Optogenetic control of cAMP oscillations reveals frequency-selective transcription factor dynamics in Dictyostelium.

Oscillatory dynamics and their modulation are crucial for cellular decision-making; however, analysing these dynamics remains challenging. Here, we present a tool that combines the light-activated adenylate cyclase mPAC with the cAMP biosensor Pink Flamindo, enabling precise manipulation and real-time monitoring of cAMP oscillation frequencies in Dictyostelium. High-frequency modulation of cAMP oscillations induced cell aggregation and multicellular formation, even at low cell densities, such as a few dozen cells.

Possible mechanism for improving the endogenous immune system through the blockade of peripheral μ-opioid receptors by treatment with naldemedine.

Background: It has been considered that activation of peripheral μ-opioid receptors (MORs) induces side effects of opioids. In this study, we investigated the possible improvement of the immune system in tumour-bearing mice by systemic administration of the peripheral MOR antagonist naldemedine.

Methods: The inhibitory effect of naldemedine on MOR-mediated signalling was tested by cAMP inhibition and β-arrestin recruitment assays using cultured cells.

Characterization of the discriminative stimulus effect of quinpirole: Further evidence for functional interaction between central dopamine D/D-receptors.

Dysfunction of the central dopamine D-receptor-related network has been proposed to play a critical role in dopamine-related diseases, such as schizophrenia and drug dependence. Generally, the stimulation of dopamine D-receptors on medium spiny neurons (MSN) induces several behavioral effects, such as sedation, hallucination, aversion and motivation. Furthermore, such physiological responses through dopamine D-receptor-containing MSN (D-MSN) may be synchronized with the activity of dopamine D-receptor-containing MSN (D-MSN), or both may exhibit dual agonistic/antagonistic innervation.

CRISPR Toolbox for Genome Editing in .

The development of new techniques to create gene knockouts and knock-ins is essential for successful investigation of gene functions and elucidation of the causes of diseases and their associated fundamental cellular processes. In the biomedical model organism , the methodology for gene targeting with homologous recombination to generate mutants is well-established. Recently, we have applied CRISPR/Cas9-mediated approaches in , allowing the rapid generation of mutants by transiently expressing sgRNA and Cas9 using an all-in-one vector.

Knock-in and precise nucleotide substitution using near-PAMless engineered Cas9 variants in Dictyostelium discoideum.

The powerful genome editing tool Streptococcus pyogenes Cas9 (SpCas9) requires the trinucleotide NGG as a protospacer adjacent motif (PAM). The PAM requirement is limitation for precise genome editing such as single amino-acid substitutions and knock-ins at specific genomic loci since it occurs in narrow editing window. Recently, SpCas9 variants (i.