Correction: Identification and Functional Characterization of a Novel Monotreme- Specific Antibacterial Protein Expressed during Lactation
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PROVIDER: S-EPMC3692951 | biostudies-literature | 2013 Jan
REPOSITORIES: biostudies-literature
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Project description:Monotremes are the only oviparous mammals and exhibit a fascinating combination of reptilian and mammalian characters. They represent a component of synapsidal reproduction by laying shelled eggs which are incubated outside the mother's body. This is accompanied by a prototherian lactation process, marking them as representatives of early mammals. The only extant monotremes are the platypus, and the short- and long- beaked echidnas, and their distributions are limited to Australia and New Guinea. Apart for a short weaning period, milk is the sole source of nutrition and protection for the hatchlings which are altricial and immunologically naive. The duration of lactation in these mammals is prolonged relative to the gestational length and period of incubation of eggs. Much of the development of monotreme young occurs in the non-sterile ex-utero environment. Therefore the role of milk in the growth, development and disease protection of the young is of significant interest. By sequencing the cDNA of cells harvested from monotreme milk, we have identified a novel monotreme- specific transcript, and the corresponding gene was designated as the EchAMP. The expression profile of this gene in various tissues revealed that it is highly expressed in milk cells. The peptides corresponding to the EchAMP protein have been identified in a sample of echidna milk In silico analysis indicated putative antimicrobial potential for the cognate protein of EchAMP. This was further confirmed by in vitro assays using a host of bacteria. Interestingly, EchAMP did not display any activity against a commensal gut floral species. These results support the hypothesis of enhancement of survival of the young by antimicrobial bioactives of mammary gland origin and thus emphasize the protective, non- nutritional role of milk in mammals.
| S-EPMC3541144 | biostudies-literature
Project description:Monotremes (platypus and echidna) are the descendants of the oldest ancestor of all extant mammals distinguished from other mammals by mode of reproduction. Monotremes lay eggs following a short gestation period and after an even briefer incubation period, altricial hatchlings are nourished over a long lactation period with milk secreted by nipple-less mammary patches located on the female's abdomen. Milk is the sole source of nutrition and immune protection for the developing young until weaning. Using transcriptome and mass spectrometry analysis of milk cells and milk proteins, respectively, a novel Monotreme Lactation Protein (MLP) was identified as a major secreted protein in milk. We show that platypus and short-beaked echidna MLP genes show significant homology and are unique to monotremes. The MLP transcript was shown to be expressed in a variety of tissues; however, highest expression was observed in milk cells and was expressed constitutively from early to late lactation. Analysis of recombinant MLP showed that it is an N-linked glycosylated protein and biophysical studies predicted that MLP is an amphipathic, α-helical protein, a typical feature of antimicrobial proteins. Functional analysis revealed MLP antibacterial activity against both opportunistic pathogenic Staphylococcus aureus and commensal Enterococcus faecalis bacteria but showed no effect on Escherichia coli, Pseudomonas aeruginosa, Staphylococcus epidermidis, and Salmonella enterica. Our data suggest that MLP is an evolutionarily ancient component of milk-mediated innate immunity absent in other mammals. We propose that MLP evolved specifically in the monotreme lineage supporting the evolution of lactation in these species to provide bacterial protection, at a time when mammals lacked nipples.
| S-EPMC4224336 | biostudies-literature
Project description:Monotreme lactation protein (MLP) is a recently identified protein with antimicrobial activity. It is present in the milk of monotremes and is unique to this lineage. To characterize MLP and to gain insight into the potential role of this protein in the evolution of lactation, the crystal structure of duck-billed platypus (Ornithorhynchus anatinus) MLP was determined at 1.82 Å resolution. This is the first structure to be reported for this novel, mammalian antibacterial protein. MLP was expressed as a FLAG epitope-tagged protein in mammalian cells and crystallized readily, with at least three space groups being observed (P1, C2 and P21). A 1.82 Å resolution native data set was collected from a crystal in space group P1, with unit-cell parameters a = 51.2, b = 59.7, c = 63.1 Å, α = 80.15, β = 82.98, γ = 89.27°. The structure was solved by SAD phasing using a protein crystal derivatized with mercury in space group C2, with unit-cell parameters a = 92.7, b = 73.2, c = 56.5 Å, β = 90.28°. MLP comprises a monomer of 12 helices and two short β-strands, with much of the N-terminus composed of loop regions. The crystal structure of MLP reveals no three-dimensional similarity to any known structures and reveals a heretofore unseen fold, supporting the idea that monotremes may be a rich source for the identification of novel proteins. It is hypothesized that MLP in monotreme milk has evolved to specifically support the unusual lactation strategy of this lineage and may have played a central role in the evolution of these mammals.
| S-EPMC5947691 | biostudies-literature
Project description:Oviduct-specific glycoprotein (OVGP1) is a high molecular weight chitinase-like protein belonging to GH18 family. It is secreted by non-ciliated epithelial cells of oviduct during estrous cycle providing an essential milieu for fertilization and embryo development. The present study reports the characterization of buffalo OVGP1 through structural modeling, carbohydrate-binding properties and evolutionary analysis. Structural model displayed the typical fold of GH18 family members till the boundary of chitinase-like domain further consisting of a large (β/α)8 TIM barrel sub-domain and a small (α+β) sub-domain. Two critical catalytic residues were found substituted in the catalytic centre (Asp to Phe118, Glu to Leu120) compared with the active chitinase. The carbohydrate-binding groove in TIM barrel was lined with various conserved aromatic residues. Molecular docking with different sugars revealed the involvement of various residues in hydrogen-bonding and non-bonded contacts. Most of the substrate-binding residues were conserved except for a few replacements (Ser13, Lys48, Asp49, Pro50, Asp167, Glu199, Gln272 and Phe275) in comparison with other GH18 members. The residues Trp10, Trp79, Asn80, Gln272, Phe275 and Trp334 were involved in recognition of all six ligands. The α+β sub-domain participated in sugar-binding through Thr270, Gln272, Tyr242 and Phe275. The binding assays revealed significant sugar-binding with purified native and recombinant OVGP1. Phylogenetic analysis revealed that OVGP1 was closely related to AMCases followed by other CLPs and evolution of OVGP1 occurred through several gene duplications. This is the first study describing the structural characteristics of OVGP1 that will further help to understand its interaction with gametes to perform crucial reproductive functions.
| S-EPMC6904773 | biostudies-literature
Project description:Phospholipases A(2) (PLA(2)s) catalyze hydrolysis of fatty acids from the sn-2 position of phospholipids. Here we report the identification and characterization of a membrane-associated intracellular calcium-dependent, adipose-specific PLA(2) that we named AdPLA (adipose-specific phospholipase A(2)). We found that AdPLA was highly expressed specifically in white adipose tissue and was induced during preadipocyte differentiation into adipocytes. Clearance of AdPLA by immunoprecipitation significantly decreased PLA activity in white adipose tissue lysates but had no effect on liver lysates, where expression was hardly detectable. In characterizing AdPLA, we employed radiochemical assays with TLC analysis of the enzyme activity of lysates from COS-7 cells overexpressing AdPLA. For kinetic studies, we produced purified recombinant AdPLA for use in a lipoxidase-coupled spectrophotometric assay. AdPLA generated free fatty acid and lysophospholipid from phosphatidylcholine with a preference for hydrolysis at the sn-2 position. Although we found low but detectable lysophospholipase activity, AdPLA showed no significant activity against a variety of other lipid substrates. Calcium was found to activate AdPLA but was not essential for activity. Studies with known phospholipase inhibitors, including bromoenolactone, methyl arachidonyl fluorophosphate, AACOCF(3), 7,7-dimethyl-5,8-eicosadienoic acid, and thioetheramide, supported that AdPLA is a phospholipase. Mutational studies showed that His-23 and Cys-113 are critical for activity of AdPLA and suggested that AdPLA is likely a His/Cys PLA(2). Overall, although AdPLA is similar to other histidine phospholipases in pH and calcium dependence, AdPLA showed different characteristics in many regards, including predicted catalytic mechanism. AdPLA may therefore represent the first member of a new group of PLA(2)s, group XVI.
| S-EPMC2533091 | biostudies-literature
Project description:Endogenous retroviruses (ERVs) have played an essential role in the evolution of mammals. ERV-derived genes are reported in the therians, many of which are involved in placental development; however, the contribution of the ERV-derived genes in monotremes, which are oviparous mammals, remains to be uncovered. Here, we conducted a comprehensive search for possible ERV-derived genes in platypus and echidna genomes and identified three reverse transcriptase-like genes named RTOM1, RTOM2, and RTOM3 clustered in the GRIP2 intron. Comparative genomic analyses revealed that RTOM1, RTOM2, and RTOM3 are strongly conserved and are under purifying selection between these species. These could be generated by tandem duplications before the divergence of platypus and echidna. All RTOM transcripts were specifically expressed in the testis, possibly suggesting their physiological importance. This is the first study reporting monotreme-specific de novo gene candidates derived from ERVs, which provides new insights into the unique evolution of monotremes.
| S-EPMC9514029 | biostudies-literature
Project description:ABCG2 encodes a transporter protein that is associated with multidrug-resistant phenotypes in many cancers, including acute myeloid leukemia (AML); high levels of expression are generally associated with a poor prognosis. To better understand how expression of ABCG2 is controlled in pediatric AML, we performed a detailed analysis of the ABCG2 transcript isoforms from a variety of tissue sources, including 85 pediatric AML samples. These studies revealed a complex 5' untranslated region (UTR) with 6 novel exons and multiple splice variants. Samples from children with acute megakaryoblastic leukemia (AML FAB-M7) not associated with Down syndrome showed uniformly higher levels of ABCG2 transcripts than samples from children with other AML subtypes. A novel 5' UTR identified 90kb upstream of the exon 2 translation initiation site was expressed only in M7 AML subtypes. An associated upstream promoter fragment was shown to be selectively expressed in megakaryoblastic leukemia cells but not in human epithelial cell lines. These findings identify a new tissue-specific ABCG2 promoter that is selectively expressed in pediatric M7 AML. We also show a relatively high incidence of ABCG2 mRNA expression in non-Down associated M7 AML, which may contribute to the relatively poor prognosis of the M7 AML subtype.
| S-EPMC3163718 | biostudies-literature
Project description:ABCG2 encodes a transporter protein that is associated with multidrug-resistant phenotypes in many cancers, including acute myeloid leukemia (AML); high levels of expression are generally associated with a poor prognosis. To better understand how expression of ABCG2 is controlled in pediatric AML, we performed a detailed analysis of the ABCG2 transcript isoforms from a variety of tissue sources, including 85 pediatric AML samples. These studies revealed a complex 5' untranslated region (UTR) with 6 novel exons and multiple splice variants. Samples from children with acute megakaryoblastic leukemia (AML FAB-M7) not associated with Down syndrome showed uniformly higher levels of ABCG2 transcripts than samples from children with other AML subtypes. A novel 5' UTR identified 90kb upstream of the exon 2 translation initiation site was expressed only in M7 AML subtypes. An associated upstream promoter fragment was shown to be selectively expressed in megakaryoblastic leukemia cells but not in human epithelial cell lines. These findings identify a new tissue-specific ABCG2 promoter that is selectively expressed in pediatric M7 AML. We also show a relatively high incidence of ABCG2 mRNA expression in non-Down associated M7 AML, which may contribute to the relatively poor prognosis of the M7 AML subtype.
| S-EPMC3163718 | biostudies-literature
Project description:In this study, circular RNAs (circRNAs) from Holstein cow mammary tissues were identified and compared between early lactation and non-lactation. After analysis, 10,684 circRNAs were identified, ranging from 48 to 99,406 bp, and the average size was 882 bp. The circRNAs were mainly distributed on chromosomes 1 to 11, and 89.89% of the circRNAs belonged to sense-overlapping circRNA. The exons contained with circRNAs ranged from 1 to 47 and were concentrated from 1 to 5. Compared with the non-lactating cows, 87 circRNAs were significantly differentially expressed in the peak lactation cows. There were 68 upregulated circRNAs and 19 downregulated circRNAs. Enrichment analysis of circRNAs showed that GO analysis mainly focused on immune response, triglyceride transport, T cell receptor signaling pathway, etc. Pathway analysis mainly focused on cytokine-cytokine receptor interaction, T helper 17 cell differentiation, fatty acid biosynthesis, the JAK-STAT signaling pathway, etc. Specific primers were designed for two proximal ends of the circRNA junction sites to allow for PCR validation of four randomly selected circRNAs and carry out circRNA-miRNA interaction research. This study revealed the expression profile and characteristics of circRNAs in mammary tissue from Holstein cows at early lactation and non-lactation, thus providing rich information for the study of circRNA functions and mechanisms, as well as potential candidate miRNA genes for studying lactation in Holstein cows.
| S-EPMC8946036 | biostudies-literature
Project description:Paternally Expressed Gene 3 (Peg3) is an imprinted gene that controls milk letdown and maternal-caring behaviors. In this study, a conditional knockout allele has been developed in Mus musculus to further characterize these known functions of Peg3 in a tissue-specific manner. The mutant line was first crossed with a germline Cre. The progeny of this cross displayed growth retardation phenotypes. This is consistent with those seen in the previous mutant lines of Peg3, confirming the usefulness of the new mutant allele. The mutant line was subsequently crossed individually with MMTV- and Nkx2.1-Cre lines to test Peg3's roles in the mammary gland and hypothalamus, respectively. According to the results, the milk letdown process was impaired in the nursing females with the Peg3 mutation in the mammary gland, but not in the hypothalamus. This suggests that Peg3's roles in the milk letdown process are more critical in the mammary gland than in the hypothalamus. In contrast, one of the maternal-caring behaviors, nest-building, was interrupted in the females with the mutation in both MMTV- and Nkx2.1-driven lines. Overall, this is the first study to introduce a conditional knockout allele of Peg3 and to further dissect its contribution to mammalian reproduction in a tissue-specific manner.
| S-EPMC4671625 | biostudies-literature