Pregnancy-specific glycoprotein function, conservation and receptor investigation

Pregnancy-specific glycoproteins (PSGs) are highly glycosylated secreted proteins encoded by multi-gene families in some placental mammals. They are carcinoembryonic antigen (CEA) family and immunoglobulin (Ig) superfamily members. PSGs are immunomodulatory, and have been demonstrated to possess antiplatelet and pro-angiogenic properties. Low serum levels of these proteins have been correlated with adverse pregnancy outcomes. Objectives: Main research goals of this thesis were: 1). To attempt to replicate previously reported cytokine responses to PSG-treatment of immune cells and subsequently to investigate functionally important amino acids within PSG1. 2). To determine whether candidate receptor, integrin αvβ3, was a binding partner for PSG1 and to investigate whether PSG1 possessed functionality in a leukocyte-endothelial interaction assay. 3). To determine whether proteins generated from recently identified putative PSG genes in the horse shared functional properties with PSGs from other species. Outcomes: 1). Sequential domain deletion of PSG1 as well as mutation of conserved residues within the PSG1 Ndomain did not affect PSG1-induced TGF-β1. The investigated response was subsequently found to be the result of latent TGF-β1 contaminating the recombinant protein. Protein further purified by SEC to remove this showed no induction of TGF-β1. The most N-terminal glycosylation site was demonstrated to have an important role in PSG N domain secretion. PSG1 attenuated LPS-induced IL-6 and TNF-α. Investigations into signalling underpinning this proved inconclusive. 2). Integrin αvβ3 was identified as a novel PSG1 receptor mediating an as yet unknown function. Preliminary investigations into a role for PSGs as inhibitors of leukocyte endothelial interactions showed no effect by PSG1. 3). Horse PSG protein, CEACAM49, was shown to be similarly contaminated by latent TGF-β1 particle and once removed did not demonstrate TGF-β1 release. Interestingly horse PSG did show anti-platelet properties through inhibition of the plateletfibrinogen interaction as previously published for mouse and human PSGs.

The relevance of bacterial isoprenoid biosynthetic pathways for host-microbe interactions

This thesis was undertaken to investigate the relevance of two bacterial isoprenoid biosynthetic pathways (Mevalonate (MVAL) and 2-C-methyl-D-erythritol 4-phosphate (MEP)) for host-microbe interactions. We determined a significant reduction in microbial diversity in the murine gut microbiota (by next generation sequencing) following oral administration of a common anti-cholesterol drug Rosuvastatin (RSV) that targets mammalian and bacterial HMG-CoA reductase (HMG-R) for inhibition of MVAL formation. In tandem we identified significant hepatic and intestinal off-target alterations to the murine metabolome indicating alterations in inflammation, bile acid profiles and antimicrobial peptide synthesis with implications on community structure of the gastrointestinal microbiota in statin-treated animals. However we found no effect on local Short Chain Fatty Acid biosynthesis (metabolic health marker in our model). We demonstrated direct inhibition of bacterial growth in-vitro by RSV which correlated with reductions in bacterial MVAL formation. However this was only at high doses of RSV. Our observations demonstrate a significant RSV-associated impact on the gut microbiota prompting similar human analysis. Successful deletion of another MVAL pathway enzyme (HMG-CoA synthase (mvaS)) involved in Listeria monocytogenes EGDe isoprenoid biosynthesis determined that the enzyme is non-essential for normal growth and in-vivo pathogenesis of this pathogen. We highlight potential evidence for alternative means of synthesis of the HMG-CoA substrate that could render mvaS activity redundant under our test conditions. Finally, we showed by global gene expression analysis (Massive Analysis of cDNA Ends (MACE RNA-seq) a significant role for the penultimate MEP pathway metabolite (E)-4-hydroxy-3-methyl-2-but-2-enyl pyrophosphate (HMBPP) in significant up regulation of genes of immunity and antigen presentation in THP-1 cells at nanomolar levels. We infected THP-1 cells with wild type or HMBPP under/over-producing L. monoctyogenes EGDe mutants and determined subtle effects of HMBPP upon overall host responses to Listeria infection. Overall our findings provide greater insights regarding bacterial isoprenoid biosynthetic pathways for host-microbe/microbe-host dialogue.

Na marbhnaí do Shéamas Óg Mac Coitir (1689-1720)

Go príomha, is tráchtas é seo a dhéanann staidéar ar ghné de litríocht iar-chlasaiceach na Gaeilge. Baineann sé go háirithe leis an sraith chaointe nó marbhnaí i bhfoirm véarsaíochta a cumadh do Shéamas Óg Mac Coitir (1689-1720), duine uasal Caitliceach ó Charraig Tuathail, Co. Chorcaí, nuair a ciontaíodh é in éigniú Elizabeth Squibb, bean de Chumann na gCarad; nuair a cuireadh pionós an bháis air; agus nuair a crochadh é i gCathair Chorcaí an 7 Bealtaine, 1720. Ó thaobh na staire de, scrúdaítear Clann Choitir mar shampla de theaghlach nár cheil a ndílseacht do chúis pholaitiúil na Stíobhartach agus a sheas an fód go cróga faoi mar a bhí a ngreim polaitiúil á dhaingniú ag an gCinsealacht Phrotastúnach ó dheireadh an 17ú haois amach. Tagraítear do sheicteachas na sochaí comhaimseartha agus don teannas idir an pobal Caitliceach agus an pobal Protastúnach ag an am. Déantar scagadh ar an véarsaíocht mar fhoinse luachmhar do dhearcadh míshásta an mhóraimh Chaitlicigh ar struchtúr polaitiúil chontae Chorcaí (agus na hÉireann) i dtosach an 18ú haois. Is feiniméan liteartha an dlús véarsaíochta seo a bhaineann go háirithe le traidisiún liteartha Chorcaí. Tá na dánta curtha in eagar agus aistriúchán go Béarla curtha ar fáil: is é seo croí an tráchtais. Tá an t-eagrán bunaithe ar scrúdú cuimsitheach ar thraidisiún na lsí; pléitear modheolaíocht na heagarthóireachta. Déantar iarracht ar na dánta a shuíomh sa traidisiún casta liteartha sa tráchtaireacht tosaigh; sa chuid eile den bhfearas scoláiriúil, scrúdaítear ceisteanna a bhaineann le cúrsaí teanga, foclóra, meadarachta agus stíle. Tá innéacsanna agus liosta foinsí le fáil i ndeireadh an tráchtais.