Biology subjects

Dobson, T.

Publications and source records attributed to Dobson, T..

3 recordsLinked to original sources

bioRxiv2025

REST elevation-dependent chromatin remodeling and alternative Grk6 transcript synthesis hyperactivates Cxcr4-Sdf1 signaling in cerebellar granule cell progenitors

RE1 Silencing Transcription Factor (REST) is a repressor of transcriptional initiation of genes involved in neurogenesis. Here, we show that conditional REST elevation in cerebellar granule cell progenitors (CGNPs) of RESTTG mice perturbed foliation, increased cell migration, and sustained C-X-C motif receptor 4 (Cxcr4) signaling, a pathway key to postnatal CGNP migration. Mechanistic studies uncovered a novel role for REST in controlling transcript diversity and exon skipping in CGNPs. Alternative transcript expression was detected in known Cxcr4 signaling regulator, G-protein-coupled receptor kinase-6 (Grk6). Further analysis of Grk6s isoform expression revealed an upregulation of a transcript lacking exon 10a (Grk6-207) in RESTTG CGNPs. Grk6-207 expression in wildtype CGNPs hyperactivated Cxcr4 signaling and increased chemotaxis. Structural modeling of Grk6-207 predicted changes in active site conformation and interactions with Cxcr4 and {beta}-Arrestin-1, supporting impairment of Cxcr4 signaling desensitization. Interestingly, REST elevation promoted increased chromatin accessibility at the exon10a-10b junction and exon 10a exclusion. Integrated multiomic analyses identified the enhancer of zeste (Ezh2) as a potential mediator of alternative transcript generation which demonstrated increased occupancy at the exon10a-10b locus in RESTTG CGNPs. Pharmacological inhibition of Ezh2 downregulated Grk6-207, confirming a role for Ezh2 in Grk6 exon10a exclusion and the increased migration in RESTTG CGNPs. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=67 SRC="FIGDIR/small/682654v1_ufig1.gif" ALT="Figure 1"> View larger version (22K): org.highwire.dtl.DTLVardef@166db32org.highwire.dtl.DTLVardef@1c0c5e0org.highwire.dtl.DTLVardef@ff2d6borg.highwire.dtl.DTLVardef@fed170_HPS_FORMAT_FIGEXP M_FIG C_FIG

neuroscience↗

bioRxiv2025

Identification of Raptor and GLI1 as USP37 substrates highlight its context-specific function in medulloblastoma cells.

USP37 gene encodes a deubiquitylase (DUB), which catalyzes the proteolytic removal of ubiquitin moieties from proteins to modulate their stability, cellular localization or activity. Its expression is downregulated in a subgroup of medulloblastomas driven by constitutive activation of sonic hedgehog (SHH) signaling. Patients with SHH-driven medulloblastomas with elevated expression of the RE1 silencing transcription factor (REST) and reduced expression of USP37 have poor outcomes. In previous studies, we showed sustained proliferation of SHH-medulloblastoma cells due to blockade of terminal cell cycle exit and neuronal differentiation stemming from a failure in USP37-dependent stabilization of its target, the cyclin-dependent kinase inhibitor (CDKI)-p27. This finding suggested a tumor suppressive function for USP37. Interestingly, the current study also uncovered Raptor, a component of the mTORC1 complex, as a novel target of USP37. Under conditions of low-USP37 expression, reduced Raptor stability and mTORC1 activity caused a decline in phosphorylation of 4E-binding protein 1 (4EBP1) and increased its interaction with eukaryotic elongation factor 4E (eIF4E), which is known to inhibit CAP-dependent translation initiation. Surprisingly, a subset of patients with SHH-driven medulloblastomas with elevated expression of USP37 and the Glioma associated Oncogene 1 (GLI1), also exhibited poor outcomes. Using genetic and biochemical analyses, we showed that USP37-mediated stabilization of GLI1, a terminal effector of SHH signaling, increases pathway activity and upregulates expression of its target oncogene products, NMYC and CCND1, to drive cell proliferation. These data indicate that USP37 elevation in SHH-driven medulloblastomas has the potential to promote non-canonical activation of SHH signaling. Overall, our findings suggest that USP37 may have context-specific oncogenic and tumor suppressive roles in medulloblastoma cells.

cancer biology↗

bioRxiv2024

A chimeric viral platform for directed evolution in mammalian cells

Directed evolution (DE) is a process of mutation and artificial selection to breed biomolecules with new or improved activity1,2. DE platforms are primarily prokaryotic or yeast-based, and stable mutagenic mammalian systems have been challenging to establish and apply3. To this end, we developed PROTein Evolution Using Selection (PROTEUS), a new platform that uses chimeric virus-like vesicles (VLVs) to enable extended mammalian DE campaigns without loss of system integrity. This platform is stable and can generate sufficient diversity for DE in mammalian systems. Using PROTEUS, we altered the doxycycline responsiveness of tetracycline-controlled transactivators, generating a more sensitive TetON-4G tool for gene regulation. PROTEUS is also compatible with intracellular nanobody evolution, and we use it to design a DNA damage-responsive anti-p53 nanobody. Overall, PROTEUS is an efficient and stable platform to direct evolution of biomolecules within mammalian cells.

molecular biology↗