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Eukaryotic chromosomes exhibit a hierarchical organization that spans a spectrum of length scales, ranging from sub-regions known as loops, which typically comprise hundreds of base pairs, to much larger chromosome territories that can encompass a few mega base pairs. Chromosome conformation capture experiments that involve high-throughput sequencing methods combined with microscopy techniques have enabled a new understanding of inter- and intra-chromosomal interactions with unprecedented details. This information also provides mechanistic insights on the relationship between genome architecture and gene expression. In this article, we review the recent findings on three-dimensional interactions among chromosomes at the compartment, topologically associating domain, and loop levels and the impact of these interactions on the transcription process. We also discuss current understanding of various biophysical processes involved in multi-layer structural organization of chromosomes. Then, we discuss the relationships between gene expression and genome structure from perturbative genome-wide association studies. Furthermore, for a better understanding of how chromosome architecture and function are linked, we emphasize the role of epigenetic modifications in the regulation of gene expression. Such an understanding of the relationship between genome architecture and gene expression can provide a new perspective on the range of potential future discoveries and therapeutic research., (© 2023 IOP Publishing Ltd.)