High-resolution confocal microscopy and nano-scale distance measurements show difference in 3D conformation between active and inactive mouse beta-major globin loci.

Genomes are one of the major foundations of life due to their role in information storage, process regulation and evolution. However, the sequential and three-dimensional structure of the human genome in the cell nucleus as well as its interplay with and embedding into the cell and organism only arise scarcely. Nano-scale distance measurements show that within an actively transcribing locus the distance between the 5’ and 3’ end of the locus is 563±201±12nm. In in-active loci the 5’ to 3’ distance is 621±266±16nm which is statistically significant. Since the spatial distance distributions are very similar we conclude that for both cell types the underlying 3D architecture is not dramatically different as earlier hypothesized. However, frequency and cumulative frequency distributions show that the distance between the 5’ and 3’ end of the locus is more distinct and stable in erythroid (active) cells. In non-erythroid cells the distribution is broader, indicating a less distinct, more dynamic organized chromatin structure. These result agree with the hypothesis that gene activation requires a different/folded conformation of the chromatin than when genes are inactive. Now that only the nano- architecture seems to be functionally influenced by gene activation/transcription, this cannot be concluded on a larger scale. Inter-allelic differences do not show preference towards (interactive) chromatic co- and/or near- localization. Distance measurements across this entire 218Kb chromatin region will elucidate the complete 3D structure of active and inactive mouse β-major globin loci. Furthermore, distance measurements combined with computer simulations (fig. 10) based on theoretical and/or empirical data, can give a better insight into high and low order chromatin architecture.