Phosphorylation of chemoattractant receptors regulates chemotaxis, actin reorganization and signal relay.

Migratory cells, including mammalian leukocytes and Dictyostelium, use G-protein-coupled receptor (GPCR) signaling to regulate MAPK/ERK, PI3K, TORC2/AKT, adenylyl cyclase and actin polymerization, which collectively direct chemotaxis. Upon ligand binding, mammalian GPCRs are phosphorylated at cytoplasmic residues, uncoupling G-protein pathways, but activating other pathways. However, connections between GPCR phosphorylation and chemotaxis are unclear. In developing Dictyostelium, secreted cAMP serves as a chemoattractant, with extracellular cAMP propagated as oscillating waves to ensure directional migratory signals. cAMP oscillations derive from transient excitatory responses of adenylyl cyclase, which then rapidly adapts. We have studied chemotactic signaling in Dictyostelium that express non-phosphorylatable cAMP receptors and show through chemotaxis modeling, single-cell FRET imaging, pure and chimeric population wavelet quantification, biochemical analyses and TIRF microscopy, that receptor phosphorylation is required to regulate adenylyl cyclase adaptation, long-range oscillatory cAMP wave production and cytoskeletal actin response. Phosphorylation defects thus promote hyperactive actin polymerization at the cell periphery, misdirected pseudopodia and the loss of directional chemotaxis. Our data indicate that chemoattractant receptor phosphorylation is required to co-regulate essential pathways for migratory cell polarization and chemotaxis. Our results significantly extend the understanding of the function of GPCR phosphorylation, providing strong evidence that this evolutionarily conserved mechanism is required in a signal attenuation pathway that is necessary to maintain persistent directional movement of Dictyostelium, neutrophils and other migratory cells.

Informing resilience building: FAO's Surveillance Evaluation Tool (SET) Biothreat Detection Module will help assess national capacities to detect agro-terrorism and agro-crime.

Attacks using animal pathogens can have devastating socioeconomic, public health and national security consequences. The livestock sector has some inherent vulnerabilities which put it at risk to the deliberate or accidental spread of disease. The growing concern of countries about the risks of agro-terrorism and agro-crime has led to efforts to prepare against potential attacks. One recent international effort is the launch of a joint OIE, FAO and INTERPOL project in 2019 to build resilience against agro-terrorism and agro-crime targeting animal health with the financial support of the Weapons Threat Reduction Programme of Global Affairs Canada. Given the importance of strong animal health surveillance systems for the early and effective response to agro-terrorism and agro-crime, the project will use the FAO Surveillance Evaluation Tool (SET) and its new Biothreat Detection Module to evaluate beneficiary countries' capacities to detect criminal or terrorist animal health events. This paper presents the development of the new SET Biothreat Detection Module and how it will be used to evaluate surveillance for agro-terrorism and agro-crime animal disease threats. The module will be piloted in early 2021 and, once finalized, will be used by beneficiary countries of the joint OIE-FAO-INTERPOL project. Results from evaluations using SET and its Biothreat Detection Module are expected to provide a baseline from which countries can build targeted capacity for animal disease surveillance including early detection and investigation of potential terrorist or criminal events involving zoonotic and non-zoonotic animal pathogens.

Using total internal reflection fluorescence (TIRF) microscopy to visualize cortical actin and microtubules in the Drosophila syncytial embryo.

The Drosophila syncytial embryo is a powerful developmental model system for studying dynamic coordinated cytoskeletal rearrangements. Confocal microscopy has begun to reveal more about the cytoskeletal changes that occur during embryogenesis. Total internal reflection fluorescence (TIRF) microscopy provides a promising new approach for the visualization of cortical events with heightened axial resolution. We have applied TIRF microscopy to the Drosophila embryo to visualize cortical microtubule and actin dynamics in the syncytial blastoderm. Here, we describe the details of this technique, and report qualitative assessments of cortical microtubules and actin in the Drosophila syncytial embryo. In addition, we identified a peak of cortical microtubules during anaphase of each nuclear cycle in the syncytial blastoderm, and using images generated by TIRF microscopy, we quantitatively analyzed microtubule dynamics during this time.

Optic nerve astrocytoma in a dog.

Intracranial astrocytomas are relatively uncommon in dogs and optic nerve astrocytomas even more so. This neoplasm should be considered as differential in canine patients with vision loss, retinal detachment, ocular mass, and histopathologic findings of infiltrative fusiform to polygonal glial cells possibly associated with glomeruloid vascular proliferation.