Molecular characterization of a human matrix attachment region epigenetic regulator.

Matrix attachment regions (MAR) generally act as epigenetic regulatory sequences that increase gene expression, and they were proposed to partition chromosomes into loop-forming domains. However, their molecular mode of action remains poorly understood. Here, we assessed the possible contribution of the AT-rich core and adjacent transcription factor binding motifs to the transcription augmenting and anti-silencing effects of human MAR 1-68. Either flanking sequences together with the AT-rich core were required to obtain the full MAR effects. Shortened MAR derivatives retaining full MAR activity were constructed from combinations of the AT-rich sequence and multimerized transcription factor binding motifs, implying that both transcription factors and the AT-rich microsatellite sequence are required to mediate the MAR effect. Genomic analysis indicated that MAR AT-rich cores may be depleted of histones and enriched in RNA polymerase II, providing a molecular interpretation of their chromatin domain insulator and transcriptional augmentation activities

DNA aptamers against the MUC1 tumour marker: design of aptamer–antibody sandwich ELISA for the early diagnosis of epithelial tumours

Aptamers are functional molecules able to bind tightly and selectively to disease markers, offering great potential for applications in disease diagnosis and therapy. MUC1 is a well-known tumour marker present in epithelial malignancies and is used in immunotherapeutic and diagnostic approaches. We report the selection of DNA aptamers that bind with high affinity and selectivity an MUC1 recombinant protein containing five repeats of the variable tandem repeat region. Aptamers were selected using the SELEX methodology from an initial library containing a 25-base-long variable region for their ability to bind to the unglycosylated form of the MUC1 protein. After ten rounds of in vitro selection and amplification, more than 90% of the pool of sequences consisted of target-binding molecules, which were cloned, sequenced and found to share no sequence consensus. The binding properties of these aptamers were quantified using ELISA and surface plasmon resonance. The lead aptamer sequence was subsequently used in the design of an aptamer–antibody hybrid sandwich ELISA for the identification and quantification of MUC1 in buffered solutions. Following optimisation of the operating conditions, the resulting enzyme immunoassay displayed an EC50 value of 25 μg/ml, a detection limit of 1 μg/ml and a linear range between 8 and 100 μg/ml for the MUC1 five tandem repeat analyte. In addition, recovery studies performed in buffer conditions resulted in averaged recoveries between 98.2 and 101.7% for all spiked samples, demonstrating the usability of the aptamer as a receptor in microtitre-based assays. Our results aim towards the formation of new diagnostic assays against this tumour marker for the early diagnosis of primary or metastatic disease in breast, bladder and other epithelial tumours

Cystic Echinococcoses in Mongolia: Molecular Identification, Serology and Risk Factors

BACKGROUND: Cystic echinococcosis (CE) is a globally distributed cestode zoonosis that causes hepatic cysts. Although Echinococcus granulosus sensu stricto (s.s.) is the major causative agent of CE worldwide, recent molecular epidemiological studies have revealed that E. canadensis is common in countries where camels are present. One such country is Mongolia. METHODOLOGY/PRINCIPAL FINDINGS: Forty-three human hepatic CE cases that were confirmed histopathologically at the National Center of Pathology (NCP) in Ulaanbaatar (UB) were identified by analysis of mitochondrial cox 1 gene as being caused by either E. canadensis (n=31, 72.1%) or E. granulosus s.s. (n=12, 27.9%). The majority of the E. canadensis cases were strain G6/7 (29/31, 93.5%). Twenty three haplotypes were identified. Sixteen of 39 CE cases with data on age, sex and province of residence were citizens of UB (41.0%), with 13 of the 16 cases from UB caused by E. canadensis (G6/7) (81.3%). Among these 13 cases, nine were children (69.2%). All pediatric cases (n  =  18) were due to E. canadensis with 17 of the 18 cases (94.4%) due to strain G6/7. Serum samples were available for 31 of the 43 CE cases, with 22 (71.0%) samples positive by ELISA to recombinant Antigen B8/1 (rAgB). Nine of 10 CE cases caused by E. granulosus s.s. (90.0%) and 13 of 20 CE cases by E. canadensis (G6/7) (65.0%) were seropositive. The one CE case caused by E. canadensis (G10) was seronegative. CE cases caused by E. granulosus s.s. showed higher absorbance values (median value 1.131) than those caused by E. canadensis (G6/7) (median value 0.106) (p  =  0.0137). CONCLUSION/SIGNIFICANCE: The main species/strains in the study population were E. canadenis and E. granulossus s.s. with E. canadensis the predominant species identified in children. The reason why E. canadensis appears to be so common in children is unknown