Biology subjects

Lundgren, S. M.

Publications and source records attributed to Lundgren, S. M..

3 recordsLinked to original sources

bioRxiv2025

Parallel CRISPR screens reveal pathways controlling the cell surface levels of the attractant receptor FPR1

Levels of receptors on the cell surface control the sensitivity of immune cells, which is central to achieving effective responses while avoiding inflammatory diseases. The chemoattractant receptor FPR1 activates strong chemotactic and cytotoxic responses in neutrophils. While its precise regulation is critical for appropriate responses, the mechanisms controlling its cell surface levels remain unclear. Here, we investigated the roles of both classic and unknown regulators. First, we found that multiple G protein-coupled receptor kinases (GRK2, GRK3, and GRK6), and both {beta}-arrestin1 and 2 are important for FPR1 internalization. However, FPR1 uses multiple endocytic pathways, as cells lacking {beta}-arrestins have strong, but incomplete defects in FPR1 internalization. Moreover, we performed two parallel genome-wide CRISPR/Cas9 screens for the regulators of FPR1 surface expression and internalization in a neutrophil-like cell line. These screens identified regulators of FPR1 surface expression, recycling, and endocytosis. We identified the formin mDia1 and the small GTPase Arf6 as specific regulators of FPR1 internalization, and we confirmed these phenotypes using chemical inhibitors in primary human neutrophils. Furthermore, our data indicates that Arf6 contributes to the {beta}-arrestin-independent pathway. Collectively, our results clarify the contributions of GRKs and {beta}-arrestins in FPR1 internalization, indicate that internalization involves multiple compensatory routes, and uncover previously unidentified regulators of FPR1 biogenesis and trafficking, providing new mechanistic insight into these processes.

cell biology↗

bioRxiv2022

Signaling dynamics distinguish high and low priority neutrophil attractant receptors

Human neutrophils respond to multiple chemical attractants to guide their migration from the vasculature to sites of infection and injury, where they clear pathogens and amplify inflammation. To properly focus their responses during this complex navigation, neutrophils distinguish between attractants to modulate their responses. They prioritize pathogen and injury derived signals over long-range inflammatory signals secreted by host cells. Different attractants can also drive qualitatively different modes of migration. As receptors recognizing both classes of attractant couple to Gi G-proteins, it remains unclear how downstream signaling pathways distinguish and prioritize between the inputs. Here, we use live-cell imaging to demonstrate that the responses differ in their signaling dynamics: low priority attractants cause transient responses, while responses to high priority attractants are sustained. We observe this difference in both primary neutrophils and differentiated HL-60 cells, for signaling outputs of calcium, a major regulator of secretion, and Cdc42, a primary regulator of polarity and cell steering. We find that the rapid attenuation of Cdc42 activation in response to LTB4 depends on the threonine 308 and serine 310 phosphorylation sites in the C-terminal tail of its receptor LTB4R, in a manner independent of endocytosis. Mutation of these residues to alanine impairs attractant prioritization, although it does not affect attractant-dependent differences in migration persistence. Our results indicate that distinct temporal regulation of shared signaling pathways distinguishes receptors and contributes to chemoattractant prioritization.

cell biology↗

bioRxiv2020

H3 K27M and EZHIP impede H3K27-methylation spreading by inhibiting allosterically stimulated PRC2

Diffuse midline gliomas and posterior fossa type-A ependymomas contain the highly recurrent histone H3 K27M mutation and the H3 K27M-mimic EZHIP, respectively. In vitro, H3 K27M and EZHIP are competitive inhibitors of Polycomb Repressive Complex 2 (PRC2) lysine methyltransferase activity. In vivo, these proteins reduce overall H3K27me3 levels, however residual peaks of H3K27me3 remain at CpG islands through an unknown mechanism. Here, we report that EZHIP and H3 K27M preferentially interact with an allosterically activated form of PRC2 in vivo. The formation of H3 K27M- and EZHIP-PRC2 complexes occurs at CpG islands containing H3K27me3 and impedes PRC2 and H3K27me3 spreading. While EZHIP is not found outside of placental mammals, we find that expression of human EZHIP reduces H3K27me3 in Drosophila melanogaster through a conserved molecular mechanism. Our results highlight the mechanistic similarities between EZHIP and H3 K27M in vivo and provide mechanistic insight for the retention of residual H3K27me3 in tumors driven by these oncogenes.

cancer biology↗