Transcriptomic and nuclear architecture of immune cells after LPS activation

Abstract

Changes in the nuclear positioning of specific genes, depending on their expression status, have been observed in a large diversity of physiological processes. However, gene position is poorly documented for immune cells which have been subjected to activation following bacterial infection. Using a pig model, we focused our study on monocyte-derived macrophages and neutrophils, as they are the first lines of defence against pathogens. We examined whether changes in gene expression due to LPS activation imply that genes have repositioned in the nuclear space. We first performed a transcriptomic analysis to identify the differentially expressed genes and then analysed the networks involved during lypopolysaccharide/interferon gamma activation in monocyte-derived macrophages. This allowed us to select four up-regulated (IL1β, IL8, CXCL10 and TNFα) and four down-regulated (VIM, LGALS3, TUBA3 and IGF2) genes. Their expression statuses were verified by quantitative real-time RT-PCR before studying their behaviour in the nuclear space during macrophage activation by means of 3D fluorescence in situ hybridization. No global correlation was found between gene activity and radial positioning. Only TNFα belonging to the highly transcribed MHC region on chromosome 7 became more peripherally localized in relation to the less decondensed state of its chromosome territory (CT) in activated macrophages. The analysis of gene positioning towards their CT revealed that IL8 increases significantly its tendency to be outside its CT during the activation process. In addition, the gene to CT edge distances increase for the three up-regulated genes (IL8, CXCL10 and TNFα) among the four analysed. Contrarily, the four down-regulated genes did not change their position. The analysis of gene behaviour towards their CT was extended to include neutrophils for three (TNFα, IL8 and IL1β) up- and two (IGF2 and TUBA3) down-regulated genes, and similar results were obtained. The analysis was completed by studying the four up-regulated genes in fibroblasts, not involved in immune response. Our data suggest that relocation in the nuclear space of genes that are differentially expressed in activated immune cells is gene and cell type specific but also closely linked to the entire up-regulation status of their chromosomal regions.

First survey and functional annotation of prohormone and convertase genes in the pig

“…In experiment GSE7314, IGF2 was over-expressed in pigs inoculated with Salmonella choleraesuis at 21 dpi relative to non-inoculated pigs ( P -value < 8.1 × 10 -4 ). These results are consistent with reports that IGF2 is down-regulated in pigs immune-challenged with lipopolysaccharide [44]. …”

Sperm Nuclear Architecture Is Locally Modified in Presence of a Robertsonian Translocation t(13;17)

“…This non-random organization probably relies on functional constrains to regulate genome activity, ie regulation of gene expression. Thus, the nuclear environment could favors gene specific interactions between distant genomic regions to activate or repress their transcriptional activities [5], [6] but also modulate genes position within their respective chromosome territories, depending on their expression status [7]–[10]. However to date little is known on how this three-dimensional organization is controlled and transmitted through cell division but also through generations.…”

Three-Dimensional Arrangement of Genes Involved in Lipid Metabolism in Nuclei of Porcine Adipocytes and Fibroblasts in Relation to Their Transcription Level

“…Recent studies have shown that transcriptional activation of genes by hormone addition was not connected with changes in the nuclear gene positioning [Kocanova et al, 2010;Hakim et al, 2011]. Similarly, variations in expression levels that occurred during bacterial infection did not induce global repositioning of genes in the nuclear space of porcine macrophages [Solinhac et al, 2011]. Studies concerning the parasite influencing the host interphase genome organization have revealed gene repositioning events, but the behavior of the up-regulated genes was different -one of the studied genes moved to the nuclear interior and the other one moved to the nuclear edge [Knight et al, 2011].…”

“…Depending on the type of detected objects (genes or chromosome territories) the global thresholds are computed differently. If necessary, the segmentation thresholds were adjusted manually to improve object detection [Iannuccelli et al, 2010;Solinhac et al, 2011]. To determine the radial position of studied genes the ratio of measurement distance was calculated ( R = distance from the FISH signal to the nucleus center by distance from the FISH signal to the nuclear border).…”