Journal: bioRxiv

Article Title: A novel sodium signaling complex regulates uterine activity

doi: 10.1101/2020.07.31.229138

Figure Lengend Snippet: During the quiescent state, progesterone binding to the progesterone receptor (PR A/B ) increases NALCN expression and activity . Sodium current through NALCN activates SLO2.1 channels, increasing K + efflux to maintain the cell in a hyperpolarized state. As a result, voltage-dependent Ca 2+ channels (VDCCs) are closed, and uterine contractions do not occur. In the contractile state, estrogen acting on ERα inhibits NALCN expression , leading to decreased SLO2.1 activity. The reduced K + efflux depolarizes the membrane, leading to VDCC activation, an increase in intracellular Ca 2+ , and uterine contractility. At labor, Oxytocin (OXT) binds to the oxytocin receptor (OTR), leading to activation of phospholipase C (PLC), production of phosphatidylinositol 4,5-bisphosphate (PIP 2 ), and production of inositol triphosphate (IP 3 ). IP 3 activates the release of Ca 2+ from intracellular stores, and PIP 2 activates protein kinase C (PKC), which inhibits SLO2.1 . This SLO2.1 inhibition further depolarizes the membrane, thus opening more VDCCs, increasing intracellular Ca 2+ , and further activating myosin to cause muscle contraction.

Article Snippet: Duolink in situ proximity ligation assay (Sigma, St. Louis, MO) labeling was performed with the following antibodies: NALCN (mouse monoclonal, 1:100, StressMarq) and SLO2.1 (rabbit polyclonal, 1:200, Alomone).

Techniques: Binding Assay, Expressing, Activity Assay, Activation Assay, Inhibition

Journal:

Article Title: Phospholipase D2 Is Localized to the Rims of the Golgi Apparatus in Mammalian Cells

doi: 10.1091/mbc.02-04-0059

Figure Lengend Snippet: PLD2 localization with different organelles in rat NRK cells. Cells were prepared for immunofluorescence microscopy and incubated with rabbit PLD2 antibody PLD2-27 (A, D, G, and J) and costained with monoclonal antibodies to the ER marker BiP (B); the transferrin receptor (E), a marker of early endosomes and the plasma membrane; and lgp120 (H), a late endosome/lysosomal marker. (K) Cells were stained with caveolin-1, a marker for caveoli; the arrow corresponds to caveolin-1 localized to the plasma membrane. Images were merged to determine overlap between PLD2 (red) and the respective marker proteins (green; C, F, I, and L). Areas of maximal overlap are yellow. (L) Inset, enlargement of the region of overlap between PLD2 and caveolin-1 in the perinuclear Golgi apparatus. The arrow indicates areas where PLD2 puncta (red) either overlap completely with caveolin-1 (green) or are directly adjacent. Images are from projected Z-series. Bars, 10 μm.

Article Snippet: Purified monoclonal anti-rat transferrin receptor antibodies were purchased from Cedarlane Laboratories (Hornby, Ontario, Canada).

Techniques: Immunofluorescence, Microscopy, Incubation, Marker, Staining