The Prolyl Isomerase Pin1 Promotes the Herpesvirus-Induced Phosphorylation-Dependent Disassembly of the Nuclear Lamina Required for Nucleocytoplasmic Egress

Abstract

The nuclear lamina lines the inner nuclear membrane providing a structural framework for the nucleus. Cellular processes, such as nuclear envelope breakdown during mitosis or nuclear export of large ribonucleoprotein complexes, are functionally linked to the disassembly of the nuclear lamina. In general, lamina disassembly is mediated by phosphorylation, but the precise molecular mechanism is still not completely understood. Recently, we suggested a novel mechanism for lamina disassembly during the nuclear egress of herpesviral capsids which involves the cellular isomerase Pin1. In this study, we focused on mechanistic details of herpesviral nuclear replication to demonstrate the general importance of Pin1 for lamina disassembly. In particular, Ser22-specific lamin phosphorylation consistently generates a Pin1-binding motif in cells infected with human and animal alpha-, beta-, and gammaherpesviruses. Using nuclear magnetic resonance spectroscopy, we showed that binding of Pin1 to a synthetic lamin peptide induces its cis/trans isomerization in vitro. A detailed bioinformatic evaluation strongly suggests that this structural conversion induces large-scale secondary structural changes in the lamin N-terminus. Thus, we concluded that a Pin1-induced conformational change of lamins may represent the molecular trigger responsible for lamina disassembly. Consistent with this concept, pharmacological inhibition of Pin1 activity blocked lamina disassembly in herpesvirus-infected fibroblasts and consequently impaired virus replication. In addition, a phospho-mimetic Ser22Glu lamin mutant was still able to form a regular lamina structure and overexpression of a Ser22-phosphorylating kinase did not induce lamina disassembly in Pin1 knockout cells. Intriguingly, this was observed in absence of herpesvirus infection proposing a broader importance of Pin1 for lamina constitution. Thus, our results suggest a functional model of similar events leading to disassembly of the nuclear lamina in response to herpesviral or inherent cellular stimuli. In essence, Pin1 represents a regulatory effector of lamina disassembly that promotes the nuclear pore-independent egress of herpesviral capsids.

The human cytomegalovirus nuclear egress complex unites multiple functions: Recruitment of effectors, nuclear envelope rearrangement, and docking to nuclear capsids

“…In addition, cellular protein kinases were found NEC‐associated, in particular, PKC and CDK1, as identified by pUL50‐specific coimmunoprecipitation . In an NEC associated form, the activity of cellular kinases PKC and CDK1, however, is most probably not directly targeted to nuclear lamins, because pUL97 was shown to be essential for high levels of site‐specific lamin phosphorylation . Instead, recruited protein kinases may be required for proper NEC formation, including events of regulatory phosphorylation of NEC‐associated proteins …”

“…This scenario, however, was challenged by the identification of the phosphorylation‐dependent lamin binding of the peptidyl‐prolyl cis/trans isomerase Pin1 and the functional validation that strongly supported a mode of Pin1‐induced conformational change facilitating lamina disassembly . Recently, a combined strategy of pharmacological Pin1 inhibition and Pin1 knockout provided conclusive evidence that Pin1 promotes the disassembly of the nuclear lamina during HCMV replication …”

“…55,56,74 In an NEC associated form, the activity of cellular kinases PKC and CDK1, however, is most probably not directly targeted to nuclear lamins, because pUL97 was shown to be essential for high levels of site-specific lamin phosphorylation. 60,116 Instead, recruited protein kinases may be required for proper NEC formation, including events of regulatory phosphorylation of NEC-associated proteins. 72,74 Site-specific phosphorylation of lamins A/C at Ser22 is known to mediate nuclear lamina disassembly both during herpesviral nuclear egress and cellular mitosis.…”

“…72,74 Site-specific phosphorylation of lamins A/C at Ser22 is known to mediate nuclear lamina disassembly both during herpesviral nuclear egress and cellular mitosis. 44,60,118 Seeking to understand the further-reaching consequences of this phosphorylation step, it has been proposed on the basis of lamin A coil 2B dimeric crystal structures that phosphorylation may interfere with electrostatic interactions. 119 cles.…”

Transmembrane Protein pUL50 of Human Cytomegalovirus Inhibits ISGylation by Downregulating UBE1L

“…The integral membrane protein pUL50 is associated with the inner nuclear membrane (INM) by its transmembrane (TM) domain, whereas the nuclear phosphoprotein pUL53 carries a classical nuclear localization signal and accumulates in the nuclear rim upon interaction with the INMassociated pUL50 protein. Once the core NEC is formed, further NEC proteins, such as p32/gC1qR, emerin, and protein kinases, such as the viral protein kinase pUL97, cellular protein kinase C isoform a (PKCa), and cellular cyclin-dependent kinase 1 (CDK1), are recruited (42)(43)(44)(45)(46) in order to phosphorylate nuclear lamins, resulting in lamina disassembly (47)(48)(49). The recently solved crystal structure of the pUL50-pUL53 complex suggests that pUL50-pUL53 heterodimers assemble to form hexameric ring-like structures, acting as a scaffold for binding of further NEC proteins (50).…”