Correlation between benzene and testosterone in workers exposed to urban pollution.

AIM: Many studies have examined the effects of benzene on testosterone. The purpose of this study was to evaluate the possible correlation between the blood levels of benzene and the levels of testosterone. MATERIALS AND METHODS: The study involved a group of 148 subjects. For every worker have been made out a blood sample for the evaluation of benzene and testosterone levels and an urine analysis for the evaluation of the levels of trans, trans-muconic acid and S-phenylmercapturic acid. We estimated the Pearson correlation coefficient between the variables in the sample and the urinary metabolites, age, length of service, gender, BMI. For the analysis of the major confounding factors it was performed a multiple linear regression. RESULTS: The Pearson correlation coefficiet showed: 1. a significant inverse correlation between the S-phenyl mercapturic acid and free testosterone; 2. a significant direct correlation between trans-trans muconic acid and BMI. After dividing the sample according to the median of blood benzene (161.0 ng / L), Pearson correlation coefficient showed a significant inverse correlation between the S-phenyl mercapturic acid and free testosterone in the group with values below this median. CONCLUSIONS: Our results, to be considered preliminary, suggest that occupational exposure to low levels of benzene, present in urban pollution, affect the blood levels of testosterone. These results need to be confirmed in future studies, with the eventual possibility of including more specific fertility tests.

Work related stress and blood glucose levels.

AIM: The aim of the study is to evaluate work-related subjective stress in a group of workers on a major Italian company in the field of healthcare through the administration of a valid "questionnaire-tool indicator" (HSE Indicator Tool), and to analyze any correlation between stress levels taken from questionnaire scores and blood glucose values. MATERIAL AND METHODS: We studied a final sample consisting of 241 subjects with different tasks. The HSE questionnaire - made up of 35 items (divided into 7 organizational dimensions) with 5 possible answers - has been distributed to all the subjects in occasion of the health surveillance examinations provided by law. The questionnaire was then analyzed using its specific software to process the results related to the 7 dimensions. These results were compared using the Pearson correlation and multiple linear regression with the blood glucose values obtained from each subject. RESULTS: From the analysis of the data the following areas resulted critical, in other words linked to an intermediate (yellow area) or high (red area) condition of stress: sustain from managers, sustain from colleagues, quality of relationships and professional changes. A significant positive correlation (p <0.05) between the mean values of all critical areas and the concentrations of glucose values have been highlighted with the correlation index of Pearson. Multiple linear regression confirmed these findings, showing that the critical dimensions resulting from the questionnaire were the significant variables that can increase the levels of blood glucose. CONCLUSION: The preliminary results indicate that perceived work stress can be statistically associated with increased levels of blood glucose.

Work-related stress in healthcare workers.

BACKGROUND: In the assessment of work-related stress it is crucial to find the factors that generate and increase it in order to identify categories of individuals at risk, to plan interventions for prevention, elimination or reduction of risk. The aim of the study is to assess the subjective stress in 68 workers of a large Italian company dealing with human health, through the use of a questionnaire-indicating tool, elaborated by the Italian National Institute for insurance against accidents at work (INAIL) and developed by the Health and Safety Executive (HSE). METHODS: We studied a final sample of 68 individuals (34 drivers/rescuers and 34 video display unit (VDU) operators). The questionnaire consists of 35 items (divided into six areas) with five possible answers each, that cover working conditions considered potential causes of stress. RESULTS: The drivers/rescuers had a better performance than the VDU operators, especially in the areas "demand", "relationships" and "role". We compared men and women in the two groups, finding that, in VDU operators, women had a better performance than men in all areas, except "role" and "changes", in which the overall scores were the same in men and women. In the drivers/rescuers women showed more critical scores in the items "relationships" and "change". CONCLUSION: The results show that: the questionnaire-indicating tool is useful, with a demonstrated effectiveness for the occupational physician during the visits and proven validity; additional future efforts should focus on understanding the psycho-social, organizational and individual problems related to stress and the consequent implementation of preventive measures.

A novel autoregulatory loop between the Gcn2-Atf4 pathway and (L)-Proline [corrected] metabolism controls stem cell identity.

Increasing evidence indicates that metabolism is implicated in the control of stem cell identity. Here, we demonstrate that embryonic stem cell (ESC) behaviour relies on a feedback loop that involves the non-essential amino acid L-Proline (L-Pro) in the modulation of the Gcn2-Eif2α-Atf4 amino acid starvation response (AAR) pathway that in turn regulates L-Pro biosynthesis. This regulatory loop generates a highly specific intrinsic shortage of L-Pro that restricts proliferation of tightly packed domed-like ESC colonies and safeguards ESC identity. Indeed, alleviation of this nutrient stress condition by exogenously provided L-Pro induces proliferation and modifies the ESC phenotypic and molecular identity towards that of mesenchymal-like, invasive pluripotent stem cells. Either pharmacological inhibition of the prolyl-tRNA synthetase by halofuginone or forced expression of Atf4 antagonises the effects of exogenous L-Pro. Our data provide unprecedented evidence that L-Pro metabolism and the nutrient stress response are functionally integrated to maintain ESC identity.

Relationship between reduced BCL-2 expression in circulating mononuclear cells and early nephropathy in type 1 diabetes.

Microalbuminuria is the earliest clinical evidence of diabetic nephropathy, but the mechanisms linking hyperglycemia and kidney complications are not clear. The aim of this study was to evaluate whether enhanced oxidative stress in patients with microalbuminuria can contribute to diabetic nephropathy development through downregulation of the antiapoptotic gene Bcl-2 that promotes in turn a pro-inflammatory status. We studied 30 patients with type 1 diabetes (15 with and 15 without microalbuminuria) compared to 15 matched healthy controls. Plasma oxidant status, and expression of Bcl-2, activated NF-kB, inducible Nitric Oxide synthase (iNOS), and monocyte chemoattractant protein (MCP)-1 in circulating monocytes were evaluated at baseline and after 8-week oral vitamin E treatment (600 mg b.i.d.). Bcl-2 expression was significantly reduced in microalbuminuric diabetic patients as a consequence of increased oxidant burden secondary to persistent hyperglycemia. Bcl-2 down-regulation was associated with enhanced expression of NF-kB, iNOS and MCP-1, and showed a strong correlation with the albumin excretion rate. Low Bcl-2 expression and high inflammatory status were normalized by vitamin E both in vivo and in vitro. Our study showed that Bcl-2 down-regulation in diabetic patients with poor glycemic control results in the activation of the NF-kB pathway leading to the development of nephropathy. Vitamin E might provide a novel form of therapy for prevention of nephropathy in diabetic patients in which an acceptable glycemic control is difficult to achieve despite insulin therapy.

High preprocedural non-HDL cholesterol is associated with enhanced oxidative stress and monocyte activation after coronary angioplasty: possible implications in restenosis.

OBJECTIVE: To investigate whether enhanced oxidant stress in patients undergoing percutaneous transluminal coronary angioplasty (PTCA) is associated with a higher concentration of non-high density lipoprotein (HDL) cholesterol at baseline, and whether this contributes to the inflammatory reaction and luminal renarrowing after PTCA. DESIGN: An ex vivo and in vitro study of 46 patients who underwent PTCA and who had repeat angiograms after six months. Blood samples were collected immediately before PTCA, and at 24 hours, 48 hours, and 15 days after. SETTING: Tertiary referral centre. SUBJECTS: 46 patients (30 male, 16 female; mean (SD) age, 62 (5) years) with stable or unstable angina who underwent elective PTCA. MAIN OUTCOME MEASURES: Continuous variable luminal loss as defined by change in minimum lumen diameter during follow up, normalised for vessel size; lag phase of low density lipoprotein to in vitro oxidation; plasma fluorescent products of lipid peroxidation (FPLP); plasma vitamin C and E; interleukin (IL) 1beta secretion from unstimulated monocytes; plasma C reactive protein (CRP). RESULTS: Restenosis occurred in 12 patients (26%). Oxidant stress after PTCA was greater (p < 0.0001 at 15 days) in the patients with restenosis and showed a significant correlation with the preprocedural concentration of non-HDL cholesterol (p < 0.001). Inflammatory reaction (as reflected by IL-1beta production and CRP) and late lumen loss were linearly correlated (p < 0.001) with lag phase and FPLP throughout the study, and inversely (p < 0.05) with vitamin C and E measured at two and 15 days after PTCA. CONCLUSIONS: This study provides evidence for the critical role of cholesterol dependent oxidant stress in the pathophysiology of restenosis after PTCA. The findings raise the possibility that drugs capable of modulating oxidant status might provide a novel form of adjuvant treatment in patients with hypercholesterolaemia undergoing PTCA.

Overexpression of functionally coupled cyclooxygenase-2 and prostaglandin E synthase in symptomatic atherosclerotic plaques as a basis of prostaglandin E(2)-dependent plaque instability.

BACKGROUND: Studies have implicated a role for prostaglandin (PG) E(2)-dependent matrix metalloproteinase (MMP) biosynthesis in the rupture of atherosclerotic plaque. Cyclooxygenase-2 (COX-2) and PGE synthase (PGES) are coregulated in nucleated cells by inflammatory stimuli. The aim of this study was to characterize the expression of COX-2 and PGES in carotid plaques and to correlate it with the extent of inflammatory infiltration and MMP activity and with clinical features of patients' presentation. METHODS AND RESULTS: Plaques were obtained from 50 patients undergoing carotid endarterectomy and divided into 2 groups (symptomatic and asymptomatic) according to clinical evidence of recent transient ischemic attack or stroke. Plaques were analyzed for COX-2, PGES, MMP-2, and MMP-9 by immunocytochemistry and Western blot, whereas zymography was used to detect MMP activity. Immunocytochemistry was used to identify CD68+ macrophages, CD3+ T lymphocytes, and HLA-DR+ cells. The percentage of macrophage-rich areas was larger (P<0.0001) in symptomatic plaques. COX-2, PGES, and MMPs were detected in all specimens; enzyme concentration, however, was significantly higher in symptomatic plaques. COX-2, PGES, and MMPs were especially noted in shoulders of symptomatic plaques, colocalizing with HLA-DR+ macrophages. All symptomatic plaques contained activated forms of MMPs. Finally, inhibition of COX-2 by NS-398 was accompanied by decreased production of MMPs that was reversed by PGE(2). CONCLUSIONS: This study demonstrates the colocalization of COX-2 and PGES in symptomatic lesions and provides evidence that synthesis of COX-2 and PGES by activated macrophages is associated with acute ischemic syndromes, possibly through metalloproteinase-induced plaque rupture.

Transcriptional cascades during spermatogenesis: pivotal role of CREM and ACT.

The gene CREM plays key physiological and developmental roles within the hypothalamic--pituitary--gonadal axis. We have previously shown that CREM is highly expressed in male postmeiotic cells. Spermiogenesis is a complex process by which postmeiotic male germ cells differentiate into mature spermatozoa. CREM regulates the expression of a number of post-meiotic genes involved in the process of spermiogenesis. Using homologous recombination we have generated CREM-mutant mice that display a complete block at the first step of spermiogenesis. The molecular mechanism by which CREM elicits its regulatory function involves ACT (Activator of CREM in Testis), a testis-specific coactivator constituted by a repeat of four and half LIM domains. ACT is coordinately expressed with CREM, associates with it and confers a powerful transcriptional activation function. It is able to bypass the classical requirement of CREM phosphorylation and recruiting of CBP.

Cloning and expression of activator of CREM in testis in human testicular tissue.

Activator of cAMP-responsive element modulator (CREM) in testis (ACT) has recently been found in the mouse testis where it activates CREM, a transcription factor essential for the differentiation of spermatids into mature spermatozoa. The importance of CREM in human spermatogenesis prompted us to examine whether ACT was also present in the human testis. Western blot analysis, performed with an anti-mouse ACT serum, showed the presence of a single immunoreactive band of a size similar to murine ACT. A library screening resulted in the isolation and characterization of the complete cDNA which showed 88% homology with the mouse counterpart. The human ACT gene is composed of five coding exons, being the first untranslated, and the mRNA spans 835 nucleotides coding for a 284 amino acid protein. Expression studies by RT-PCR confirmed that ACT is present in normal human testis. The human ACT gene is localized on the chromosome 6.

CREM, a master-switch of the transcriptional cascade in male germ cells.

In eukaryotes, transcriptional regulation upon stimulation of the adenylyl cyclase signalling pathway is mediated by a family of cAMP-responsive nuclear factors. The CREB and CREM transcription factors are activated by phosphorylation of a key serine residue by kinase stimulated by cyclic AMP, calcium, growth factors and stress signals. Phosphorylation allows recruitment of CBP (CREB Binding Protein), a large co-activator that contacts the general transcriptional machinery. The CREM gene plays a key physiological and developmental role within the hypothalamic-pituitary-gonadal axis. CREM is highly expressed in postmeiotic cells upon a striking developmental switch regulated by the pituitary hormone FSH. CREM-mutant mice generated by homologous recombination reveal that spermatogenesis stops at the first step of spermiogenesis. Late spermatids are completely absent while there is a significant increase in apoptotic germ cells. Mutant male mice completely lack spermatozoa, a phenotype reminiscent of cases of human infertility. Interestingly, in male germ cells, CREM is not phosphorylated but associates with ACT, a member of the LIM-only class of proteins that has intrinsic transcriptional activity. Thus, in some circumstance, CREM can bypass the classical requirement for phosphorylation and association with CBP.

A family of LIM-only transcriptional coactivators: tissue-specific expression and selective activation of CREB and CREM.

Transcription factors of the CREB family control the expression of a large number of genes in response to various signaling pathways. Regulation mediated by members of the CREB family has been linked to various physiological functions. Classically, activation by CREB is known to occur upon phosphorylation at an essential regulatory site (Ser133 in CREB) and the subsequent interaction with the ubiquitous coactivator CREB-binding protein (CBP). However, the mechanism by which selectivity is achieved in the identification of target genes, as well as the routes adopted to ensure tissue-specific activation, remains unrecognized. We have recently described the first tissue-specific coactivator of CREB family transcription factors, ACT (activator of CREM in testis). ACT is a LIM-only protein which associates with CREM in male germ cells and provides an activation function which is independent of phosphorylation and CBP. Here we characterize a family of LIM-only proteins which share common structural organization with ACT. These are referred to as four-and-a-half-LIM-domain (FHL) proteins and display tissue-specific and developmentally regulated expression. FHL proteins display different degrees of intrinsic activation potential. They provide powerful activation function to both CREB and CREM when coexpressed either in yeast or in mammalian cells, specific combinations eliciting selective activation. Deletion analysis of the ACT protein shows that the activation function depends on specific arrangements of the LIM domains, which are essential for both transactivation and interaction properties. This study uncovers the existence of a family of tissue-specific coactivators that operate through novel, CBP-independent routes to elicit transcriptional activation by CREB and CREM. The future identification of additional partners of FHL proteins is likely to reveal unappreciated aspects of tissue-specific transcriptional regulation.

Oct-1 specifically binds the UEF4 site of the human AP1-regulated urokinase enhancer.

The inducible urokinase enhancer contains three essential elements: a combined PEA3/AP1 and a downstream AP1 site, separated by a 74-bp DNA region called COM (cooperation mediator), that is required for the synergism between the three sites. The 5' half of COM (uCOM) forms four retarded complexes with HeLa or HepG2 nuclear proteins (UEF1-4). We now demonstrate that the UEF4 complex is the transcription factor Oct-1. Because of functional redundancy of the UEF sites, single mutations in UEF4 have no phenotype; we have changed UEF4 from a low to a high affinity binding site for Oct-1. In vitro, this mutation increases the DNA binding of Oct-1 and disturbs the binding of the Prep-Pbx complexes to the nearby UEF3 site. In vivo, this mutation reduces the basal transcriptional activity of the urokinase enhancer, while not affecting its phorbol ester inducibility. This is in keeping with the effect of the deletions of the COM region, which result in an increase in the basal level and, as a consequence, in the loss of 4beta-phorbol 12-myristate 13-acetate inducibility. Oct-1 therefore is not involved in the inducibility of the urokinase enhancer but only in determining its basal activity level.

Transcriptional regulation by cyclic AMP-responsive factors.

In eukaryotes, transcriptional regulation on stimulation of the adenylate cyclase signaling pathway is mediated by a family of cyclic AMP-responsive nuclear factors, including CREB, CREM, and ATF-1. These factors contain the basic domain/leucine zipper motifs and bind as dimers to cAMP-responsive elements (CREs). The activation function of CRE-binding proteins is modulated by phosphorylation by several kinases and is mediated by coactivators such as CBP and p300. Activation might also be independent of CBP and phosphorylation in some specific cell types, such as male germ cells, wherein the protein ACT confers a powerful activation function to CREM. The inducible cAMP early repressor (ICER) protein is the only inducible member of this family. The induction of this powerful repressor is likely to be important for the transient nature of cAMP-induced gene expression. CRE-binding proteins have been found to play an important role in the physiology of the pituitary gland, in regulating spermatogenesis, in the response to circadian rhythms, and in the molecular basis of memory.

Increased systemic oxidative stress after elective endarterectomy: relation to vascular healing and remodeling.

It has been reported that systemic and local redox state may have an important role in the functional and organic changes characterizing the process of vascular response to injury. Carotid endarterectomy to remove atherosclerotic plaque is followed by a long lasting healing and remodeling process that can be carefully followed over time with noninvasive ultrasonography. Plasma vitamin C concentration and native LDL (n-LDL) content in lipid peroxides, vitamin E, beta-carotene, and lycopene as well as LDL susceptibility to peroxidation were assessed in 45 patients undergoing elective endarterectomy for internal carotid stenosis, at baseline, 24 hours, 3 and 15 days, and 1 month after surgery. Serial duplex scans were performed in all patients postoperatively and 3, 6, and 12 months. The changes in far wall thickness (FW) and % renarrowing from postoperatively to 12 months were used as remodeling indices. Plasma antioxidant vitamins and lag-phase showed a sharp and significant decrease during the first 24-hours after surgery remaining unchanged until the third day, whereas, an opposite trend was evidenced for n-LDL content in lipid peroxides and serum ceruloplasmin. After the third day all the parameters returned progressively to baseline within one month from endarterectomy. Interestingly, the n-LDL lipid peroxide content, the serum ceruloplasmin and the plasma vitamin C concentration, measured at 24 and 3 days from surgery, were significantly associated to the change in % renarrowing from postoperatively to 12 months. The higher the LDL content in lipid peroxides, the higher the serum level of ceruloplasmin, the lower the plasma content in vitamin C and the higher the % of vessel renarrowing. In conclusion, carotid endarterectomy with atherosclerotic plaque removal is associated with an acute and prolonged increase in systemic oxidative stress that influences vascular healing and late luminal loss.

Role of distinct mitogen-activated protein kinase pathways and cooperation between Ets-2, ATF-2, and Jun family members in human urokinase-type plasminogen activator gene induction by interleukin-1 and tetradecanoyl phorbol acetate.

We have investigated the in vivo and in vitro regulation of the human urokinase-type plasminogen activator (uPA) gene by interleukin-1 (IL-1) and analyzed the transcription factors and signalling pathways involved in the response of the -2.0-kb uPA enhancer to IL-1 induction and to tetradecanoyl phorbol acetate (TPA) induction. Mutational analysis showed the cooperative activity of the Ets-binding site (EBS) and the two AP-1 elements of the enhancer. The results reveal that the EBS is required for the response to both inducers mediated by Ets-2, which is regulated at a level subsequent to DNA binding, by an IL-1- and phorbol ester-inducible transactivation domain. Both the IL-1 and the TPA-mediated induction result in a drastic increase of AP-1 binding to the downstream site of the enhancer (uPA 3' TPA-responsive element), while a mostly qualitative change, resulting from the interplay between ATF-2 homodimers and c-Jun-ATF-2 heterodimers, takes place at the upstream AP-1 element. The analysis of two distinct mitogen-activated protein kinase pathways shows that stress-activated protein kinase-Jun N-terminal kinase activation, resulting in the phosphorylation of ATF-2, c-Jun, and JunD, is required not only for the IL-1- but also for the TPA-dependent induction, while the extracellular signal-related kinase 1 (ERK-1) and ERK-2 activation is involved in the TPA- but not in the IL-1-dependent stimulation of the uPA enhancer.

Signaling routes to CREM and CREB: plasticity in transcriptional activation.

The CREB and CREM transcription factors are activated by phosphorylation of a key serine residue by kinases stimulated by cyclic AMP, Ca2+, growth factors and stress signals. Phosphorylation allows recruitment of CREB-binding protein (CBP), a large co-activator that contacts the general transcriptional machinery. Studies of the physiological roles played by CREB and CREM have uncovered novel routes of transcriptional activation. For example, in male germ cells CREM is not phosphorylated but associates with ACT, a member of the LIM-only class of proteins that has intrinsic transcriptional activity. Thus, in some circumstances, CREM can bypass the classical requirement for phosphorylation and association with CBP.

Blunted nocturnal fall in blood pressure and oxidative stress in men and women with essential hypertension.

Low-density lipoprotein oxidation and antioxidant vitamins E and C were investigated in dipper (nocturnal blood pressure fall > 10%) and nondipper (nocturnal blood pressure fall < 10%) hypertensives. We studied 40 dippers and 28 nondippers balanced for gender, age, and body mass index. Blood samples were drawn for lipid profile determination, assessment of thiobarbituric acid-reactive substances, and fluorescent products of lipid peroxidation in native low-density lipoprotein, evaluation of susceptibility to low-density lipoprotein oxidation in vitro (lag phase and propagation rate), and determination of low-density lipoprotein vitamin E and plasma vitamins E and C contents. Compared with dippers, nondippers had significantly higher thiobarbituric acid-reactive substances and fluorescent products of lipid peroxidation (0.63 +/- 0.1 v 0.77 +/- 0.08 nmol malondialdehyde/mg low-density lipoprotein protein, and 14.5 +/- 6 v 17.9 +/- 4 units of relative fluorescence/mg low-density lipoprotein protein, respectively, both P < .05), shorter lag phase (56 +/- 13 v 49 +/- 9 min, P < .05), and lower plasma vitamin C content (42 +/- 9 v 35 +/- 10 micromol/L, P < .05). When gender was taken into account, differences were not significant between dipper and nondipper men, whereas, compared with dipper women, nondipper women showed significantly higher thiobarbituric acid-reactive substances and fluorescent products of lipid peroxidation (0.56 +/- 0.1 v 0.77 +/- 0.07 nmol malondialdehyde/mg low-density lipoprotein protein, and 12.5 +/- 4 v 17.5 +/- 4.6 units of relative fluorescence/mg low-density lipoprotein protein, respectively, both P < .05), shorter lag phase (62.5 +/- 11 v 49 +/- 9.5 min, P < .05), and lower plasma vitamin C content (44.9 +/- 10 v 34.7 +/- 10.8 micromol/L, P < .05). Given the role of low-density lipoprotein oxidation in the pathogenesis of atherosclerosis and that of vitamin C in protecting against it, our data suggest that nondippers, especially among women, show higher atherogenic risk than dippers.

Gamma-glutamyl transpeptidase-dependent iron reduction and LDL oxidation--a potential mechanism in atherosclerosis.

BACKGROUND: gamma-Glutamyl transpeptidase (gamma-GT) is found in serum and in the plasma membranes of virtually all cell types. Its physiologic role is to initiate the hydrolysis of extracellular glutathione (GSH), a tripeptide in which cysteine lies between alpha-glycine and gamma-glutamate residues. Cysteine and other thiol compounds are known to promote LDL oxidation by reducing Fe(III) to redox active Fe(II); therefore, we sought to determine whether similar reactions can be sustained by GSH and influenced by gamma-GT. METHODS: Fe(III) reduction and LDL oxidation were studied by monitoring the formation bathophenanthroline-chelatable Fe(II) and the accumulation of thiobarbituric acid-reactive substances, respectively. Human atheromatous tissues were examined by histochemical techniques for the presence of oxidized LDL and their colocalization with cells expressing gamma-GT activity. RESULTS: A series of experiments showed that the gamma-glutamate residue of GSH affected interactions of the juxtaposed cysteine thiol with iron, precluding Fe(III) reduction and hence LDL oxidation. Both processes increased remarkably after addition of purified gamma-GT, which acts by removing the gamma-glutamate residue. GSH-dependent LDL oxidation was similarly promoted by gamma-GT associated with the plasma membrane of human monoblastoid cells, and this process required iron traces that can be found in advanced or late stage atheromas. Collectively, these findings suggested a possible role for gamma-GT in the cellular processes of LDL oxidation and atherogenesis. Histochemical analyses confirmed that this may be the case, showing that gamma-GT activity is expressed by macrophage-derived foam cells within human atheromas, and that these cells colocalize with oxidized LDL. CONCLUSIONS: Biochemical and histochemical correlates indicate that gamma-GT can promote LDL oxidation by hydrolyzing GSH into more potent iron reductants. These findings may provide mechanistic clues to the epidemiologic evidence for a possible correlation between persistent elevation of gamma-GT and the risk of fatal reinfarction in patients with ischemic heart disease.

CBP-independent activation of CREM and CREB by the LIM-only protein ACT.

Transcriptional activation by CREB and CREM requires phosphorylation of a serine residue within the activation domain (Ser 133 in CREB; Ser 117 in CREM) which as a result interacts with the coactivator CBP. The activator CREM is highly expressed in male germ cells and is required for post-meiotic gene expression. Using a two-hybrid screen, we have isolated a testis-derived complementary DNA encoding a protein that we term ACT (for activator of CREM in testis), a LIM-only protein which specifically associates with CREM. ACT is expressed coordinately with CREM in a tissue- and developmentally regulated manner. It strongly stimulates CREM transcriptional activity in yeast and mammalian cells and contains an intrinsic activation function. As ACT bypasses the classical requirements for activation, namely phosphorylation of Ser 117 and interaction with CBP, it represents a new route for transcriptional activation by CREM and CREB. ACT may define a previously undiscovered class of tissue-specific coactivators whose function could be specific for distinct cellular differentiation programmes.