Controlling periodic long-range signalling to drive a morphogenetic transition

Abstract

Cells use signal relay to transmit information across tissue scales. However, the production of information carried by signal relay remains poorly characterised. To determine how the coding features of signal relay are generated, we used the classic system for long-range signalling: the periodic cAMP waves that drive Dictyostelium collective migration. Combining imaging and optogenetic perturbation of cell signalling states, we find that migration is triggered by an increase in wave frequency generated at the signalling centre. Wave frequency is regulated by cAMP wave circulation, which organises the long-range signal. To determine the mechanisms modulating wave circulation, we combined mathematical modelling, the general theory of excitable media and mechanical perturbations to test competing models. Models in which cell density and spatial patterning modulate the wave frequency cannot explain the temporal evolution of signalling waves. Instead, our evidence leads to a model where wave circulation increases the ability for cells to relay the signal, causing further increase in the circulation rate. This positive feedback between cell state and signalling pattern regulates the long-range signal coding that drives morphogenesis.

Collective signalling drives rapid jumping between cell states

“…The induction was not fully penetrant, with strong transcription induced in three out of eight experimental replicates, however the results are suggestive that the cafA gene is responsive to cAMP, but at lower amplitudes or frequencies of stimulation. This would be consistent with its expression earlier in development than carA , when cAMP signalling is more unstructured and infrequent ( Ford et al, 2023 ).…”

“…To test this, we imaged transcription of jump genes in the colony ( ), with parallel tracking of cAMP signalling, by mixing the transcriptional reporter cells with cells expressing the cAMP reporter Flamindo2. Flamindo2 is an intensiometric cAMP reporter that dims in fluorescence when it binds to cAMP ( Ford et al, 2023 ; Hashimura et al, 2019 ; Kundert et al, 2020 ) ( ). We obtained time series data simultaneously recording the physiological dynamics of both transcription and signalling, in the unperturbed colony, from the undifferentiated cells through to the cells at the aggregation stage of differentiation, over millimetre length scales ( Fig.…”

Collective signalling drives rapid jumping between cell states

“…To test this, we imaged transcription of jump genes in the colony (Movie S1), with parallel tracking of cAMP signalling, by mixing the transcriptional reporter cells with cells expressing the cAMP reporter, Flamindo2. Flamindo2 is an intensiometric cAMP reporter that dims in fluorescence when it binds to cAMP (Ford et al, 2023;Hashimura et al, 2019;Kundert et al, 2020)(Movie S2). We obtained simultaneous time series data recording the physiological dynamics of both transcription and signalling, in the unperturbed colony, from the undifferentiated cells through to the cells at the aggregation stage of differentiation (Figure 3A).…”

“…To detect MS2 and PP7 stem loops we expressed GFP and TdTomato-tagged MCP and PCP proteins (Antolovic et al, 2019). For stable uniform Flamindo2 expression, we targeted a codon-optimised Flamindo2 gene into the act5 gene of Ax3 cells as previously described (Ford et al, 2023). To disrupt the acaA gene, we used the acaA targeting vector from (Tweedy et al, 2020).…”

“…To disrupt the acaA gene, we used the acaA targeting vector from (Tweedy et al, 2020). For bPAC, a codon-optimised bPAC gene (Ford et al, 2023) was expressed from the extrachromosomal vector pDM1203 (Paschke et al, 2018), in Flamindo2-expressing acaA-cells.…”