rme1 is required for Mi-1-mediated resistance and acts either at the same step in the signal transduction pathway as Mi-1 or upstream of Mi-1.

Animals↗

PubMed1972

Comparison of cysteine and tryptophan content of insoluble proteins derived from wild-type and mi-1 strains of Neurospora crassa.

The possibility of an amino acid substitution (cysteine for tryptophan) in a membrane protein of the [mi-1] strain of Neurospora crassa has been investigated in detail by using a double radioactive labeling procedure. Auxotrophic strains of Neurospora having wild-type [+] or [mi-1] cytoplasm have been grown under conditions which result in the specific labeling of protein tryptophan with (3)H and protein cysteine with (35)S. Although the least soluble 1 to 20% of the [mi-1] mitochondrial membrane protein was usually found to have a higher Cys/Trp ratio (ratio of cysteine plus half-cystine to tryptophan) than the corresponding [+] fraction, it has been shown that these differences were due mainly to the presence of differential amounts of a very insoluble, cysteine-rich (Cys-rich) material. The same Cys-rich material was found in variable amounts in both [+] and [mi-1] cultures, but the concentration was usually higher in the [mi-1] cultures. The Cys-rich material is clearly distinct from "structural protein" on the basis of amino acid composition and appears to have no direct relationship to the [mi-1] phenotype. In the absence of the Cys-rich material, no difference between the Cys/Trp ratios of corresponding [+] and [mi-1] membrane proteins could be detected. We conclude, therefore, that the previously postulated amino acid substitution of cysteine for tryptophan in [mi-1] membrane protein is incorrect.

Autoanalysis↗