Identification of a Carcinoembryonic Antigen Gene Family in the Rat

The existence of a carcinoembryonic antigen (CEA)-like gene family in rat has been demonstrated through isolation and sequencing of the N- terminal domain exons of presumably five discrete genes (rnCGM1-5). This finding will allow for the first time the study of functional and clinical aspects of the tumor marker CEA and related antigens in an animal model. Sequence comparison with the corresponding regions of members of the human CEA gene family revealed a relatively low similarity at the amino acid level, which indicates rapid divergence of the CEA gene family during evolution and explains the lack of cross- reactivity of rat CEA-like antigens with antibodies directed against human CEA. The N-terminal domains of the rat CEA-like proteins show structural similarity to immunoglobulin variable domains, including the presence of hypervariable regions, which points to a possible receptor function of the CEA family members. Although so far only one of the five rat CEA-like genes could be shown to be transcriptionally active, multiple mRNA species derived from other members of the rat CEA-like gene family have been found to be differentially expressed in rat placenta and liver

Cloning of a Carcinoembryonic Antigen Gene Family Member Expressed in Leukocytes of Chronic Myeloid Leukemia Patients and Bone Marrow

The carcinoembryonic antigen (CEA) gene family belongs to the immunoglobulin superfamily and can be subdivided into the CEA and pregnancy-specific glycoprotein subgroups. The basic structure of the encoded proteins consists of, in addition to a leader, one IgV-like and 2, 3, or 6 IgC-like domains. These domains are followed by varying COOH-terminal regions responsible for secretion, transmembrane anchoring, or insertion into the membrane by a glycosyl phosphatidylinositol tail. Here we report on the characterization of CGM6, a new member of the CEA gene subgroup, by complementary DNA cloning. The deduced coding region comprises 349 amino acids and consists of a leader, one IgV-like, two IgC-like domains, and a hydrophobic region, which is replaced by a glycosyl phosphatidylinositol moiety in the mature protein. CGM6 transcripts were only found thus far in leukocytes of chronic myeloid leukemia patients, in normal bone marrow, and in marginal amounts in normal granulocytes. The CGM6 gene product might, therefore, represent a myeloid marker. Analyses of CGM6 protein-expressing HeLa transfectants with monoclonal antibodies strongly indicate that the CGM6 gene codes for the CEA family member NCA-95

Mice Transgenic for the Human Carcinoembryonic Antigen Gene Maintain Its Spatiotemporal Expression Pattern

The tumor marker carcinoembryonic antigen (CEA) is predominantly expressed in epithelial cells along the gastrointestinal tract and in a variety of adenocarcinomas. As a basis for investigating its in vivo regulation and for establishing an animal model for tumor immunotherapy, transgenic mice were generated with a 33-kilobase cosmid clone insert containing the complete human CEA gene and flanking sequences. CEA was found in the tongue, esophagus, stomach, small intestine, cecum, colon, and trachea and at low levels in the lung, testis, and uterus of adult mice of independent transgenic strains. CEA was first detected at day 10.5 of embryonic development (embryonic day 10.5) in primary trophoblast giant cells and was found in the developing gut, urethra, trachea, lung, and nucleus pulposus of the vertebral column from embryonic day 14.5 onwards. From embryonic day 16.5 CEA was also visible in the nasal mucosa and tongue. Because this spatiotemporal expression pattern correlates well with that known for humans, it follows that the transferred genomic region contains all of the regulatory elements required for the correct expression of CEA. Furthermore, although mice apparently lack an endogenous CEA gene, the entire repertoire of transcription factors necessary for correct expression of the CEA transgene is conserved between mice and humans. After tumor induction, these immunocompetent mice will serve as a model for optimizing various forms of immunotherapy, using CEA as a target antigen

Chromosomal Localization of the Carcinoembryonic Antigen Gene Family and Differential Expression in Various Tumors

Carcinoembryonic antigen (CEA) is a glycoprotein which is important as a tumor marker for a number of human cancers. It is a member of a gene family comprising about 10 closely related genes. In order to characterize mUNAs transcribed from individual genes we have identified by DNA and RNA hybridization experiments, gene-specific sequences from the 3 ' noncoding regions of CEA, and of nonspecific cross-reacting antigen (NCA) mRNAs, which have been recently cloned. With these probes, CEA mRNAs with lengths of 3.5 and 3.0 kilobases and an NCA mRNA species of 2.5 kilobases were identified in various human tumors. A 2.2-kilobase mRNA species, however, could only be detected in leu kocytes of patients with chronic myeloid leukemia by hybridization with a probe from the immunoglobulin-like repeat domain of CEA. This region is known to be very similar among the various members of the CEA gene family, and indeed the probe hybridizes with all four mRNA species. In situ hybridization with a cross-hybridizing probe from the NCA gene localized the members of the CEA gene family to the short and to the long arm of chromosome 19. In addition, a CEA cDNA probe was found to hybridize to the long arm of chromosome 19 only

Cloning of the Complete Gene for Carcinoembryonic Antigen

Carcinoembryonic antigen (CEA) is a widely used tumor marker, especially in the surveillance of colonic cancer patients. Although CEA is also present in some normal tissues, it is apparently expressed at higher levels in tumorous tissues than in corresponding normal tissues. As a first step toward analyzing the regulation of expression of CEA at the transcriptional level, we have isolated and characterized a cosmid clone (cosCEA1), which contains the entire coding region of the CEA gene. A close correlation exists between the exon and deduced immunoglobulin-like domain borders. We have determined a cluster of transcriptional starts for CEA and the closely related nonspecific cross-reacting antigen (NCA) gene and have sequenced their putative promoters. Regions of sequence homology are found as far as approximately 500 nucleotides upstream from the translational starts of these genes, but farther upstream they diverge completely. In both cases we were unable to find classic TATA or CAAT boxes at their expected positions. To characterize the CEA and NCA promoters, we carried out transient transfection assays with promoter-indicator gene constructs in the CEA-producing adenocarcinoma cell line SW403, as well as in nonproducing HeLa cells. A CEA gene promoter construct, containing approximately 400 nucleotides upstream from the translational start, showed nine times higher activity in the SW403 than in the HeLa cell line. This indicates that cis-acting sequences which convey cell type-specific expression of the CEA gene are contained within this region

Investigation of a Limited but Explosive COVID-19 Outbreak in a German Secondary School

The role of schools as a source of infection and driver in the coronavirus-pandemic has been controversial and is still not completely clarified. To prevent harm and disadvantages for children and adolescents, but also adults, detailed data on school outbreaks is needed, especially when talking about open schools employing evidence-based safety concepts. Here, we investigated the first significant COVID-19 school outbreak in Hamburg, Germany, after the re-opening of schools in 2020. Using clinical, laboratory, and contact data and spatial measures for epidemiological and environmental studies combined with whole-genome sequencing (WGS) analysis, we examined the causes and the course of the secondary school outbreak. The potential index case was identified by epidemiological tracking and the lessons in classrooms with presumably high virus spreading rates and further infection chains in the setting. Sequence analysis of samples detected one sample of a different virus lineage and 25 virus genomes with almost identical sequences, of which 21 showed 100% similarity. Most infections occurred in connection with two lesson units of the primary case. Likely, 31 students (12–14 years old), two staff members, and three family members were infected in the school or the typical household. Sequence analysis revealed an outbreak cluster with a single source that was epidemiologically identified as a member of the educational staff. In lesson units, two superspreading events of varying degrees with airborne transmission took place. These were influenced by several parameters including the exposure times, the use of respiratory masks while speaking and spatial or structural conditions at that time.Peer Reviewe

Investigation of a Limited but Explosive COVID-19 Outbreak in a German Secondary School

The role of schools as a source of infection and driver in the coronavirus-pandemic has been controversial and is still not completely clarified. To prevent harm and disadvantages for children and adolescents, but also adults, detailed data on school outbreaks is needed, especially when talking about open schools employing evidence-based safety concepts. Here, we investigated the first significant COVID-19 school outbreak in Hamburg, Germany, after the re-opening of schools in 2020. Using clinical, laboratory, and contact data and spatial measures for epidemiological and environmental studies combined with whole-genome sequencing (WGS) analysis, we examined the causes and the course of the secondary school outbreak. The potential index case was identified by epidemiological tracking and the lessons in classrooms with presumably high virus spreading rates and further infection chains in the setting. Sequence analysis of samples detected one sample of a different virus lineage and 25 virus genomes with almost identical sequences, of which 21 showed 100% similarity. Most infections occurred in connection with two lesson units of the primary case. Likely, 31 students (12–14 years old), two staff members, and three family members were infected in the school or the typical household. Sequence analysis revealed an outbreak cluster with a single source that was epidemiologically identified as a member of the educational staff. In lesson units, two superspreading events of varying degrees with airborne transmission took place. These were influenced by several parameters including the exposure times, the use of respiratory masks while speaking and spatial or structural conditions at that time

Enrichment of Leishmania donovani ATP-binding proteins using a staurosporine capture compound.

International audienceTrypanosomatid parasites of the genus Leishmania cause severe human diseases collectively termed leishmaniasis. Parasite ATP-binding proteins have emerged as potent targets for chemotherapeutic intervention. However, many parasite-specific ATP-binding proteins may escape current efforts in drug target identification, validation and deconvolution due to the lack of sequence conservation and functional annotation of these proteins in early branching eukaryotic trypanosomatids. Here, we selectively enriched for ATP-binding proteins from Leishmania donovani axenic promastigote and amastigote total protein extracts utilizing a Capture Compound™ (CC) linked to the ATP-competitive inhibitor staurosporine. As judged by in-gel kinase activity assay and competitive inhibition with free staurosporine, the CC specifically enriched for parasite phosphotransferases. Comparative nanoLC-MS(n) analysis identified 70 captured proteins, including 24 conserved protein kinases, and 32 hypothetical proteins with potential ATP-binding function. We identified conserved signature sequence motifs characteristic for staurosporine-binding protein kinases, and identified the hypothetical proteins LinJ.20.0280 and LinJ.09.1630 as novel ATP-binding proteins. Thus, functional enrichment procedures such as described here, combined with bio-informatics analyses and activity assays, provide powerful tools for the discovery of parasite-specific ATP-binding proteins that escape homology-based identification, which can be subsequently targeted for pharmacological intervention.BIOLOGICAL SIGNIFICANCE: Functional enrichment using a Capture Compound™ linked to the ATP-competitive inhibitor staurosporine provides a powerful new tool for the discovery of parasite-specific ATP-binding proteins that escape homology-based identification, which can be subsequently targeted for pharmacological intervention