Biology subjects

Wolff, F.

Publications and source records attributed to Wolff, F..

3 recordsLinked to original sources

bioRxiv2024

PCMD-1 stabilizes the PCM scaffold and facilitates centriole separation

Centrosomes are highly dynamic organelles, and maintaining their stability is crucial for spindle pole integrity and bipolar spindle formation. Centrosomes consist of a pair of centrioles surrounded by the pericentriolar material (PCM). In Caenorhabditis elegans, interactions between the PCM scaffold protein SPD-5 and the regulatory kinase PLK-1 are essential for PCM formation and disassembly. However, how PCM stability is established and maintained remains an open question. Here, we address this question by analyzing the function of PCMD-1, a protein that predominantly localizes to centrioles. Mutations in the predicted PLK-1 binding sites of PCMD-1 result in a reduction of centrosomal PLK-1 and PCMD-1 levels, leading to severe distortion of the PCM scaffold. The disorganization of the PCM is already evident during its formation and results in the assembly of a structurally unstable mitotic centrosome, which is unable to resist the microtubule pulling forces exerted on the spindle pole. As a consequence of a weakened PCM scaffold, the pulling forces are not effectively relayed to the entrapped centriole pair, resulting in delayed centriole separation in late anaphase. Together, these findings show that PCMD-1, which predominantly localizes to centrioles and tethers the PCM scaffold to them, is essential for stabilizing the entire PCM scaffold and ensuring timely centriole separation during PCM disassembly. We propose a model in which PCMD-1 initiates the ordered assembly of the PCM scaffold by biasing the PCM core to a certain intrinsic order around the centrioles. This intrinsic order acts as a seed that propagates throughout the surrounding micron-scale PCM scaffold, providing the necessary strength and structural integrity for the centrosome.

cell biology↗

bioRxiv2023

Spatially distinct inputs modulate the amount of active Mitotic-phase GAP to locally restrict RhoA signaling for successful cell division

At the end of mitosis a contractile ring consisting of filamentous actin (F-actin) assembles at the cell equator and ring constriction equally partitions the cellular content. Inhibitory and stimulatory signaling cascades spatially limit RhoA activity to a narrow central zone to promote F-actin polymerization only at the cell equator. While the stimulatory signal is well-characterized, the mechanisms by which inhibitory signals restrict active RhoA to a narrow equatorial zone and prevent improper RhoA activity at the cell poles are not known. Here, we identify two regulatory inputs that control the activation and cortical targeting of the Mitotic-phase GTPase activating protein (MP-GAP) for RhoA which limits RhoA activity both at the cell poles and at the cell equator. We show that at cell poles, MP-GAP is a direct phosphorylation target of spindle-pole associated Aurora A kinase. We identify three Aurora A target residues in a region of MP-GAP that binds directly to its catalytic GAP domain suggesting the Aurora A phosphorylation releases MP-GAP autoinhibition. We show that phosphorylation of these sites is required to keep RhoA activity low at the cell poles. At the cell equator, F-actin facilitates MP-GAP enrichment, which opposes RhoA GEF activity and thereby accelerates RhoA flux through the GTPase cycle for the formation of a narrow equatorial zone of active RhoA. Thus, the amount of active MP-GAP is modulated by two distinct regulatory inputs that function in spatially restricted locations: Aurora A phosphorylation relieves MP-GAP autoinhibition to limit RhoA activity at the cell poles, while F-actin polymerization promotes MP-GAP targeting to prevent distribution of active RhoA in a broad zone at the cell equator. By determining the mechanism of spatially confining RhoA activity at the equator and the cell poles during cytokinesis, our work has broad implications to how Rho activity zones are formed and maintained during cytokinesis and how defects in their formation impact animal development and disease.

cell biology↗

bioRxiv2020

Oncogenic GLI1 and STAT1/3 activation drives immunosuppressive tryptophan/kynurenine metabolism via synergistic induction of IDO1 in skin cancer cells

Pharmacological targeting of Hedgehog (HH)/GLI has proven effective for certain blood, brain and skin cancers including basal cell carcinoma (BCC). However, limited response rates and the development of drug resistance call for improved anti-HH therapies that take into account synergistic crosstalk mechanisms and immune evasion strategies. In previous work, we demonstrated that crosstalk of HH/GLI with pro-inflammatory Interleukin-6 (IL6) signaling drives BCC by promoting tumor cell proliferation [1]. In the present study, we screened for possible mechanisms of cancer immune evasion regulated by synergistic HH-IL6 signaling and identified the immunosuppressive enzyme indoleamine 2,3-dioxygenase 1 (IDO1) as a novel transcriptional target co-regulated by HH-IL6 signaling. Analysis of the cis-regulatory region of IDO1 by chromatin-immunoprecipitation revealed co-occupancy of this region by HH- IL6 induced GLI1 and STAT3 transcription factors along with active chromatin marks at the histone level. Elevated expression of IDO1 in human BCC with high-level HH and IL6 signatures supports the clinical relevance of our mechanistic data. Genetic inhibition of GLI1 expression prevented the induction of IDO1 expression in response to IL6/STAT3 and IFN{gamma}/STAT1 signaling in human melanoma cells. Pharmacological targeting of HH signaling at the level of GLI proteins interfered with IDO1 expression and consequently prevented the production of the immunosuppressive metabolite kynurenine generated by active IDO1 from tryptophan. Further, inhibition of GLI1 enhanced the efficacy of the selective IDO1 inhibitor epacadostat. Of note, inhibition of HH/GLI signaling in melanoma cells not only reduced IDO1 expression but also interfered with the repression of T cell activation by attenuating IDO1/kynurenine-mediated immunosuppression. These data identify the immunosuppressive IDO1-kynurenine pathway as a novel pro-tumorigenic effector of oncogenic HH-IL6 and GLI-STAT cooperation. Our data suggest simultaneous pharmacological targeting of the HH/GLI, JAK/STAT and IDO1- kynurenine axis as rational combination therapy in skin cancers.

cancer biology↗