“cAMP Sponge”: A Buffer for Cyclic Adenosine 3′, 5′-Monophosphate

Background: While intracellular buffers are widely used to study calcium signaling, no such tool exists for the other major second messenger, cyclic AMP (cAMP). Methods/Principal Findings: Here we describe a genetically encoded buffer for cAMP based on the high-affinity cAMP-binding carboxy-terminus of the regulatory subunit $RI\beta$ of protein kinase A (PKA). Addition of targeting sequences permitted localization of this fragment to the extra-nuclear compartment, while tagging with mCherry allowed quantification of its expression at the single cell level. This construct (named “cAMP sponge”) was shown to selectively bind cAMP in vitro. Its expression significantly suppressed agonist-induced cAMP signals and the downstream activation of PKA within the cytosol as measured by FRET-based sensors in single living cells. Point mutations in the cAMP-binding domains of the construct rendered the chimera unable to bind cAMP in vitro or in situ. Cyclic AMP sponge was fruitfully applied to examine feedback regulation of gap junction-mediated transfer of cAMP in epithelial cell couplets. Conclusions: This newest member of the cAMP toolbox has the potential to reveal unique biological functions of cAMP, including insight into the functional significance of compartmentalized signaling events

Exceptionally Preserved Jellyfishes from the Middle Cambrian

Cnidarians represent an early diverging animal group and thus insight into their origin and diversification is key to understanding metazoan evolution. Further, cnidarian jellyfish comprise an important component of modern marine planktonic ecosystems. Here we report on exceptionally preserved cnidarian jellyfish fossils from the Middle Cambrian (∼505 million years old) Marjum Formation of Utah. These are the first described Cambrian jellyfish fossils to display exquisite preservation of soft part anatomy including detailed features of structures interpreted as trailing tentacles and subumbrellar and exumbrellar surfaces. If the interpretation of these preserved characters is correct, their presence is diagnostic of modern jellyfish taxa. These new discoveries may provide insight into the scope of cnidarian diversity shortly after the Cambrian radiation, and would reinforce the notion that important taxonomic components of the modern planktonic realm were in place by the Cambrian period

Calcium-Dependent Increases in Protein Kinase-A Activity in Mouse Retinal Ganglion Cells Are Mediated by Multiple Adenylate Cyclases

Neurons undergo long term, activity dependent changes that are mediated by activation of second messenger cascades. In particular, calcium-dependent activation of the cyclic-AMP/Protein kinase A signaling cascade has been implicated in several developmental processes including cell survival, axonal outgrowth, and axonal refinement. The biochemical link between calcium influx and the activation of the cAMP/PKA pathway is primarily mediated through adenylate cyclases. Here, dual imaging of intracellular calcium concentration and PKA activity was used to assay the role of different classes of calcium-dependent adenylate cyclases (ACs) in the activation of the cAMP/PKA pathway in retinal ganglion cells (RGCs). Surprisingly, depolarization-induced calcium-dependent PKA transients persist in barrelless mice lacking AC1, the predominant calcium-dependent adenylate cyclase in RGCs, as well as in double knockout mice lacking both AC1 and AC8. Furthermore, in a subset of RGCs, depolarization-induced PKA transients persist during the inhibition of all transmembrane adenylate cyclases. These results are consistent with the existence of a soluble adenylate cyclase that plays a role in calcium-dependent activation of the cAMP/PKA cascade in neurons

REVISITING THE PUBLIC STRUCTURE ARTIFACT PATTERN: CULTURAL PATTERNING AT TWO EIGHTEENTH CENTURY GOVERNMENT SITES.

This thesis presents a comparison of the artifact assemblages from two eighteenth century government sites: Delaware's Old State House of 1787 located in Dover , Delaware; and The Chowan County Courthouse of 1767 located in Edenton , North Carolina. The main purpose of this comparison is to test the validity of the Public Structure Artifact Pattern first proposed in 1978 by Cara Wise. The identification of this pattern followed Wise's original analysis of Delaware's Old State House. Wise compared the functional groups of artifacts from the State House with two other sites that operated in a public capacity and found them to display similar frequencies. This study introduces data from excavations of the Chowan County Courthouse , a public site comparable to Delaware's Old State House , as a test implication. The proportions of functional groups of artifacts from the Chowan County site were compared to those displayed by Delaware's Old State House and were assessed for their conformity to the expected Public Structure Pattern. The ceramic assemblages from both sites were also compared at the minimum vessel level both by ware and functional type. This comparison was offered as an independent , complimentary test to further assess similarity in patterning between the sites and to further evaluate the Public Structure Pattern. The overall results of this analysis found general support for the validity of the Public Pattern. The functional groups of artifacts from both sites display remarkable consistency and conform closely to the expected pattern. Additionally , the ceramic assemblages from both sites display consistent proportions of vessels both by ware and functional type

REVISITING THE PUBLIC STRUCTURE ARTIFACT PATTERN: CULTURAL PATTERNING AT TWO EIGHTEENTH CENTURY GOVERNMENT SITES

This thesis presents a comparison of the artifact assemblages from two eighteenth century government sites: Delaware's Old State House of 1787 located in Dover, Delaware\; and The Chowan County Courthouse of 1767 located in Edenton, North Carolina. The main purpose of this comparison is to test the validity of the Public Structure Artifact Pattern first proposed in 1978 by Cara Wise. The identification of this pattern followed Wise's original analysis of Delaware's Old State House. Wise compared the functional groups of artifacts from the State House with two other sites that operated in a public capacity and found them to display similar frequencies. This study introduces data from excavations of the Chowan County Courthouse, a public site comparable to Delaware's Old State House, as a test implication. The proportions of functional groups of artifacts from the Chowan County site were compared to those displayed by Delaware's Old State House and were assessed for their conformity to the expected Public Structure Pattern. The ceramic assemblages from both sites were also compared at the minimum vessel level both by ware and functional type. This comparison was offered as an independent, complimentary test to further assess similarity in patterning between the sites and to further evaluate the Public Structure Pattern. The overall results of this analysis found general support for the validity of the Public Pattern. The functional groups of artifacts from both sites display remarkable consistency and conform closely to the expected pattern. Additionally, the ceramic assemblages from both sites display consistent proportions of vessels both by ware and functional type