Research Article

Nucleocytoplasmic Shuttling of a GATA Transcription Factor Functions as a Development Timer

Science  21 Mar 2014:
Vol. 343, Issue 6177,
DOI: 10.1126/science.1249531

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Structured Abstract

Introduction

Biological oscillations are universally found in nature and are critical at many levels of cellular organization. In the social amoeba Dictyostelium discoideum, starvation-triggered cell-cell aggregation and the early stages of developmental morphogenesis are orchestrated by periodic extracellular cAMP (3',5'-cyclic adenosine monophosphate) waves, which provide both chemotactic cues and developmental signals. Repeated occupancy of G protein–coupled cAMP receptors promotes optimal gene expression, whereas continuous stimulation suppresses the program. Although this activity was recognized nearly 40 years ago, the underlying mechanism for the stimulus-response pattern has not been elucidated.

Embedded Image

GtaC shuttling modulates developmental gene expression. (Top) Periodic nuclear enrichment of GtaC at times indicated in response to cAMP waves. (Bottom) In each cAMP stimulation cycle, GtaC transits through successive states of nuclear active (green), cytosolic (blue), and nuclear inactive (red), promoting transcription only in the active state. Repeated stimuli lead to reoccurring activations and incremental accumulation of gene products. The mechanism enables cells to count effective stimuli and modulate gene expression accordingly.

Rationale

We reasoned that the mechanism decoding cyclic stimuli might be traced to a receptor-mediated regulation of a gene that is required for developmental progression. In a genetic screen for mutants that affect proper development, we discovered a GATA family transcription factor, GtaC, which is essential in this process. Furthermore, in different cells at different times, GtaC was found to either accumulate in the nucleus or disperse in the cytosol, suggesting that the dynamic cellular localization of GtaC could be a key to the relay of oscillatory signaling.

Results

We report here that GtaC exhibits rapid nucleocytoplasmic shuttling in response to periodic cAMP waves. Persistent occupancy of the receptors by cAMP causes GtaC to translocate and remain out of the nucleus. This behavior requires a negative regulation of an intrinsic nuclear localization signal (NLS) by the N-terminal region of GtaC and receptor-mediated phosphorylation by the glycogen synthase kinase GskA. When the exit of GtaC from the nucleus is inhibited by adding an exogenous NLS or by mutating the residues involved in phosphorylation, cAMP stimulation, either pulsatile or continuous, induces precocious gene expression and cell differentiation. The two apparently opposing effects of cAMP—it acts as a positive regulator of developmental gene expression but drives GtaC out of the nucleus—suggest that GtaC may be briefly activated during each stimulation cycle before it leaves the nucleus, while its return resensitizes the system. Consistently, we observe that each cAMP cycle generates a transient burst of GtaC-dependent transcription of the contact site A gene (csaA), a well-characterized developmental marker. We further demonstrate that this regulatory mechanism enables cells to modulate gene expression by counting low-frequency stimulations but filtering out high-frequency signals; hence, the steady-state expression level of csaA is determined by the number of effective cAMP pulses experienced by the cell rather than the concentration of the cAMP stimuli or the overall time since the initiation of development. Computer simulations based on a corresponding logical circuit recapitulate GtaC-mediated accumulation of gene products under various stimulation regimes.

Conclusion

This work reveals a decoding mechanism by which oscillatory signals are used to guide gene expression and promote timely development. Tuning transcription to the number rather than the level of the external stimuli allows large populations of cells over an expanded territory to be developmentally synchronized. Similar mechanisms may operate in other circumstances where cellular plasticity is linked to repeated experience.

Oscillate to Synchronize

In the social amoeba Dictyostelium discoideum, periodic waves of the small-molecule cyclic adenosine monophosphate (cAMP) guide chemotactic migration and cell differentiation. Cai et al. (10.1126/science.1249531; see the Perspective by Wollman) found that a GATA family transcription factor shuttles between the nucleus and the cytoplasm of the amoeba in response to oscillatory cAMP signals acting through G protein–coupled receptors. Each oscillation generates a transient burst of gene activation, such that gene expression is linked to the number rather than the level of stimulus, which then brings about transcriptional synchrony in populations of cells.

Abstract

Biological oscillations are observed at many levels of cellular organization. In the social amoeba Dictyostelium discoideum, starvation-triggered multicellular development is organized by periodic cyclic adenosine 3′,5′-monophosphate (cAMP) waves, which provide both chemoattractant gradients and developmental signals. We report that GtaC, a GATA transcription factor, exhibits rapid nucleocytoplasmic shuttling in response to cAMP waves. This behavior requires coordinated action of a nuclear localization signal and reversible G protein (heterotrimeric guanine nucleotide–binding protein)–coupled receptor–mediated phosphorylation. Although both are required for developmental gene expression, receptor occupancy promotes nuclear exit of GtaC, which leads to a transient burst of transcription at each cAMP cycle. We demonstrate that this biological circuit filters out high-frequency signals and counts those admitted, thereby enabling cells to modulate gene expression according to the dynamic pattern of the external stimuli.

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