Biology subjects

Ruppert, B.

Publications and source records attributed to Ruppert, B..

4 recordsLinked to original sources

bioRxiv2026

Cell cycle-coupled CK1δ turnover, autoinhibition, and activity

Casein kinase 1{delta} (CK1{delta}) is a ubiquitously expressed kinase involved in diverse cellular processes, including cell cycle regulation. CK1{delta} activity is attenuated by (auto)phosphorylation. However, inhibitory phosphorylation is efficiently opposed by cellular phosphatases as CK1{delta} accumulates in its hypophosphorylated, active state. CK1{delta} is a target of the nuclear ubiquitin ligase APC/C-CDH1, yet the kinase is apparently stable. Thus, the physiological relevance of CK1{delta} (auto)phosphorylation, autoinhibition, and regulated turnover has remained unclear. Here we show that CK1{delta} activity and abundance are coordinated in a cell cycle-dependent manner. During G1, assembled CK1{delta} kinase is stable while free active kinase is degraded. In S phase, unassembled CK1{delta} is no longer degraded, likely to support functions in DNA damage signaling. Upon mitotic entry, the downregulation of phosphatases promotes CK1{delta} (auto)phosphorylation and consequent autoinhibition, thereby preserving a pool of kinase to rapidly reestablish the post-mitotic steady state.

biochemistry↗

bioRxiv2025

CK1a-Mediated Two-Step Subunit Remodeling of the Circadian FRQ-FRH Complex

The circadian clock of Neurospora operates through a negative feedback loop in which FRQ, along with FRH and CK1a, inhibits its transcriptional activator, WCC, via phosphorylation. CK1a, anchored to FRQ, hyperphosphorylates FRQ at its IDRs in a slow, temperature-independent manner, forming a module suited for molecular timekeeping. However, the molecular processes triggered by FRQs hyperphosphorylation have remained unclear. We show that FRH, the folded binding partner of disordered FRQ, decodes FRQs time-dependent phosphorylation state by triggering a two-step remodeling of the FRQ-FRH complex: initially, two FRH molecules bind a FRQ dimer, keeping it inactive by blocking its interaction with WCC. Gradual phosphorylation induces with a delay the dissociation of one FRH, exposing a binding site for WCC and activating the complex. Following a time delay, attributable to the slow and stochastic nature of phosphorylation, the release of the second FRH promotes nuclear export and subsequent degradation of FRQ. This stepwise remodeling ensures precise activation and inactivation of FRQ and positions FRH as a hub for decoding temporal phosphorylation information.

cell biology↗

bioRxiv2025

Nuclear CK1δ as a Critical Determinant of PER:CRY Complex Dynamics and Circadian Period

The mammalian circadian clock is governed by a feedback loop in which the transcription activator CLOCK:BMAL1 induces expression of its inhibitors, PERs and CRYs, which form a complex with CK1{delta}, the main circadian kinase. However, the spatiotemporal dynamics of this feedback loop and the precise role of CK1{delta} remain incompletely understood. Using an inducible overexpression system, we show that nuclear availability of CK1{delta} is limited by both rapid nuclear degradation and active export of unassembled kinase, while cytoplasmic kinase is readily available for association with PERs. We demonstrate that CK1{delta}-mediated phosphorylation may disrupt PER2-CRY1 interaction thereby resulting in cytoplasmic PER2 dimers containing substoichiometric amounts of CRY1. Analysis of endogenous PER2 localization in the context of an intact circadian clock reveals that PER2 accumulates in the cytoplasm late in the circadian cycle. Based on these findings, we propose that cytoplasmic accumulation of PER:CRY:CK1{delta} complexes contributes to the clearance of nuclear PER2, while the CK1{delta}-dependent release of CRY1 into the nucleus may sustain CLOCK:BMAL1 repression on DNA supporting the transition from the early to the late repressive phase.

biochemistry↗

bioRxiv2023

CK1δ homeostasis by activity-dependent shuttling and degradation of orphan kinase

Casein kinase 1{delta} (CK1{delta}) is a simple monomeric enzyme involved in the regulation of a variety of functions, including signal transduction, the circadian clock, and the cell cycle. Although CK1{delta} is targeted by the ubiquitin ligase APC/CCdh1 is not understood how CK1{delta} expression is regulated to support its multiple functions. Here, we show that kinase activity controls CK1{delta} homeostasis by coordinating two competing processes: export from the nucleus to ensure distribution of CK1{delta} between its assembly partners, and proteasomal degradation of unassembled CK1{delta} in the nucleus to keep the amount of active, potentially deleterious orphan kinase low. During mitosis, CK1{delta} is released from centrosomes and stabilized by (auto)phosphorylation to preserve it for the subsequent G1 phase. TeaserCompetitive nuclear export and nuclear degradation of active CK1{delta} ensure efficient partner interaction and keep unassembled kinase levels low.

biochemistry↗