Serum neurofilament light chain at time of diagnosis is an independent prognostic factor of survival in amyotrophic lateral sclerosis.

BACKGROUND AND PURPOSE: The prognostic value of serum neurofilament light chain (sNfL), a biomarker of neurodegeneration, compared to other prognostic factors of amyotrophic lateral sclerosis (ALS) at the time of diagnosis, remains unclear. METHODS: Sera from ALS patients were prospectively collected at the first diagnostic visit in our centre. sNfL levels were determined by single molecule array in 207 ALS patients and in 21 healthy controls. The prognostic value of sNfL was compared with that of other known clinical prognostic factors using a Cox regression model and multivariate analysis. RESULTS: Serum neurofilament light chain levels were higher in ALS patients than in controls (P < 0.0001). Seven parameters were predictive of death in ALS: older age, bulbar onset, higher ALS Functional Rating Scale revised (ALSFRS-R) score, greater weight loss, lower maximal inspiratory pressure, forced vital capacity and higher sNfL levels. A Cox regression model showed that sNfL (P < 0.0001), weight loss (P = 0.040) and site at onset (P = 0.048) were independent predictive factors of death. In a sub-cohort restricted to 139 patients with complete spirometry data, sNfL level (P < 0.005) and forced vital capacity (P = 0.022) were independent factors predictive of death. In a subgroup of 142 patients in whom ALSFRS-R score was available at several time points, sNfL levels positively correlated with ALSFRS-R rate of decline (r = 0.571, P < 10-12 ). CONCLUSIONS: Higher sNfL concentration is a strong and independent prognostic factor of death in ALS as early as the time of diagnosis.

Cranberry capsules to prevent nosocomial urinary tract bacteriuria after pelvic surgery: a randomised controlled trial.

OBJECTIVE: To evaluate whether cranberries are able to prevent postoperative urinary bacteriuria in patients undergoing pelvic surgery and receiving transurethral catheterisation. DESIGN: Randomised, double-blind, placebo-controlled trial. SETTINGS: French tertiary Care centre, University Hospital. POPULATION: A total of 272 women undergoing pelvic surgery aged 18 or older. METHODS: Participants undergoing pelvic surgery were randomised to 36 mg cranberry (proanthocyanidins, PAC) or placebo once daily for 10 days. Statistical analysis was performed by a chi-square test. MAIN OUTCOME MEASURES: The primary and secondary outcomes were postoperative bacteriuria, defined by a positive urine culture, within the first 15 and 40 days, respectively. RESULTS: Two hundred and fifty-five participants received the intended treatment: 132 (51.8%) received PAC and 123 (48.2%) received placebo. There were no significant differences in baseline demographics, intra-operative characteristics or duration and type of catheterisation between the two groups. PAC prophylaxis did not reduce the risk of bacteriuria treatment within 15 days of surgery [27% bacteriuria with PAC compared with 25% bacteriuria with placebo: relative risk 1.05, 95% CI 0.78-1.4, P = 0.763). The same result was observed on day 40. Bacteriuria occurred more often in older women with increased length of catheterisation. CONCLUSION: Immediate postoperative prophylaxis with PAC does not reduce the risk of postoperative bacteriuria in patients receiving short-term transurethral catheterisation after pelvic surgery. TWEETABLE ABSTRACT: PAC prophylaxis does not reduce the risk of postoperative bacteriuria in patients undergoing pelvic surgery.

Link between nasal carriage of Staphylococcus aureus and infected diabetic foot ulcers.

AIMS: Nasal carriage of Staphylococcus aureus in diabetic patients may be a risk factor for diabetic foot lesion infections. The aims of this study were to compare the genotypic profiles of S. aureus strains isolated from nares and diabetic foot ulcers (DFUs) using microarray technology. METHODS: Patients were included if they were admitted for diabetic foot infection (DFI) at any of three diabetology departments of Montpellier and Nîmes University Hospitals between 1 September 2010 to 30 June 2012. All S. aureus isolates were analyzed using oligonucleotides arrays; S. aureus resistance and virulence genes were determined and each isolate was affiliated to a clonal complex. RESULTS: The prevalence of S. aureus nasal carriage among the 276 included patients was 39.5% (n=109), while 36.6% (n=101) had S. aureus at both sites (nares and foot wounds) and, of these patients, 65.3% of patients harboured the same strain at both sites. In addition, the spread of the methicillin-resistant S. aureus (MRSA) ST398 clone in DFI and its tropism for bone were also further confirmed. CONCLUSION: These findings appear to provide new arguments in favour of the systematic detection of nasal S. aureus carriage to anticipate the management of DFI.

Chitinase 3-like proteins as diagnostic and prognostic biomarkers of multiple sclerosis.

BACKGROUND: Despite sensitivity of MRI to diagnose multiple sclerosis (MS), prognostic biomarkers are still needed for optimized treatment. OBJECTIVE: The objective of this paper is to identify cerebrospinal fluid (CSF) diagnostic biomarkers of MS using quantitative proteomics and to analyze their expression at different disease stages. METHODS: We conducted differential analysis of the CSF proteome from control and relapsing-remitting MS (RRMS) patients followed by verification by ELISA of candidate biomarkers in CSF and serum in control, clinically isolated syndrome (CIS), RRMS and progressive MS (PMS) patients. RESULTS: Twenty-two of the 527 quantified proteins exhibited different abundances in control and RRMS CSF. These include chitinase 3-like protein 1 (CHI3L1) and 2 (CHI3L2), which showed a strong expression in brain of MS patients, especially in astrocytes and microglial cells from white matter plaques. CSF and serum CHI3L1 levels increased with the disease stage and CIS patients with high CSF (>189 ng/ml) and serum (>33 ng/ml) CHI3L1 converted more rapidly to RRMS (log rank test, p < 0.05 and p < 0.001, respectively). In contrast, CSF CHI3L2 levels were lower in PMS than in RRMS patients. Accordingly, CSF CHI3L1/CHI3L2 ratio accurately discriminated PMS from RRMS. CONCLUSIONS: CSF CHI3L1 and CHI3L2 and serum CHI3L1 might help to define MS disease stage and have a prognostic value in CIS.

Single-shot intraoperative local anaesthetic infiltration does not reduce morphine consumption after total hip arthroplasty: a double-blinded placebo-controlled randomized study.

BACKGROUND: The infiltration of local anaesthetic (LA), ketorolac, and epinephrine has been suggested to be effective for analgesia after total hip arthroplasty (THA). The part of action of each component of the mixture remains unclear. We investigated the contribution of infiltration of ropivacaine alone on the morphine consumption during the first 24 h after surgery. METHODS: Sixty patients undergoing primary THA were included in this prospective randomized double-blinded placebo-controlled trial, after IRB approval and informed consent. Surgical and general anaesthetic management were standardized. At the end of surgery, 80 ml of ropivacaine 0.2% (160 mg) or saline was infiltrated. The primary endpoint was morphine consumption 24 h after surgery. The secondary endpoints were: visual analogue scale scores and opioid side-effects at H2, H4, H8, H12, H24, D1, D2, D3, D4, D5, rehabilitation programme progress, chronic pain level, analgesic consumption, and surgical result at 3 months and 1 yr after surgery. The observation period was 1 yr. RESULTS: Groups were similar for patient characteristic and perioperative characteristics. The ropivacaine wound infiltration did not reduce morphine consumption at 24 h [median (25th and 75th inter-quartile) 27 (17-37) mg in the ropivacaine group vs 24 (18-34) mg in the placebo group, P=0.51] or its side-effects. No effect was found on rehabilitation progress or chronic pain after 3 months or 1 yr, but these were not the main endpoints of the study. CONCLUSIONS: Ropivacaine infiltration alone did not reduce morphine consumption at 24 h after operation nor did it improve postoperative rehabilitation.

Accuracy of prenatal three-dimensional ultrasound in the diagnosis of cleft hard palate when cleft lip is present.

OBJECTIVE: To investigate the accuracy of prenatal axial three-dimensional (3D) ultrasound in predicting the absence or presence of cleft palate in the presence of cleft lip. METHODS: Between March 2005 and January 2009, there were 81 cases with a prenatal two-dimensional (2D) ultrasound screening diagnosis of unilateral or bilateral cleft lip at 22-25 weeks of gestation referred to our tertiary care center. Of these, 79 fetuses were included in this prospective study and two were excluded. Axial 3D ultrasound imaging of the fetal palate was performed and the diagnoses were compared with clinical findings at delivery. The frequencies of intact and cleft palate, the degree of association between the prenatal predictions and postnatal findings and the probability of detection of cleft lip and palate were determined. RESULTS: Of 79 prenatal predictions, 77 (97%) were correct and the association between the prenatal predictions and postnatal findings was strong. The sensitivity for detection of cleft lip and palate within this high-risk population was 100% and the specificity was 90%. In one of the excluded cases, the palate could not be visualized due to a fetal prone position. There were chromosomal anomalies in 4% of cases and associated structural or growth anomalies in 23%, termination of pregnancy was carried out in 4% and intrauterine fetal demise occurred in 3%. CONCLUSION: Axial 3D ultrasound of the fetal palate has high accuracy in identifying prenatal cleft palate when cleft lip is diagnosed at mid-trimester 2D ultrasound screening.

Validation of a fully automatic photoplethysmographic device for toe blood pressure measurement.

OBJECTIVES: This study was designed to assess the accuracy and reliability of a new, portable, fully automated photoplethysmography (PPG) device for toe blood pressure (TBP) measurement. DESIGN: Within-subject comparison with conventional laser Doppler (LD) measurement. MATERIALS AND METHODS: Four TBP measurements were performed on both lower limbs, alternatively with LD and PPG in 200 patients recruited at the Nîmes University Hospital. Reproducibility was assessed by the intraclass correlation coefficient (ICC). The concordance between the two methods was evaluated by Lin's concordance correlation coefficient (CCC), in the whole population as well as in comorbidity subgroups. A potential bias was investigated with the Bland and Altman method. RESULTS: The ICC was 0.887 (95% confidence interval (CI) 0.852-0.913) and 0.893 (0.860-0.918) on the right side (n = 193), 0.905 (0.875-0.928) and 0.898 (0.866-0.922) on the left side (n = 188) for PPG and LD measurements, respectively. The CCC was 0.913 (0.885-0.934) on the right side and 0.915 (0.888-0.937) on the left side, and remained >0.8 regardless of co-morbidities. CONCLUSIONS: This new, fully automatic, photoplethysmographic device yielded reliable TBP measurements and showed good agreement with the reference LD system over a wide range of values.

Association between caries experience and body mass index in 12-year-old French children.

The prevalence of overweight and obesity reached 19.7% in 12-year-old French children in the year 2005. Recently, nationwide programs have been broadly implemented in France to reduce the overconsumption of sugars, salt and fat. The aims of this study were to assess the distribution of body mass index (BMI) and D(3+4)MFT index in a sample of 12-year-old French children, and to compare several regression models in order to analyze the association between these two indices. A cross-sectional study was conducted in Montpellier, France, and the height, weight, D(3+4)MFT, sugar and soft drink consumption were recorded in a randomly selected sample of 835 schoolchildren. In order to analyze the association between BMI and DMFT, four models of regression were tested: logistic, Poisson, zero-inflated Poisson (ZIP) and zero-inflated negative binomial (ZINB). The mean BMI was 18.9 for the whole sample and the corresponding DMFT value was 1.47. The caries prevalence was 51.7%. The best fitted models for testing the association between BMI and DMFT were ZIP and ZINB models. They showed a significant association between DMFT and sugar consumption, but not with BMI. As a result of the best fitted models (ZIP and ZINB), where BMI was not statistically associated with DMFT, we conclude, within the limits of a cross-sectional survey, that there is no association between these two variables.

Assessment and predictor determination of indoor aldehyde levels in Paris newborn babies' homes.

UNLABELLED: Exposure to indoor chemical air pollutants expected to be potentially involved in allergic respiratory diseases in infants is poorly documented. A specific environmental investigation included in a birth cohort study was carried out to first assess indoor airborne aldehyde levels, using passive devices and their variability within 1 year (1, 6, 9 and 12 months) in the bedroom of 196 Paris infants, and second, to identify predictors for aldehyde concentrations using interviewer administered questionnaires about housing factors. Comfort parameters and carbon dioxide levels were measured simultaneously. Aldehydes were detected in almost all dwellings and geometric mean levels (geometric standard deviation) at the first visit were respectively for formaldehyde, acetaldehyde, hexanal, and pentanal 19.4 (1.7) microg/m(3), 8.9 (1.8) microg/m(3), 25.3 (3.1) microg/m(3), 3.7 (2.3) microg/m(3), consistent with earlier published results. Generalized Estimating Equation multivariate analyses showed that, apart from comfort parameters, aeration and season, the main indoor aldehyde sources were either continuous (building materials and coverings especially when they were new) or discontinuous (smoking, use of air fresheners and cleaning products, DIY etc...). Finally, the data collected by questionnaires should be sufficient to enable us to classify each infant in our cohort study according to his/her degree of exposure to the main aldehydes. PRACTICAL IMPLICATIONS: This analysis contributed to document indoor aldehyde levels in Parisian homes and to identify factors determining these levels. In the major part of newborn babies' homes, indoor formaldehyde levels were above the guideline value of 10 microg/m(3) proposed by the French Agency for Environmental and Occupational Health Safety for long-term exposure. Given this result, it is essential to study the health impact of exposure to aldehydes especially formaldehyde on the incidence of respiratory and allergic symptoms, particularly during the first months of life.

[Hematocrit measurement: comparison of conductimetry to microcentrifugation]. / Mesure de l'hématocrite: comparaison de la conductimétrie à la microcentrifugation.

In general, blood gas analysers can also determine the value of haematocrite by measuring the blood's conductivity. The question to ask is whether this value is reliable. In this study, hematocrit obtained via conductivity from 6 different pieces of equipment were compared with those measured using the gold standard method, which is microcentrifugation. By interpreting the results of 320 arterial blood samples taken in the intensive care unit DAR "B" we can see that the reliability between two measurements on the same piece of equipment is very good, in general > 0.95 whatever the equipment. The reliability between the means of the two measurements and the gold standard is slightly lower but remains very satisfactory, most often between 0.8 and 0.9. The Gem Premier 3000 (IL) analyser and the Roche OMNI S gave the best reliability compared with centrifugation. The Spearman coefficients between the mean values of the analysers and those of centrifugation were high, with the exception of the Rapidpoint 405. They are all statistically different from zero (p<0.0001).

Plasma proteasome level is a reliable early marker of malignant transformation of liver cirrhosis.

BACKGROUND: Proteasomes are the main non-lysosomal proteolytic structures which regulate crucial cellular processes. Circulating proteasome levels can be measured using an ELISA test and can be considered as a tumour marker in several types of malignancy. Given that there is no sensitive marker of hepatocellular carcinoma (HCC) in patients with cirrhosis, we measured plasma proteasome levels in 83 patients with cirrhosis (33 without HCC, 50 with HCC) and 40 controls. METHODS AND RESULTS: Patients with HCC were sub-classified into three groups according to tumour mass. alpha-Fetoprotein (AFP) was also measured. Plasma proteasome levels were significantly higher in patients with HCC compared to controls (4841 (SEM 613) ng/ml vs 2534 (SEM 187) ng/ml; p<0.001) and compared to patients with cirrhosis without HCC (2077 (SEM 112) ng/ml; p<0.001). This difference remained significant when the subgroup of patients with low tumour mass (proteasome level 3970 (SEM 310) ng/ml, p<0.001) was compared to controls and patients with cirrhosis without HCC. Plasma proteasome levels were independent of the cause of cirrhosis and were weakly correlated with AFP levels. With a cut-off of 2900 ng/ml, diagnostic specificity for HCC was 97% with a sensitivity of 72%, better than results obtained with AFP. Diagnostic relevance of plasma proteasome measurement was also effective in low tumour mass patients (sensitivity 76.2% vs 57.1% for AFP). CONCLUSION: The plasma proteasome level is a reliable marker of malignant transformation in patients with cirrhosis, even when there is a low tumour mass.

Indoor airborne endotoxin assessment in homes of Paris newborn babies.

The aims of this study were first to assess airborne endotoxin levels in the dwellings of 162 newborns living in Paris twice during a 1-year period, and second, to identify predictors for endotoxin concentrations using questionnaire data in relation to housing factors and living conditions. Air samples were collected on a glass fiber filter in polystyrene filter holders, using a pump at a flow rate of 3.5 l/min for 24 h placed in the main room of the home. Endotoxin levels were measured using a chromogenic kinetic Limulus Amoebocyte Lysate test. Geometric means (geometric standard deviation) of airborne endotoxin levels at two different visits were respectively 0.509 (4.289) EU/m3 and 0.557 (3.029) EU/m3. Airborne endotoxin levels were significantly increased: (i) in cold season (P = 0.024), with (ii) the presence of visible cockroaches in the previous 12 months at home (P < 0.001), (iii) increased number of inhabitants per square meter (P = 0.012), (iv) the high frequency of cleaning with the floor cloths (P = 0.0014), and (v) the low frequency of vacuuming (P = 0.0045). This study provided for the first time airborne endotoxin levels issued from repeated measurements in Paris dwellings. PRACTICAL IMPLICATIONS This analysis contributed to identify a few factors that determined indoor airborne endotoxin levels. However, the predictive model including housing factors and living conditions poorly estimated endotoxin levels. Consequently, multiple samples and longer sampling periods might improve the estimate of long-term airborne endotoxin exposure especially its variability, in cohort studies.

A longitudinal study of the evolution of cognitive function and affective state in patients with amyotrophic lateral sclerosis.

OBJECTIVES: The study aimed to evaluate cognitive function and emotional reactivity in 18 patients with ALS, compared to 19 matched controls, and assess their evolution over a 12-month period. METHODS: 18 ALS patients and 19 matched controls were included, and assessed at inclusion, six months and twelve months later. Depression was evaluated with the Geriatric Depression Scale, and cognitive function with the Folstein Mini Mental State. A battery of psychometric tests (Wisconsin Card Sorting Test (WCST), the numerical Empan test, the Trail-making test, the Boston Naming Test, the 15-word Rey memory test, the Benton visual retention test and the Raven Progressive Matrix) was used to measure frontal processing and non-frontal function. Emotional reactivity was measured with the film-evoked emotions test. RESULTS: ALS patients were significantly more depressed than controls, as measured on the Geriatric Depression Scale, and depression increased over the study period. There was a very mild defect in cognitive function, and a performance deficit in the Trail-making test, a measure of frontal processing. These deficits, unlike neuromuscular function and depression, did not aggravate over the 12 months of the study. There was no observable change in non-frontal function. Emotional reactivity did not differ significantly between ALS patients and controls. CONCLUSIONS: This study provides further evidence for a mild defect in frontal cognitive processing in ALS patients that evolves only slowly, if at all, with time.

Point mutations identify a conserved region of the saccharomyces cerevisiae AFR1 gene that is essential for both the pheromone signaling and morphogenesis functions.

Mating pheromone receptors activate a G protein signal pathway that leads to the conjugation of the yeast Saccharomyces cerevisiae. This pathway also induces the production of Afr1p, a protein that negatively regulates pheromone receptor signaling and is required to form pointed projections of new growth that become the site of cell fusion during mating. Afr1p lacks strong similarity to any well-characterized proteins to help predict how it acts. Therefore, we investigated the relationship between the different functions of Afr1p by isolating and characterizing seven mutants that were defective in regulating pheromone signaling. The AFR1 mutants were also defective when expressed as fusions to STE2, the alpha-factor receptor, indicating that the mutant Afr1 proteins are defective in function and not in co-localizing with receptors. The mutant genes contained four distinct point mutations that all occurred between codons 254 and 263, identifying a region that is critical for AFR1 function. Consistent with this, we found that the corresponding region is very highly conserved in the Afr1p homologs from the yeasts S. uvarum and S. douglasii. In contrast, there were no detectable effects on pheromone signaling caused by deletion or overexpression of YER158c, an open reading frame with overall sequence similarity to Afr1p that lacks this essential region. Interestingly, all of the AFR1 mutants showed a defect in their ability to form mating projections that was proportional to their defect in regulating pheromone signaling. This suggests that both functions may be due to the same action of Afr1p. Thus, these studies identify a specific region of Afr1p that is critical for its function in both signaling and morphogenesis.

Combining mutations in the incoming and outgoing pheromone signal pathways causes a synergistic mating defect in Saccharomyces cerevisiae.

Mating pheromones stimulate Saccharomyces cerevisiae yeast cells to form a pointed projection that becomes the site of cell fusion during conjugation. To investigate the role of mating projections, we screened for mutations that enhanced the weak mating defect of MATa ste2-T326 cells that are defective in forming pointed projections. These cells are also 10-fold more sensitive to alpha-factor pheromone because ste2-T326 encodes truncated alpha-factor receptors that are not regulated properly. Mutations in AXL1, STE6 and FUS3 were identified in the screen. AXL1 was studied further because it is required for efficient a-factor pheromone production and for selecting the site for bud morphogenesis. Mutation of AXL1 did not enhance the morphogenesis or pheromone sensitivity defects of ste2-T326. Instead, the synergistic mating defect was apparently due to decreased a-factor production because the axl1Delta ste2-T326 cells mated well with a sst2 alpha mating partner that is supersensitive to a-factor. When combined with a wild-type mating partner, the ste2-T326 axl1Delta cells failed to mate because they did not lock cell walls, one of the earliest steps in conjugation. Analysis of axl1Delta in combination with other mutations that cause defects in morphogenesis or pheromone sensitivity (e.g. bar1, sst2, afr1) indicated that both phenotypes of ste2-T326 cells, supersensitivity to alpha-factor and the defect in forming pointed projections, contributed to the synergistic mating defect. We suggest a model that the synergistic mating defect is caused by the combined effects of ste2-T326 and axl1Delta on the presentation of a-factor to partner cells. Altogether, these results demonstrate an important linkage between the incoming and outgoing pheromone signals during the intercellular communication that promotes yeast mating.

AFR1 promotes polarized apical morphogenesis in Saccharomyces cerevisiae.

The G protein-coupled alpha-factor receptor promotes polarized growth toward a mating partner. alpha-Factor induces the expression of AFR1, which acts together with the receptor C terminus to promote normal morphogenesis. The function of AFR1 was investigated by engineering cells to constitutively express AFR1 without alpha-factor. Constitutive AFR1 expression caused cells to form elongated buds that demonstrate that AFR1 can also interact with the morphogenesis components that promote bud formation. A similar elongated bud phenotype is caused by mutation of the CDC3, CDC10, CDC11, and CDC12 genes, which encode putative filament proteins that form a ring at the bud neck. AFR1 may act directly on the filament proteins, since immunolocalization detected AFR1 at the bud neck and interaction of AFR1 and CDC12 was detected in the two-hybrid protein assay. AFR1 localized to the base of pheromone-induced projections. These results suggest that AFR1 and the putative filament proteins act together with the receptor to facilitate proper localization of components during mating.

Retinoic acid receptors initiate induction of the cytomegalovirus enhancer in embryonal cells.

Reactivation of latent virus is believed to result from a signal transduction event that induces immediate-early (IE) gene transcription. Evidence is presented that the major IE promoter (MIEP) of human cytomegalovirus (hCMV) is activated by physiological levels of retinoic acid (RA) in human embryonal carcinoma cells. Mutagenesis experiments localized in the MIEP enhancer, a retinoic acid-responsive element composed of a direct repeat separated by five nucleotides. Protein-DNA binding experiments revealed that this element functions as a specific target site for the direct interaction of nuclear receptor proteins for RA. These findings implicate the biologically active derivative of vitamin A (RA) as a potential modulator of hCMV pathogenesis in infants and immunocompromised adults.

A discrete cis element in the human immunodeficiency virus long terminal repeat mediates synergistic trans activation by cytomegalovirus immediate-early proteins.

The major immediate-early (IE) promoter of human cytomegalovirus directs the expression of several differentially spliced and polyadenylated mRNAs that encode isoformic proteins with apparent molecular masses of 55, 72, and 86 kDa. All of these proteins are potent transcriptional regulatory proteins. We are interested in the collateral interactions between human cytomegalovirus and human immunodeficiency virus (HIV) in the context of dual infection of a cell. The roles of the specific IE protein isoforms and their respective response elements involved in trans activation of the HIV long terminal repeat (LTR) are not known. Here we present evidence that major IE proteins IE86, IE72, and IE55 are capable of trans-activating the HIV LTR in a T-cell line, HUT-78. The IE55 isoform noncooperatively stimulates the HIV LTR in the presence of either isoform IE72 or IE86. Interactions between isoforms IE72 and IE86, however, result in strong synergistic activation of the LTR. Our results suggest that a specific 155-amino-acid protein domain that is unique for the IE86 protein participates in this synergic interaction. Point mutational analysis of the LTR identified a distinct cis-acting target site, located between nucleotide positions -174 and -163, that mediates exclusively synergistic trans activation by the IE72 and IE86 proteins. Finally, this study underscores the role of a cellular intermediate(s) for communicating the synergic interactions between two IE trans activators.