Abstract
BackgroundThe crucial role of the major histocompatibility complex (MHC) for the immune response to infectious diseases is well-known, but no information is available on the 3D nuclear organization of this gene-dense region in immune cells, whereas nuclear architecture is known to play an essential role on genome function regulation. We analyzed the spatial arrangement of the three MHC regions (class I, III and II) in macrophages using 3D-FISH. Since this complex presents major differences in humans and pigs with, notably, the presence of the centromere between class III and class II regions in pigs, the analysis was implemented in both species to determine the impact of this organization on the 3D conformation of the MHC. The expression level of the three genes selected to represent each MHC region was assessed by quantitative real-time PCR. Resting and lipopolysaccharide (LPS)-activated states were investigated to ascertain whether a response to a pathogen modifies their expression level and their 3D organization.ResultsWhile the three MHC regions occupy an intermediate radial position in porcine macrophages, the class I region was clearly more peripheral in humans. The BAC center-to-center distances allowed us to propose a 3D nuclear organization of the MHC in each species. LPS/IFNγ activation induces a significant decompaction of the chromatin between class I and class III regions in pigs and between class I and class II regions in humans. We detected a strong overexpression of TNFα (class III region) in both species. Moreover, a single nucleus analysis revealed that the two alleles can have either the same or a different compaction pattern. In addition, macrophage activation leads to an increase in alleles that present a decompacted pattern in humans and pigs.ConclusionsThe data presented demonstrate that: (i) the MHC harbors a different 3D organization in humans and pigs; (ii) LPS/IFNγ activation induces chromatin decompaction, but it is not the same area affected in the two species. These findings were supported by the application of an original computation method based on the geometrical distribution of the three target genes. Finally, the position of the centromere inside the swine MHC could influence chromatin reorganization during the activation process.
Highlights
The crucial role of the major histocompatibility complex (MHC) for the immune response to infectious diseases is well-known, but no information is available on the 3D nuclear organization of this gene-dense region in immune cells, whereas nuclear architecture is known to play an essential role on genome function regulation
We previously investigated the spatial arrangements of genes differentially expressed in porcine macrophages when the immune cells are activated by LPS/ IFNγ [28]
We found the same results in neutrophils, another type of immune cell, but not in fibroblasts, except for TNFα, which retains its tendency to be at the edge or outside of its chromosome territories (CT) in the three cell types tested
Summary
The crucial role of the major histocompatibility complex (MHC) for the immune response to infectious diseases is well-known, but no information is available on the 3D nuclear organization of this gene-dense region in immune cells, whereas nuclear architecture is known to play an essential role on genome function regulation. The first line of defense is provided by an innate immune response consisting of barriers such as skin, tears, saliva and mucus, as well as an inflammatory response This is closely followed by defensive mechanisms due to adaptive immune responses that include both a humoral response produced by antibodies, and a cell-mediated response produced by T cells that have the ability to destroy other cells. Neutrophils are the first cells to intervene at infection sites but have a short lifespan and are taken over by monocytes that differentiate into macrophages and phagocyte both pathogens and apoptotic neutrophils. To do this, these immune cells undergo a series of distinct functional changes both at the cytoplasm and nucleus levels. Most of these changes imply variations in gene expression revealed by transcriptomic analyses [3]
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