In Vivo Expression of MHC Class I Genes Depends on the Presence of a Downstream Barrier Element

Regulation of MHC class I gene expression is critical to achieve proper immune surveillance. In this work, we identify elements downstream of the MHC class I promoter that are necessary for appropriate in vivo regulation: a novel barrier element that protects the MHC class I gene from silencing and elements within the first two introns that contribute to tissue specific transcription. The barrier element is located in intergenic sequences 3′ to the polyA addition site. It is necessary for stable expression in vivo, but has no effect in transient transfection assays. Accordingly, in both transgenic mice and stably transfected cell lines, truncation of the barrier resulted in transcriptional gene silencing, increased nucleosomal density and decreased histone H3K9/K14 acetylation and H3K4 di-methylation across the gene. Significantly, distinct sequences within the barrier element govern anti-silencing and chromatin modifications. Thus, this novel barrier element functions to maintain transcriptionally permissive chromatin organization and prevent transcriptional silencing of the MHC class I gene, ensuring it is poised to respond to immune signaling

Upregulation of matrix metalloproteinases in a model of T cell mediated tissue injury in the gut: analysis by gene array and in situ hybridisation

Background and aim: Matrix metalloproteinases (MMPs) have been implicated in tissue remodelling and ulceration in inflammatory bowel disease and coeliac disease. Studies to date have concluded that stromelysin 1 is functionally involved in mucosal degradation. However, there are many other MMPs whose function in the gut is currently unknown. This work had two aims: firstly, to use gene array technology to measure changes in MMP and tissue inhibitor of metalloproteinase (TIMP) expression in a model of T cell mediated injury in the gut, and secondly, to correlate data from gene arrays with that generated by in situ hybridisation.Methods: T cells in explants of human fetal gut were activated with pokeweed mitogen or anti-CD3 plus interleukin 12. Gene array analysis and in situ hybridisation were performed to investigate changes in MMP gene expression.Results: Both gene array analysis and in situ hybridisation indicated marked upregulation of stromelysin 2 and macrophage metalloelastase expression in the explants associated with mucosal destruction. The arrays also confirmed our previous observation that interstitial collagenase (MMP-1), stromelysin 1 (MMP-3), and gelatinase B (MMP-9) are upregulated but there was no change in MMP-2, -7, -8, -9, -11, -13, -14–17, or -19. Following T cell activation, transcripts for TIMPs were reduced.Conclusions: These results show that there is differential upregulation of MMPs during T cell responses in the gut and suggest that further studies on the role of stromelysin 2 and macrophage metalloelastase may show that they have a functional role. In addition, the increase in MMPs and reduction in TIMPs suggest that the protease/antiprotease balance in the mucosa may determine the extent of mucosal degradation

HMG Box Transcriptional Repressor HBP1 Maintains a Proliferation Barrier in Differentiated Liver Tissue

We previously isolated HBP1 as a target of the retinoblastoma (RB) and p130 family members and as the first of the HMG box transcriptional repressors. Our subsequent work demonstrated that HBP1 coordinates differentiation in cell culture models. In the present study, we show that HBP1 regulates proliferation in a differentiated tissue of an animal. Using transgenic mice in which HBP1 expression was specifically increased in hepatocytes under control of the transthyretin promoter, we determined the impact of HBP1 on synchronous cell cycle reentry following partial hepatectomy. Modest overexpression of HBP1 yielded a detectable cell cycle phenotype. Following a mitogenic stimulus induced by two-thirds partial hepatectomy, mice expressing the HBP1 transgene showed a 10- to 12-h delay in progression through G(1) to the peak of S phase. There was a concomitant delay in mid-G(1) events, such as the induction of cyclin E. While the delay in G(1) and S phases correlated with the slight overexpression of transgenic HBP1, the level of the endogenous HBP1 protein itself declined in S phase. In contrast, the onset of the immediate-early response following partial hepatectomy was unchanged in HBP1 transgenic mice. This observation indicated that the observed delay in S phase did not result from changes in signaling pathways leading into the G(0)-to-G(1) transition. Finally, transgenic mice expressing a mutant HBP1 lacking the N-terminal RB interacting domain showed a stronger S-phase response following partial hepatectomy. These results provide the first evidence that HBP1 can regulate cell cycle progression in differentiated tissues