Two-step colocalization of MORC3 with PML nuclear bodies

Abstract

SummaryMany functional subdomains, including promyelocytic leukemia nuclear bodies (PML NBs), are formed in the mammalian nucleus. Various proteins are constitutively or transiently accumulated in PML NBs in a PML-dependent manner. MORC3 (microrchidia family CW-type zinc-finger 3), also known as NXP2, which consists of GHL-ATPase, a CW-type zinc-finger and coiled-coil domains, is localized in PML NBs, where it recruits and activates p53 to induce cellular senescence. Interestingly, we found that MORC3 can form PML-independent nuclear domains (NDs) in mouse hematopoietic cells and even in Pml-deficient cells. Here, we show that MORC3 colocalizes with PML by a two-step molecular mechanism: the PML-independent formation of MORC3 NDs by the ATPase cycle, and the association of MORC3 with PML via the SUMO1-SUMO-interacting motif (SIM). Similarly to other members of the GHLATPase family, MORC3 functions as a 'molecular clamp'. ATP binding induces conformational changes in MORC3, leading to the formation of MORC3 NDs, and subsequent ATP hydrolysis mediates the diffusion and binding of MORC3 to the nuclear matrix. MORC3 might clamp DNA or nucleosomes in MORC3 NDs via the CW domain. Furthermore, the SUMOylation of MORC3 at five sites was involved in the association of MORC3 with PML, and SUMO1-unmodified MORC3 formed NDs independently of PML.

Mouse MORC3 is a GHKL ATPase that localizes to H3K4me3 marked chromatin

“…It is also likely that other MORCs similarly use dimer formation and dissolution in their modes of action. Our data are consistent with an earlier proposal that MORC3 may act as a molecular clamp for DNA, with constitutive dimerization through the C-terminal coiled-coil domain and ATP-dependent dimerization through the ATPase domains (6). Indeed, bacterial GHKL ATPases, such as topoisomerase VI and gyrase, can trap DNA through dimerization of their ATPase domains.…”

“…This is consistent with other GHKL ATPases described in the literature, many of which have been reported to undergo ATP-dependent dimerization (8)(9)(10). Both plant and animal MORCs are capable of forming nuclear bodies, and mutations that impair ATP binding and/or hydrolysis disrupt nuclear body formation of human MORC3 (6).…”

“…MORCs have been reported to form functional homomultimers or heteromultimers, where multimerization is likely mediated by the N-and/or the C-terminal domains (4,6,7). The coiled-coil region has been proposed to promote constitutive dimerization, whereas N-terminal ATPase head dimerization occurs only on ATP binding (6). This is consistent with other GHKL ATPases described in the literature, many of which have been reported to undergo ATP-dependent dimerization (8)(9)(10).…”

“…Consistent with this hypothesis, human and murine MORC3 have been identified in mass spectrometry screens as H3K4me3 readers (12)(13)(14). Mutation of a critical tryptophan residue in the human MORC3 CW domain disrupts localization in the nucleus, suggesting that CW domain-mediated recognition of H3K4me3 is also critical for MORC3 targeting to chromatin (6).…”

“…The N terminus contains a GHKL (gyrase, heat-shock protein 90, histidine kinase, MutL) type ATPase domain, and at the C terminus is a coiled-coil segment. MORCs have been reported to form functional homomultimers or heteromultimers, where multimerization is likely mediated by the N-and/or the C-terminal domains (4,6,7). The coiled-coil region has been proposed to promote constitutive dimerization, whereas N-terminal ATPase head dimerization occurs only on ATP binding (6).…”