Fraser Child

Prolyl-hydroxylases (PHDs) are oxygen-sensing enzymes that mediate the hydroxylation of proline residues. In mammals, three PHD isoforms (PHD1–3) are responsible for proline hydroxylation of hypoxia-inducible factor (HIF) alpha, a key regulator of the hypoxia response. In the accompanying paper (Jiang et al., 2025), we report development of a mass spectrometry-based method to reliably identify proline hydroxylation (OH-Pro) sites on proteins and use this to identify a PHD-dependent OH-Pro modification at Pro604 on the protein RepoMan (CDCA2), a regulatory subunit for protein phosphatase PP1γ with important roles in mitotic progression and cell viability. Here, we investigate the functional significance of hydroxylation of RepoMan at P604. During M phase, the PP1-RepoMan complex dephosphorylates Thr3 of Histone H3 (H3T3) on chromosome arms to ensure the correct localisation of the chromosomal passenger complex (CPC) at centromeres. We show that siRNA depletion of PHD1, but not PHD2, increases H3T3 phosphorylation in prometaphase-arrested cells. In cells depleted of endogenous RepoMan, exogenous expression of wild-type RepoMan, but not a RepoMan-P604A mutant, restored normal H3T3 phosphorylation localisation in prometaphase arrested cells. RepoMan-P604 is located proximal to the short linear motifs (SLiMs) that function as binding sites for the serine/threonine protein phosphatase 2A (PP2A). The interaction of RepoMan and PP2A-B56γ is reduced in cells expressing RepoMan-P604A. Moreover, analyses in both fixed and live cells released from a prometaphase arrest show that expression of the RepoMan-P604A mutant delays completion of mitosis, results in defects in chromosome alignment and segregation, and increases levels of cell death. These data support a role for PHD1-mediated prolyl hydroxylation in controlling progression through mitosis, acting, at least in part, via hydroxylation of RepoMan at P604 regulating the interaction of RepoMan with PP2A during chromosome alignment and thereby controlling the levels of Histone H3 phosphorylation at Thr3.

Immune-checkpoint inhibitors (ICIs) targeting programmed cell death 1 (PD1) and programmed cell death 1 ligand 1 (PDL1) have improved survival for patients with different types of solid tumour. However, clinical response in patients with muscle-invasive bladder cancer (MIBC) is limited, with only 20–30% demonstrating a sustained response. An improved understanding of ICI mechanisms and robust biomarkers will increase efficacy and enable patient stratification in MIBC. Hypoxia (low oxygen tension) and neutrophil infiltration are prevalent in MIBC and are associated with immunotherapy resistance. Hypoxia-associated extracellular matrix (ECM) remodelling can induce pro-tumour or anti-tumour neutrophil polarization through biomechanical and biochemical signalling. Hypoxia-associated ECM mechanisms alter neutrophil recruitment, polarization, activation and affect T cell-centric immunotherapies. However, the specific mechanisms by which hypoxia, ECM and neutrophils confer immunotherapy resistance in MIBC are not yet fully understood. ICI resistance could be overcome by targeting specific ECM remodelling-related and neutrophil-related pathways to elicit durable and efficacious responses in 70–80% of patients with MIBC who are currently non-responsive to ICIs.

The cell cycle is an important cellular process whereby the cell attempts to replicate its genome in an error-free manner. As such, mechanisms must exist for the cell cycle to respond to stress signals such as those elicited by hypoxia or reduced oxygen availability. This review focuses on the role of transcriptional and post-transcriptional mechanisms initiated in hypoxia that interface with cell cycle control. In addition, we discuss how the cell cycle can alter the hypoxia response. Overall, the cellular response to hypoxia and the cell cycle are linked through a variety of mechanisms, allowing cells to respond to hypoxia in a manner that ensures survival and minimal errors throughout cell division.