Chromosome Structure I: Loop extrusion or boundary:boundary pairing?

Abstract

Two different models have been proposed to explain how the endpoints of chromatin looped domains (“TADs”) in eukaryotic chromosomes are determined. In the first, a cohesin complex extrudes a loop until it encounters a boundary element roadblock, generating a stem-loop (and an unanchored loop). In this model, boundaries are functionally autonomous: they have an intrinsic ability to halt the movement of incoming cohesin complexes that is independent of the properties of neighboring boundaries. In the second, loops are generated by boundary:boundary pairing. In this model, boundaries are functionally non-autonomous, and their ability to form a loop depends upon how well they match with their neighbors. Moreover, unlike the loop-extrusion model, pairing interactions can generate both stem-loops and circle-loops. We have used a combination of MicroC to analyze how TADs are organized and experimental manipulations of theeven skippedTAD boundary,homie, to test the predictions of the “loop-extrusion” and the “boundary-pairing” models. Our findings are incompatible with the loop-extrusion model and instead suggest that endpoints of TADs in flies are determined by a mechanism in which boundary elements physically pair with their partners, either head-to-head, or head-to-tail, with varying degrees of specificity. How the partners find each other is not clear but is unlikely to require a loop extrusion mechanism.

Chromosome Structure II: Stem-loops and circle-loops

“…If, as seems likely, a significant fraction of the TADs genome-wide are circle-loops, this would effectively exclude cohesin-based loop extrusion as a general mechanism for TAD formation in flies. In addition, though stem-loops could be generated by a cohesin-dependent mechanism, it is unlikely that this mechanism is operational in flies, as we have shown here and in Bing et al (Bing et al ., 2023) that stem-loops in flies are formed by orientation-dependent boundary:boundary pairing.…”

“…The orientation of boundary:boundary pairing interactions determines the topology of each chromatin loop (Bing et al ., 2023; Fujioka et al ., 2016). Since nhomie and homie pair with each other head-to-tail, the endogenous eve TAD is a stem-loop.…”

“…PRE pairing in cis typically takes place within the context of a larger chromosomal domain, or TAD. In contrast, boundary elements are responsible for defining the endpoints of these looped domains (Arzate-Mejía et al, 2020; Batut et al ., 2022; Bing et al ., 2023; Chetverina et al ., 2017; Ibragimov et al, 2023; Stadler et al, 2017). While not much is known about the parameters governing PRE pairing, the pairing interactions of fly boundaries have been studied in some detail.…”

“…An alternating pattern of TADs connected by DNA segments that crosslink to each other with reduced frequency (c.f., 11 DNA below) is not observed in MicroC experiments. Instead, most TADs appeared to be directly connected to both of their neighbors without an intervening unanchored loop (Batut et al, 2022; Bing et al, 2023; Levo et al, 2022): see also below). This would suggest that TAD boundaries are typically linked to both neighbors, either simultaneously or as alternating pair-wise interactions.…”

Thehomieinsulator has sub-elements with different insulating and long-range pairing properties

“…This constraint arises because homie in the transgene pairs with homie in the eve locus head-to-head, and it pairs with eve -locus nhomie head-to-tail (F ujioka et al . 2016; B ing et al . 2024).…”