PCTK3/CDK18 regulates cell migration and adhesion by negatively modulating FAK activity

PCTAIRE kinase 3 (PCTK3) is a member of the cyclin dependent kinase family, but its physiological function remains unknown. We previously reported that PCTK3-knockdown HEK293T cells showed actin accumulation at the leading edge, suggesting that PCTK3 is involved in the regulation of actin reorganization. In this study, we investigated the physiological function and downstream signal transduction molecules of PCTK3. PCTK3 knockdown in HEK293T cells increased cell motility and RhoA/Rho-associated kinase activity as compared with control cells. We also found that phosphorylation at residue Tyr-397 in focal adhesion kinase (FAK) was increased in PCTK3-knockdown cells. FAK phosphorylation at Tyr-397 was increased in response to fibronectin stimulation, whereas its phosphorylation was suppressed by PCTK3. In addition, excessive expression of PCTK3 led to the formation of filopodia during the early stages of cell adhesion in HeLa cells. These results indicate that PCTK3 controls actin cytoskeleton dynamics by negatively regulating the FAK/Rho signaling pathway

Alisol B, a triterpene from Alismatis rhizoma (dried rhizome of Alisma orientale), inhibits melanin production in murine B16 melanoma cells

To develop new whitening agents from natural products, we screened 80 compounds derived from crude drugs in Kampo medicine in a melanin synthesis inhibition assay using murine B16 melanoma cells. The screen revealed that treatment with alisol B, a triterpene from Alismatis rhizoma, significantly decreased both melanin content and cellular tyrosinase activity in B16 cells. However, alisol B did not directly inhibit mushroom tyrosinase activity in vitro. Therefore, we investigated the mechanism underlying the inhibitory effect of alisol B on melanogenesis. Alisol B suppressed mRNA induction of tyrosinase and its transcription factor, microphthalmia-associated transcription factor (MITF). Furthermore, alisol B reduced the phosphorylation of CREB and maintained the activation of ERK1/2. These results suggest that the reduction in melanin production by alisol B is due to the downregulation of MITF through the suppression of CREB and activation of ERK, and that alisol B may be useful as a new whitening agent

Enhanced stability of hippocampal place representation caused by reduced magnesium block of NMDA receptors in the dentate gyrus

BACKGROUND: Voltage-dependent block of the NMDA receptor by Mg(2+) is thought to be central to the unique involvement of this receptor in higher brain functions. However, the in vivo role of the Mg(2+) block in the mammalian brain has not yet been investigated, because brain-wide loss of the Mg(2+) block causes perinatal lethality. In this study, we used a brain-region specific knock-in mouse expressing an NMDA receptor that is defective for the Mg(2+) block in order to test its role in neural information processing. RESULTS: We devised a method to induce a single amino acid substitution (N595Q) in the GluN2A subunit of the NMDA receptor, specifically in the hippocampal dentate gyrus in mice. This mutation reduced the Mg(2+) block at the medial perforant path–granule cell synapse and facilitated synaptic potentiation induced by high-frequency stimulation. The mutants had more stable hippocampal place fields in the CA1 than the controls did, and place representation showed lower sensitivity to visual differences. In addition, behavioral tests revealed that the mutants had a spatial working memory deficit. CONCLUSIONS: These results suggest that the Mg(2+) block in the dentate gyrus regulates hippocampal spatial information processing by attenuating activity-dependent synaptic potentiation in the dentate gyrus

SLPI is a critical mediator that controls PTH-induced bone formation

Osteoclastic bone resorption and osteoblastic bone formation/replenishment are closely coupled in bone metabolism. Anabolic parathyroid hormone (PTH), which is commonly used for treating osteoporosis, shifts the balance from osteoclastic to osteoblastic, although it is unclear how these cells are coordinately regulated by PTH. Here, we identify a serine protease inhibitor, secretory leukocyte protease inhibitor (SLPI), as a critical mediator that is involved in the PTH-mediated shift to the osteoblastic phase. Slpi is highly upregulated in osteoblasts by PTH, while genetic ablation of Slpi severely impairs PTH-induced bone formation. Slpi induction in osteoblasts enhances its differentiation, and increases osteoblast–osteoclast contact, thereby suppressing osteoclastic function. Intravital bone imaging reveals that the PTH-mediated association between osteoblasts and osteoclasts is disrupted in the absence of SLPI. Collectively, these results demonstrate that SLPI regulates the communication between osteoblasts and osteoclasts to promote PTH-induced bone anabolism.Morimoto A., Kikuta J., Nishikawa K., et al. SLPI is a critical mediator that controls PTH-induced bone formation. Nature Communications 12, 2136 (2021); https://doi.org/10.1038/s41467-021-22402-x

Osteoclasts adapt to physioxia perturbation through DNA demethylation

Oxygen plays an important role in diverse biological processes. However, since quantitation of the partial pressure of cellular oxygen in vivo is challenging, the extent of oxygen perturbation in situ and its cellular response remains underexplored. Using two-photon phosphorescence lifetime imaging microscopy, we determine the physiological range of oxygen tension in osteoclasts of live mice. We find that oxygen tension ranges from 17.4 to 36.4 mmHg, under hypoxic and normoxic conditions, respectively. Physiological normoxia thus corresponds to 5% and hypoxia to 2% oxygen in osteoclasts. Hypoxia in this range severely limits osteoclastogenesis, independent of energy metabolism and hypoxia-inducible factor activity. We observe that hypoxia decreases ten-eleven translocation (TET) activity. Tet2/3 cooperatively induces Prdm1 expression via oxygen-dependent DNA demethylation, which in turn activates NFATc1 required for osteoclastogenesis. Taken together, our results reveal that TET enzymes, acting as functional oxygen sensors, regulate osteoclastogenesis within the physiological range of oxygen tension, thus opening new avenues for research on in vivo response to oxygen perturbation.Nishikawa K., Seno S., Yoshihara T., et al. Osteoclasts adapt to physioxia perturbation through DNA demethylation. EMBO Reports 22, e53035 (2021); https://doi.org/10.15252/embr.202153035